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Sample records for interspecies nuclear transfer

  1. Cloning Endangered Felids by Interspecies Somatic Cell Nuclear Transfer.

    PubMed

    Gómez, Martha C; Pope, C Earle

    2015-01-01

    In 2003, the first wild felid was produced by interspecies somatic cell nuclear transfer. Since then other wild felid clone offspring have been produced by using the same technique with minor modifications. This chapter describes detailed protocols used in our laboratory for (1) the isolation, culture, and preparation of fibroblast cells as donor nucleus, and (2) embryo reconstruction with domestic cat enucleated oocytes to produce cloned embryos that develop to the blastocyst stage in vitro and, after transfer into synchronized recipients, establish successful pregnancies.

  2. Interspecies Somatic Cell Nuclear Transfer: Advancements and Problems

    PubMed Central

    Lagutina, Irina; Fulka, Helena; Lazzari, Giovanna

    2013-01-01

    Abstract Embryologists working with livestock species were the pioneers in the field of reprogramming by somatic cell nuclear transfer (SCNT). Without the “Dolly experiment,” the field of cellular reprogramming would have been slow and induced plutipotent cells (iPSCs) would not have been conceived. The major drive of the work in mammalian cloning was the interest of the breeding industry to propagate superior genotypes. Soon it was realized that the properties of oocytes could be used also to clone endangered mammalian species or to reprogram the genomes of unrelated species through what is known as interspecies (i) SCNT, using easily available oocytes of livestock species. iSCNT for cloning animals works only for species that can interbreed, and experiments with taxonomically distant species have not been successful in obtaining live births or deriving embryonic stem cell (ESC) lines to be used for regenerative medicine. There are controversial reports in the literature, but in most cases these experiments have underlined some of the cellular and molecular mechanisms that are incomplete during cell nucleus reprogramming, including the failure to organize nucleoli, silence somatic cell genes, activate the embryonic genome, and resume mitochondrial replication and function, thus indicating nucleus–cytoplasmic incompatibility. PMID:24033141

  3. Two-staged nuclear transfer can enhance the developmental ability of goat-sheep interspecies nuclear transfer embryos in vitro.

    PubMed

    Ma, Li-Bing; Cai, Lu; Li, Jia-Jia; Chen, Xiu-Li; Ji, Feng-Yu

    2011-02-01

    The technique of interspecies somatic cell nuclear transfer, in which interspecies cloned embryos can be reconstructed by using domestic animal oocytes as nuclear recipients and endangered animal or human somatic cells as nuclear donors, can afford more opportunities in endangered animal rescue and human tissue transplantation, but the application of this technique is limited by extremely low efficiency which may be attributed to donor nucleus not fully reprogrammed by xenogenic cytoplasm. In this study, goat fetal fibroblasts (GFFs) were used as nuclear donors, in vitro-matured sheep oocytes were used as nuclear recipients, and a two-stage nuclear transfer procedure was performed to improve the developmental ability of goat-sheep interspecies clone embryos. In the first stage nuclear transfer (FSNT), GFFs were injected into the ooplasm of enucleated sheep metaphase-II oocytes, then non-activated reconstructed embryos were cultured in vitro, so that the donor nucleus could be exposed to the ooplasm for a period of time. Subsequently, in the second stage nuclear transfer, FSNT-derived non-activated reconstructed embryo was centrifuged, and the donor nucleus was then transferred into another freshly enucleated sheep oocyte. Compared with the one-stage nuclear transfer, two-stage nuclear transfer could significantly enhance the blastocyst rate of goat-sheep interspecies clone embryos, and this result indicated that longtime exposure to xenogenic ooplasm benefits the donor nucleus to be reprogrammed. The two-stage nuclear transfer procedure has two advantages, one is that the donor nucleus can be exposed to the ooplasm for a long time, the other is that the problem of oocyte aging can be solved. PMID:21082282

  4. In vitro development of bison embryos using interspecies somatic cell nuclear transfer.

    PubMed

    Seaby, R P; Alexander, B; King, W A; Mastromonaco, G F

    2013-12-01

    Interspecies somatic cell nuclear transfer (interspecies SCNT) has been explored in many domestic and non-domestic animal species. However, problems arise during the development of these embryos, which may be related to species-specific differences in nuclear-cytoplasmic communication. The objectives of this study were to investigate the possibility of producing bison embryos in vitro using interspecies SCNT and assess the developmental potential of these embryos. Treatment groups consisted of cattle in vitro fertilization (IVF) and cattle SCNT as controls and wood bison SCNT, plains bison SCNT and wisent SCNT as experimental groups. Cleavage and blastocyst rates were assessed, and blastocyst quality was determined using total cell number, apoptotic incidence and relative quantification of mitochondria-related genes NRF1, MT-CYB and TFAM. These results indicate that embryos can be produced by interspecies SCNT in all bison species/subspecies (13.34-33.54% blastocyst rates). Although increased incidence of apoptosis was observed in bison SCNT blastocysts compared to cattle SCNT controls (10.45-12.69 vs 8.76, respectively) that corresponded with significantly lower cell numbers (80-87 cells vs >100 cells, respectively), no major differences were observed in the expression of NRF1, MT-CYB and TFAM. This study is the first to report the production of bison embryos by interspecies SCNT. Blastocyst development in all three bison species/subspecies was greater than the rates obtained in previous studies by IVF, which supports the potential role of SCNT for in vitro embryo production in this species. Yet, further investigation of developmental competence and the factors influencing blastocyst quality and viability is required.

  5. Successful cloning of coyotes through interspecies somatic cell nuclear transfer using domestic dog oocytes.

    PubMed

    Hwang, Insung; Jeong, Yeon Woo; Kim, Joung Joo; Lee, Hyo Jeong; Kang, Mina; Park, Kang Bae; Park, Jung Hwan; Kim, Yeun Wook; Kim, Woo Tae; Shin, Taeyoung; Hyun, Sang Hwan; Jeung, Eui-Bae; Hwang, Woo Suk

    2013-01-01

    Interspecies somatic cell nuclear transfer (iSCNT) is an emerging assisted reproductive technology (ART) for preserving Nature's diversity. The scarcity of oocytes from some species makes utilisation of readily available oocytes inevitable. In the present study, we describe the successful cloning of coyotes (Canis latrans) through iSCNT using oocytes from domestic dogs (Canis lupus familiaris or dingo). Transfer of 320 interspecies-reconstructed embryos into 22 domestic dog recipients resulted in six pregnancies, from which eight viable offspring were delivered. Fusion rate and cloning efficiency during iSCNT cloning of coyotes were not significantly different from those observed during intraspecies cloning of domestic dogs. Using neonatal fibroblasts as donor cells significantly improved the cloning efficiency compared with cloning using adult fibroblast donor cells (P<0.05). The use of domestic dog oocytes in the cloning of coyotes in the present study holds promise for cloning other endangered species in the Canidae family using similar techniques. However, there are still limitations of the iSCNT technology, as demonstrated by births of morphologically abnormal coyotes and the clones' inheritance of maternal domestic dog mitochondrial DNA.

  6. Successful cloning of coyotes through interspecies somatic cell nuclear transfer using domestic dog oocytes.

    PubMed

    Hwang, Insung; Jeong, Yeon Woo; Kim, Joung Joo; Lee, Hyo Jeong; Kang, Mina; Park, Kang Bae; Park, Jung Hwan; Kim, Yeun Wook; Kim, Woo Tae; Shin, Taeyoung; Hyun, Sang Hwan; Jeung, Eui-Bae; Hwang, Woo Suk

    2013-01-01

    Interspecies somatic cell nuclear transfer (iSCNT) is an emerging assisted reproductive technology (ART) for preserving Nature's diversity. The scarcity of oocytes from some species makes utilisation of readily available oocytes inevitable. In the present study, we describe the successful cloning of coyotes (Canis latrans) through iSCNT using oocytes from domestic dogs (Canis lupus familiaris or dingo). Transfer of 320 interspecies-reconstructed embryos into 22 domestic dog recipients resulted in six pregnancies, from which eight viable offspring were delivered. Fusion rate and cloning efficiency during iSCNT cloning of coyotes were not significantly different from those observed during intraspecies cloning of domestic dogs. Using neonatal fibroblasts as donor cells significantly improved the cloning efficiency compared with cloning using adult fibroblast donor cells (P<0.05). The use of domestic dog oocytes in the cloning of coyotes in the present study holds promise for cloning other endangered species in the Canidae family using similar techniques. However, there are still limitations of the iSCNT technology, as demonstrated by births of morphologically abnormal coyotes and the clones' inheritance of maternal domestic dog mitochondrial DNA. PMID:23217630

  7. Function of donor cell centrosome in intraspecies and interspecies nuclear transfer embryos

    SciTech Connect

    Zhong Zhisheng; Zhang Gang; Meng Xiaoqian; Zhang Yanling; Chen Dayuan; Schatten, Heide; Sun Qingyuan . E-mail: sunqy1@yahoo.com

    2005-05-15

    Centrosomes, the main microtubule-organizing centers (MTOCs) in most animal cells, are important for many cellular activities such as assembly of the mitotic spindle, establishment of cell polarity, and cell movement. In nuclear transfer (NT), MTOCs that are located at the poles of the meiotic spindle are removed from the recipient oocyte, while the centrosome of the donor cell is introduced. We used mouse MII oocytes as recipients, mouse fibroblasts, rat fibroblasts, or pig granulosa cells as donor cells to construct intraspecies and interspecies nuclear transfer embryos in order to observe centrosome dynamics and functions. Three antibodies against centrin, {gamma}-tubulin, and NuMA, respectively, were used to stain the centrosome. Centrin was not detected either at the poles of transient spindles or at the poles of first mitotic spindles. {gamma}-tubulin translocated into the two poles of the transient spindles, while no accumulated {gamma}-tubulin aggregates were detected in the area adjacent to the two pseudo-pronuclei. At first mitotic metaphase, {gamma}-tubulin was translocated to the spindle poles. The distribution of {gamma}-tubulin was similar in mouse intraspecies and rat-mouse interspecies embryos. The NuMA antibody that we used can recognize porcine but not murine NuMA protein, so it was used to trace the NuMA protein of donor cell in reconstructed embryos. In the pig-mouse interspecies reconstructed embryos, NuMA concentrated between the disarrayed chromosomes soon after activation and translocated to the transient spindle poles. NuMA then immigrated into pseudo-pronuclei. After pseudo-pronuclear envelope breakdown, NuMA was located between the chromosomes and then translocated to the spindle poles of first mitotic metaphase. {gamma}-tubulin antibody microinjection resulted in spindle disorganization and retardation of the first cell division. NuMA antibody microinjection also resulted in spindle disorganization. Our findings indicate that (1) the

  8. Reprogrammed Transcriptome in Rhesus-Bovine Interspecies Somatic Cell Nuclear Transfer Embryos

    PubMed Central

    Wang, Kai; Otu, Hasan H.; Chen, Ying; Lee, Young; Latham, Keith; Cibelli, Jose B.

    2011-01-01

    Background Global activation of the embryonic genome (EGA), one of the most critical steps in early mammalian embryo development, is recognized as the time when interspecies somatic cell nuclear transfer (iSCNT) embryos fail to thrive. Methodology/Principal Findings In this study, we analyzed the EGA-related transcriptome of rhesus-bovine iSCNT 8- to 16-cell embryos and dissected the reprogramming process in terms of embryonic gene activation, somatic gene silencing, and maternal RNA degradation. Compared with fibroblast donor cells, two thousand and seven genes were activated in iSCNT embryos, one quarter of them reaching expression levels comparable to those found in in vitro fertilized (IVF) rhesus embryos. This suggested that EGA in iSCNT embryos had partially recapitulated rhesus embryonic development. Eight hundred and sixty somatic genes were not silenced properly and continued to be expressed in iSCNT embryos, which indicated incomplete nuclear reprogramming. We compared maternal RNA degradation in bovine oocytes between bovine-bovine SCNT and iSCNT embryos. While maternal RNA degradation occurred in both SCNT and iSCNT embryos, we saw more limited overall degradation of maternal RNA in iSCNT embryos than in SCNT embryos. Several important maternal RNAs, like GPF9, were not properly processed in SCNT embryos. Conclusions/Significance Our data suggested that iSCNT embryos are capable of triggering EGA, while a portion of somatic cell-associated genes maintain their expression. Maternal RNA degradation seems to be impaired in iSCNT embryos. Further understanding of the biological roles of these genes, networks, and pathways revealed by iSCNT may expand our knowledge about cell reprogramming, pluripotency, and differentiation. PMID:21799794

  9. Production of wild buffalo (Bubalus arnee) embryos by interspecies somatic cell nuclear transfer using domestic buffalo (Bubalus bubalis) oocytes.

    PubMed

    Priya, D; Selokar, N L; Raja, A K; Saini, M; Sahare, A A; Nala, N; Palta, P; Chauhan, M S; Manik, R S; Singla, S K

    2014-04-01

    The objective of this study was to explore the possibility of producing wild buffalo embryos by interspecies somatic cell nuclear transfer (iSCNT) through handmade cloning using wild buffalo somatic cells and domestic buffalo (Bubalus bubalis) oocytes. Somatic cells derived from the ear skin of wild buffalo were found to express vimentin but not keratin and cytokeratin-18, indicating that they were of fibroblast origin. The population doubling time of skin fibroblasts from wild buffalo was significantly (p < 0.05) higher, and the cell proliferation rate was significantly (p < 0.05) lower compared with that of skin fibroblasts from domestic buffalo. Neither the cleavage (92.6 ± 2.0% vs 92.8 ± 2.0%) nor the blastocyst rate (42.4 ± 2.4% vs 38.7 ± 2.8%) was significantly different between the intraspecies cloned embryos produced using skin fibroblasts from domestic buffalo and interspecies cloned embryos produced using skin fibroblasts from wild buffalo. However, the total cell number (TCN) was significantly (p < 0.05) lower (192.0 ± 25.6 vs 345.7 ± 42.2), and the apoptotic index was significantly (p < 0.05) higher (15.1 ± 3.1 vs 8.0 ± 1.4) for interspecies than that for intraspecies cloned embryos. Following vitrification in open-pulled straws (OPS) and warming, although the cryosurvival rate of both types of cloned embryos, as indicated by their re-expansion rate, was not significantly different (34.8 ± 1.5% vs 47.8 ± 7.8), the apoptotic index was significantly (p < 0.05) higher for vitrified-warmed interspecies than that for corresponding intraspecies cloned embryos (48.9 ± 7.2 vs 23.9 ± 2.8). The global level of H3K18ac was significantly (p < 0.05) lower in interspecies cloned embryos than that in intraspecies cloned embryos. The expression level of HDAC1, DNMT3a and CASPASE3 was significantly (p < 0.05) higher, that of P53 was significantly (p < 0.05) lower in interspecies than in intraspecies embryos, whereas that of DNMT1 was similar between the two

  10. Production of interspecies handmade cloned embryos by nuclear transfer of cattle, goat and rat fibroblasts to buffalo (Bubalus bubalis) oocytes.

    PubMed

    Selokar, N L; George, A; Saha, A P; Sharma, R; Muzaffer, M; Shah, R A; Palta, P; Chauhan, M S; Manik, R S; Singla, S K

    2011-02-01

    The possibility of producing interspecies handmade cloned (iHMC) embryos by nuclear transfer from donor cells of cattle, goat and rat using buffalo oocytes as recipient cytoplasts was explored. Zona-free buffalo oocytes were enucleated by protrusion cone-guided bisection with a microblade. After electrofusion with somatic cells, reconstructed oocytes were activated by calcimycin A23187, treated with 6-dimethylaminopurine and were cultured in K-RVCL-50® medium for 8 days. Although the cleavage rate was not significantly different when buffalo, cattle, goat or rat cells were used as donor nuclei (74.6 ± 3.8, 82.8 ± 5.3, 86.0 ± 4.9 and 82.3 ± 3.6%, respectively), the blastocyst rate was significantly higher (P<0.01) for buffalo (51.4 ± 2.6) than for cattle (3.5 ± 1.0) or the goat (2.2 ± 0.9), whereas none of the embryos crossed the 32-cell stage when rat cells were used. However, the total cell number was similar for buffalo-buffalo (175.0 ± 5.07) and cattle-buffalo embryos (178.0 ± 11.84). Following transfer of 3 buffalo-buffalo embryos each to 6 recipients, 3 were found to be pregnant, though the pregnancies were not carried to full term. These results suggest that interspecies blastocyst stage embryos can be produced by iHMC using buffalo cytoplasts and differentiated somatic cells from cattle and goat and that the source of donor nucleus affects the developmental competence of interspecies embryos.

  11. Production of rhesus monkey cloned embryos expressing monomeric red fluorescent protein by interspecies somatic cell nuclear transfer

    SciTech Connect

    Zhu, Hai-Ying; Kang, Jin-Dan; Li, Suo; Jin, Jun-Xue; Hong, Yu; Jin, Long; Guo, Qing; Gao, Qing-Shan; Yan, Chang-Guo; Yin, Xi-Jun

    2014-02-21

    Highlights: • Rhesus monkey cells were electroporated with a plasmid containing mRFP1, and an mRFP1-expressing cell line was generated. • For the first time, mRFP1-expressing rhesus monkey cells were used as donor cells for iSCNT. • The effect of VPA on the development of embryos cloned using iSCNT was determined. - Abstract: Interspecies somatic cell nuclear transfer (iSCNT) is a promising method to clone endangered animals from which oocytes are difficult to obtain. Monomeric red fluorescent protein 1 (mRFP1) is an excellent selection marker for transgenically modified cloned embryos during somatic cell nuclear transfer (SCNT). In this study, mRFP-expressing rhesus monkey cells or porcine cells were transferred into enucleated porcine oocytes to generate iSCNT and SCNT embryos, respectively. The development of these embryos was studied in vitro. The percentage of embryos that underwent cleavage did not significantly differ between iSCNT and SCNT embryos (P > 0.05; 71.53% vs. 80.30%). However, significantly fewer iSCNT embryos than SCNT embryos reached the blastocyst stage (2.04% vs. 10.19%, P < 0.05). Valproic acid was used in an attempt to increase the percentage of iSCNT embryos that developed to the blastocyst stage. However, the percentages of embryos that underwent cleavage and reached the blastocyst stage were similar between untreated iSCNT embryos and iSCNT embryos treated with 2 mM valproic acid for 24 h (72.12% vs. 70.83% and 2.67% vs. 2.35%, respectively). These data suggest that porcine-rhesus monkey interspecies embryos can be generated that efficiently express mRFP1. However, a significantly lower proportion of iSCNT embryos than SCNT embryos reach the blastocyst stage. Valproic acid does not increase the percentage of porcine-rhesus monkey iSCNT embryos that reach the blastocyst stage. The mechanisms underling nuclear reprogramming and epigenetic modifications in iSCNT need to be investigated further.

  12. Absence of nucleolus formation in raccoon dog-porcine interspecies somatic cell nuclear transfer embryos results in embryonic developmental failure.

    PubMed

    Jeon, Yubyeol; Nam, Yeong-Hee; Cheong, Seung-A; Kwak, Seong-Sung; Lee, Eunsong; Hyun, Sang-Hwan

    2016-08-25

    Interspecies somatic cell nuclear transfer (iSCNT) can be a solution for preservation of endangered species that have limited oocytes. It has been reported that blastocyst production by iSCNT is successful even if the genetic distances between donors and recipients are large. In particular, domestic pig oocytes can support the development of canine to porcine iSCNT embryos. Therefore, we examined whether porcine oocytes may be suitable recipient oocytes for Korean raccoon dog iSCNT. We investigated the effects of trichostatin A (TSA) treatment on iSCNT embryo developmental patterns and nucleolus formation. Enucleated porcine oocytes were fused with raccoon dog fibroblasts by electrofusion and cleavage, and blastocyst development and nucleolus formation were evaluated. To our knowledge, this study is the first in which raccoon dog iSCNT was performed using porcine oocytes; we found that 68.5% of 158 iSCNT embryos had the ability to cleave. However, these iSCNT embryos did not develop past the 4-cell stage. Treatment with TSA did not affect iSCNT embryonic development; moreover, the nuclei failed to form nucleoli at 48 and 72 h post-activation (hpa). In contrast, pig SCNT embryos of the control group showed 18.8% and 87.9% nucleolus formation at 48 and 72 hpa, respectively. Our results demonstrated that porcine cytoplasts efficiently supported the development of raccoon dog iSCNT embryos to the 4-cell stage, the stage of porcine embryonic genome activation (EGA); however, these embryos failed to reach the blastocyst stage and showed defects in nucleolus formation. PMID:27064112

  13. Absence of nucleolus formation in raccoon dog-porcine interspecies somatic cell nuclear transfer embryos results in embryonic developmental failure.

    PubMed

    Jeon, Yubyeol; Nam, Yeong-Hee; Cheong, Seung-A; Kwak, Seong-Sung; Lee, Eunsong; Hyun, Sang-Hwan

    2016-08-25

    Interspecies somatic cell nuclear transfer (iSCNT) can be a solution for preservation of endangered species that have limited oocytes. It has been reported that blastocyst production by iSCNT is successful even if the genetic distances between donors and recipients are large. In particular, domestic pig oocytes can support the development of canine to porcine iSCNT embryos. Therefore, we examined whether porcine oocytes may be suitable recipient oocytes for Korean raccoon dog iSCNT. We investigated the effects of trichostatin A (TSA) treatment on iSCNT embryo developmental patterns and nucleolus formation. Enucleated porcine oocytes were fused with raccoon dog fibroblasts by electrofusion and cleavage, and blastocyst development and nucleolus formation were evaluated. To our knowledge, this study is the first in which raccoon dog iSCNT was performed using porcine oocytes; we found that 68.5% of 158 iSCNT embryos had the ability to cleave. However, these iSCNT embryos did not develop past the 4-cell stage. Treatment with TSA did not affect iSCNT embryonic development; moreover, the nuclei failed to form nucleoli at 48 and 72 h post-activation (hpa). In contrast, pig SCNT embryos of the control group showed 18.8% and 87.9% nucleolus formation at 48 and 72 hpa, respectively. Our results demonstrated that porcine cytoplasts efficiently supported the development of raccoon dog iSCNT embryos to the 4-cell stage, the stage of porcine embryonic genome activation (EGA); however, these embryos failed to reach the blastocyst stage and showed defects in nucleolus formation.

  14. Absence of nucleolus formation in raccoon dog-porcine interspecies somatic cell nuclear transfer embryos results in embryonic developmental failure

    PubMed Central

    JEON, Yubyeol; NAM, Yeong-Hee; CHEONG, Seung-A; KWAK, Seong-Sung; LEE, Eunsong; HYUN, Sang-Hwan

    2016-01-01

    Interspecies somatic cell nuclear transfer (iSCNT) can be a solution for preservation of endangered species that have limited oocytes. It has been reported that blastocyst production by iSCNT is successful even if the genetic distances between donors and recipients are large. In particular, domestic pig oocytes can support the development of canine to porcine iSCNT embryos. Therefore, we examined whether porcine oocytes may be suitable recipient oocytes for Korean raccoon dog iSCNT. We investigated the effects of trichostatin A (TSA) treatment on iSCNT embryo developmental patterns and nucleolus formation. Enucleated porcine oocytes were fused with raccoon dog fibroblasts by electrofusion and cleavage, and blastocyst development and nucleolus formation were evaluated. To our knowledge, this study is the first in which raccoon dog iSCNT was performed using porcine oocytes; we found that 68.5% of 158 iSCNT embryos had the ability to cleave. However, these iSCNT embryos did not develop past the 4-cell stage. Treatment with TSA did not affect iSCNT embryonic development; moreover, the nuclei failed to form nucleoli at 48 and 72 h post-activation (hpa). In contrast, pig SCNT embryos of the control group showed 18.8% and 87.9% nucleolus formation at 48 and 72 hpa, respectively. Our results demonstrated that porcine cytoplasts efficiently supported the development of raccoon dog iSCNT embryos to the 4-cell stage, the stage of porcine embryonic genome activation (EGA); however, these embryos failed to reach the blastocyst stage and showed defects in nucleolus formation. PMID:27064112

  15. Interspecies nuclear transfer using fibroblasts from leopard, tiger, and lion ear piece collected postmortem as donor cells and rabbit oocytes as recipients.

    PubMed

    Yelisetti, Uma Mahesh; Komjeti, Suman; Katari, Venu Charan; Sisinthy, Shivaji; Brahmasani, Sambasiva Rao

    2016-06-01

    Skin fibroblast cells were obtained from a small piece of an ear of leopard, lion, and tiger collected postmortem and attempts were made to synchronize the skin fibroblasts at G0/G1 of cell cycle using three different approaches. Efficiency of the approaches was tested following interspecies nuclear transfer with rabbit oocytes as recipient cytoplasm. Fluorescence-activated cell sorting revealed that the proportion of G0/G1 cells increased significantly (P < 0.05) when cells subjected to serum starvation, contact inhibition, and 3 mM sodium butyrate (NaBu) treatment when compared with cycling cells. However, 3 mM NaBu treatment caused alterations in cell morphology and increase in dead cells. Thus, interspecies nuclear transfer was carried out using fibroblast cells subjected to contact inhibition for 72 h, serum starvation for 48 h, and cells treated with 1.0 mM NaBu for 48 h. The fusion rates, the proportion of fused couplets that cleaved to two-cell and developed to blastocyst, were highest in all three species when the donor cells were treated with 1.0 mM NaBu for 48 h. But, the blastocyst percentage of interspecies nuclear embryos (5-6%) was significantly lower when compared with rabbit-rabbit nuclear transfer embryos (22.9%). In conclusion, fibroblast cells of leopard, lion, and tiger were successfully synchronized and used for the development of blastocysts using rabbit oocytes as recipient cytoplasm. PMID:27071624

  16. Embryo aggregation does not improve the development of interspecies somatic cell nuclear transfer embryos in the horse.

    PubMed

    Gambini, Andrés; De Stéfano, Adrián; Jarazo, Javier; Buemo, Carla; Karlanian, Florencia; Salamone, Daniel Felipe

    2016-09-01

    The low efficiency of interspecies somatic cell nuclear transfer (iSCNT) makes it necessary to investigate new strategies to improve embryonic developmental competence. Embryo aggregation has been successfully applied to improve cloning efficiency in mammals, but it remains unclear whether it could also be beneficial for iSCNT. In this study, we first compared the effect of embryo aggregation over in vitro development and blastocyst quality of porcine, bovine, and feline zona-free (ZF) parthenogenetic (PA) embryos to test the effects of embryo aggregation on species that were later used as enucleated oocytes donors in our iSCNT study. We then assessed whether embryo aggregation could improve the in vitro development of ZF equine iSCNT embryos after reconstruction with porcine, bovine, and feline ooplasm. Bovine- and porcine-aggregated PA blastocysts had significantly larger diameters compared with nonaggregated embryos. On the other hand, feline- and bovine-aggregated PA embryos had higher blastocyst cell number. Embryo aggregation of equine-equine SCNT was found to be beneficial for embryo development as we have previously reported, but the aggregation of three ZF reconstructed embryos did not improve embryo developmental rates on iSCNT. In vitro embryo development of nonaggregated iSCNT was predominantly arrested around the stage when transcriptional activation of the embryonic genome is reported to start on the embryo of the donor species. Nevertheless, independent of embryo aggregation, equine blastocyst-like structures could be obtained in our study using domestic feline-enucleated oocytes. Taken together, these results reported that embryo aggregation enhance in vitro PA embryo development and embryo quality but effects vary depending on the species. Embryo aggregation also improves, as expected, the in vitro embryo development of equine-equine SCNT embryos; however, we did not observe positive effects on equine iSCNT embryo development. Among oocytes

  17. Promoting interspecies electron transfer with biochar.

    PubMed

    Chen, Shanshan; Rotaru, Amelia-Elena; Shrestha, Pravin Malla; Malvankar, Nikhil S; Liu, Fanghua; Fan, Wei; Nevin, Kelly P; Lovley, Derek R

    2014-01-01

    Biochar, a charcoal-like product of the incomplete combustion of organic materials, is an increasingly popular soil amendment designed to improve soil fertility. We investigated the possibility that biochar could promote direct interspecies electron transfer (DIET) in a manner similar to that previously reported for granular activated carbon (GAC). Although the biochars investigated were 1000 times less conductive than GAC, they stimulated DIET in co-cultures of Geobacter metallireducens with Geobacter sulfurreducens or Methanosarcina barkeri in which ethanol was the electron donor. Cells were attached to the biochar, yet not in close contact, suggesting that electrons were likely conducted through the biochar, rather than biological electrical connections. The finding that biochar can stimulate DIET may be an important consideration when amending soils with biochar and can help explain why biochar may enhance methane production from organic wastes under anaerobic conditions.

  18. Low oxygen tension and relative defined culture medium with 3, 4-dihydroxyflavone are beneficial for yak-bovine interspecies somatic cell nuclear transfer embryo.

    PubMed

    Xiong, X; Li, J; Wang, L; Zhong, J; Zi, X; Wang, Y

    2014-02-01

    With an aim to improve the efficiency of yak-bovine interspecies somatic cell nuclear transfer (iSCNT), this study investigated the effect of different culture systems on the development, quality and gene expression profile of yak-bovine iSCNT embryo. Reconstructed embryos were cultured in modified synthetic oviductal fluid (mSOF) or relative defined culture medium (RDCM) with 5% or 20% oxygen tension. Relative mRNA abundance of Oct-4, IFNT, IGF-2, Bax, GPX-1, SOD-1, CAT and GSS was analysed in blastocysts with qRT-PCR. The blastocyst formation rate in RDCM under 5% oxygen tension was significantly higher than that under 20% oxygen tension (P < 0.05). The total cell number of blastocyst derived from RDCM with 20% oxygen tension was lower than that of other groups, whereas the group of RDCM with 5% oxygen tension showed a beneficial effect on apoptosis index and tolerance to cryopreservation (P < 0.05). However, under the same oxygen tension, the mRNA abundance of IFNT of RDCM groups was higher than that of the mSOF groups. In addition, high oxygen tension during in vitro culture (IVC) with RDCM significantly increases the mRNA expression of oxidative stress-related genes (GPX-1, SOD-1, CAT and GSS) (P < 0.05). 3, 4-Dihydroxyflavone (DHF) during high oxygen tension was able to improve the cloned blastocyst formation rate in RDCM (P < 0.05). These results for the first time showed that low oxygen tension and RDCM could improve the developmental competence and quality and alleviate the oxidative stress for yak-bovine iSCNT embryo during IVC.

  19. Cloned endangered species takin (Budorcas taxicolor) by inter-species nuclear transfer and comparison of the blastocyst development with yak (Bos grunniens) and bovine.

    PubMed

    Li, Yanxin; Dai, Yunping; Du, Weihua; Zhao, Chunjiang; Wang, Haiping; Wang, Lili; Li, Rong; Liu, Ying; Wan, Rong; Li, Ning

    2006-02-01

    Interspecies cloning might be used as an effective method to conserve endangered species and to support the study of nuclear-cytoplasm interaction. In this study, we describe the development of takin-bovine embryos in vitro produced by fusing takin ear fibroblasts with enucleated bovine oocytes and examine the fate of mitochondrial DNA in these embryos. We also compare the blastocyst development of takin-bovine embryos with yak-bovine and bovine-bovine embryos and compare the cell numbers of the blastocyst. Our results indicate that: (1) takin-bovine cloned embryos can develop to the blastocyst stage in vitro (5%), (2) blastocyst mitochondria DNA are derived primarily from bovine oocytes in spite of a little takin donor cell mitochondrial DNA, (3) using the same cloned protocol, development efficiency is significantly different between bovine-bovine cloning, yak-bovine, and takin-bovine cloning (48 vs. 28% vs. 5%, P < 0.01), and (4) cell numbers in the blastocysts of the three species of embryos were not different. These results suggest that the bovine oocytes can reprogram the takin, yak, and bovine fibroblast nuclei. However, the development efficiency of intra-species cloning tends to be higher than inter-species cloning; the more close the species of the donor cell is to the recipient oocyte (yak versus takin), the greater the blastocyst development in vitro.

  20. Hardwiring microbes via direct interspecies electron transfer: mechanisms and applications.

    PubMed

    Cheng, Qiwen; Call, Douglas F

    2016-08-10

    Multicellular microbial communities are important catalysts in engineered systems designed to treat wastewater, remediate contaminated sediments, and produce energy from biomass. Understanding the interspecies interactions within them is therefore essential to design effective processes. The flow of electrons within these communities is especially important in the determination of reaction possibilities (thermodynamics) and rates (kinetics). Conventional models of electron transfer incorporate the diffusion of metabolites generated by one organism and consumed by a second, frequently referred to as mediated interspecies electron transfer (MIET). Evidence has emerged in the last decade that another method, called direct interspecies electron transfer (DIET), may occur between organisms or in conjunction with electrically conductive materials. Recent research has suggested that DIET can be stimulated in engineered systems to improve desired treatment goals and energy recovery in systems such as anaerobic digesters and microbial electrochemical technologies. In this review, we summarize the latest understanding of DIET mechanisms, the associated microorganisms, and the underlying thermodynamics. We also critically examine approaches to stimulate DIET in engineered systems and assess their effectiveness. We find that in most cases attempts to promote DIET in mixed culture systems do not yield the improvements expected based on defined culture studies. Uncertainties of other processes that may be co-occurring in real systems, such as contaminant sorption and biofilm promotion, need to be further investigated. We conclude by identifying areas of future research related to DIET and its application in biological treatment processes. PMID:27349520

  1. Direct Interspecies Electron Transfer between Geobacter metallireducens and Methanosarcina barkeri

    PubMed Central

    Shrestha, Pravin Malla; Liu, Fanghua; Markovaite, Beatrice; Chen, Shanshan; Nevin, Kelly P.; Lovley, Derek R.

    2014-01-01

    Direct interspecies electron transfer (DIET) is potentially an effective form of syntrophy in methanogenic communities, but little is known about the diversity of methanogens capable of DIET. The ability of Methanosarcina barkeri to participate in DIET was evaluated in coculture with Geobacter metallireducens. Cocultures formed aggregates that shared electrons via DIET during the stoichiometric conversion of ethanol to methane. Cocultures could not be initiated with a pilin-deficient G. metallireducens strain, suggesting that long-range electron transfer along pili was important for DIET. Amendments of granular activated carbon permitted the pilin-deficient G. metallireducens isolates to share electrons with M. barkeri, demonstrating that this conductive material could substitute for pili in promoting DIET. When M. barkeri was grown in coculture with the H2-producing Pelobacter carbinolicus, incapable of DIET, M. barkeri utilized H2 as an electron donor but metabolized little of the acetate that P. carbinolicus produced. This suggested that H2, but not electrons derived from DIET, inhibited acetate metabolism. P. carbinolicus-M. barkeri cocultures did not aggregate, demonstrating that, unlike DIET, close physical contact was not necessary for interspecies H2 transfer. M. barkeri is the second methanogen found to accept electrons via DIET and the first methanogen known to be capable of using either H2 or electrons derived from DIET for CO2 reduction. Furthermore, M. barkeri is genetically tractable, making it a model organism for elucidating mechanisms by which methanogens make biological electrical connections with other cells. PMID:24837373

  2. Syntrophic growth via quinone-mediated interspecies electron transfer

    PubMed Central

    Smith, Jessica A.; Nevin, Kelly P.; Lovley, Derek R.

    2015-01-01

    The mechanisms by which microbial species exchange electrons are of interest because interspecies electron transfer can expand the metabolic capabilities of microbial communities. Previous studies with the humic substance analog anthraquinone-2,6-disulfonate (AQDS) suggested that quinone-mediated interspecies electron transfer (QUIET) is feasible, but it was not determined if sufficient energy is available from QUIET to support the growth of both species. Furthermore, there have been no previous studies on the mechanisms for the oxidation of anthrahydroquinone-2,6-disulfonate (AHQDS). A co-culture of Geobacter metallireducens and G. sulfurreducens metabolized ethanol with the reduction of fumarate much faster in the presence of AQDS, and there was an increase in cell protein. G. sulfurreducens was more abundant, consistent with G. sulfurreducens obtaining electrons from acetate that G. metallireducens produced from ethanol, as well as from AHQDS. Co-cultures initiated with a citrate synthase-deficient strain of G. sulfurreducens that was unable to use acetate as an electron donor also metabolized ethanol with the reduction of fumarate and cell growth, but acetate accumulated over time. G. sulfurreducens and G. metallireducens were equally abundant in these co-cultures reflecting the inability of the citrate synthase-deficient strain of G. sulfurreducens to metabolize acetate. Evaluation of the mechanisms by which G. sulfurreducens accepts electrons from AHQDS demonstrated that a strain deficient in outer-surface c-type cytochromes that are required for AQDS reduction was as effective at QUIET as the wild-type strain. Deletion of additional genes previously implicated in extracellular electron transfer also had no impact on QUIET. These results demonstrate that QUIET can yield sufficient energy to support the growth of both syntrophic partners, but that the mechanisms by which electrons are derived from extracellular hydroquinones require further investigation. PMID

  3. Multidimensional modelling to investigate interspecies hydrogen transfer in anaerobic biofilms.

    PubMed

    Batstone, D J; Picioreanu, C; van Loosdrecht, M C M

    2006-09-01

    Anaerobic digestion is a multistep process, mediated by a functionally and phylogenetically diverse microbial population. One of the crucial steps is oxidation of organic acids, with electron transfer via hydrogen or formate from acetogenic bacteria to methanogens. This syntrophic microbiological process is strongly restricted by a thermodynamic limitation on the allowable hydrogen or formate concentration. In order to study this process in more detail, we developed an individual-based biofilm model which enables to describe the processes at a microbial resolution. The biochemical model is the ADM1, implemented in a multidimensional domain. With this model, we evaluated three important issues for the syntrophic relationship: (i) Is there a fundamental difference in using hydrogen or formate as electron carrier? (ii) Does a thermodynamic-based inhibition function produced substantially different results from an empirical function? and; (iii) Does the physical co-location of acetogens and methanogens follow directly from a general model. Hydrogen or formate as electron carrier had no substantial impact on model results. Standard inhibition functions or thermodynamic inhibition function gave similar results at larger substrate field grid sizes (> 10 microm), but at smaller grid sizes, the thermodynamic-based function reduced the number of cells with long interspecies distances (> 2.5 microm). Therefore, a very fine grid resolution is needed to reflect differences between the thermodynamic function, and a more generic inhibition form. The co-location of syntrophic bacteria was well predicted without a need to assume a microbiological based mechanism (e.g., through chemotaxis) of biofilm formation.

  4. Incompatibility between Nuclear and Mitochondrial Genomes Contributes to an Interspecies Reproductive Barrier.

    PubMed

    Ma, Hong; Marti Gutierrez, Nuria; Morey, Robert; Van Dyken, Crystal; Kang, Eunju; Hayama, Tomonari; Lee, Yeonmi; Li, Ying; Tippner-Hedges, Rebecca; Wolf, Don P; Laurent, Louise C; Mitalipov, Shoukhrat

    2016-08-01

    Vertebrate cells carry two different genomes, nuclear (nDNA) and mitochondrial (mtDNA), both encoding proteins involved in oxidative phosphorylation. Because of the extensive interactions, adaptive coevolution of the two genomes must occur to ensure normal mitochondrial function. To investigate whether incompatibilities between these two genomes could contribute to interspecies reproductive barriers, we performed reciprocal mtDNA replacement (MR) in zygotes between widely divergent Mus m. domesticus (B6) and conplastic Mus m. musculus (PWD) mice. Transfer of MR1 cybrid embryos (B6nDNA-PWDmtDNA) supported normal development of F1 offspring with reduced male fertility but unaffected reproductive fitness in females. Furthermore, donor PWD mtDNA was faithfully transmitted through the germline into F2 and F3 generations. In contrast, reciprocal MR2 (PWDnDNA-B6mtDNA) produced high embryonic loss and stillborn rates, suggesting an association between mitochondrial function and infertility. These results strongly suggest that functional incompatibility between nuclear and mitochondrial genomes contributes to interspecies reproductive isolation in mammals. PMID:27425585

  5. Carbon cloth stimulates direct interspecies electron transfer in syntrophic co-cultures.

    PubMed

    Chen, Shanshan; Rotaru, Amelia-Elena; Liu, Fanghua; Philips, Jo; Woodard, Trevor L; Nevin, Kelly P; Lovley, Derek R

    2014-12-01

    This study investigated the possibility that the electrical conductivity of carbon cloth accelerates direct interspecies electron transfer (DIET) in co-cultures. Carbon cloth accelerated metabolism of DIET co-cultures (Geobacter metallireducens-Geobacter sulfurreducens and G.metallireducens-Methanosarcina barkeri) but did not promote metabolism of co-cultures performing interspecies H2 transfer (Desulfovibrio vulgaris-G.sulfurreducens). On the other hand, DIET co-cultures were not stimulated by poorly conductive cotton cloth. Mutant strains lacking electrically conductive pili, or pili-associated cytochromes participated in DIET only in the presence of carbon cloth. In co-cultures promoted by carbon cloth, cells were primarily associated with the cloth although the syntrophic partners were too far apart for cell-to-cell biological electrical connections to be feasible. Carbon cloth seemingly mediated interspecies electron transfer between the distant syntrophic partners. These results suggest that the ability of carbon cloth to accelerate DIET should be considered in anaerobic digester designs that incorporate carbon cloth.

  6. Assessing functional annotation transfers with inter-species conserved coexpression: application to Plasmodium falciparum

    PubMed Central

    2010-01-01

    Background Plasmodium falciparum is the main causative agent of malaria. Of the 5 484 predicted genes of P. falciparum, about 57% do not have sufficient sequence similarity to characterized genes in other species to warrant functional assignments. Non-homology methods are thus needed to obtain functional clues for these uncharacterized genes. Gene expression data have been widely used in the recent years to help functional annotation in an intra-species way via the so-called Guilt By Association (GBA) principle. Results We propose a new method that uses gene expression data to assess inter-species annotation transfers. Our approach starts from a set of likely orthologs between a reference species (here S. cerevisiae and D. melanogaster) and a query species (P. falciparum). It aims at identifying clusters of coexpressed genes in the query species whose coexpression has been conserved in the reference species. These conserved clusters of coexpressed genes are then used to assess annotation transfers between genes with low sequence similarity, enabling reliable transfers of annotations from the reference to the query species. The approach was used with transcriptomic data sets of P. falciparum, S. cerevisiae and D. melanogaster, and enabled us to propose with high confidence new/refined annotations for several dozens hypothetical/putative P. falciparum genes. Notably, we revised the annotation of genes involved in ribosomal proteins and ribosome biogenesis and assembly, thus highlighting several potential drug targets. Conclusions Our approach uses both sequence similarity and gene expression data to help inter-species gene annotation transfers. Experiments show that this strategy improves the accuracy achieved when using solely sequence similarity and outperforms the accuracy of the GBA approach. In addition, our experiments with P. falciparum show that it can infer a function for numerous hypothetical genes. PMID:20078859

  7. Collisionless inter-species energy transfer and turbulent heating in drift wave turbulence

    SciTech Connect

    Zhao, L.; Diamond, P. H.

    2012-08-15

    We reconsider the classic problems of calculating 'turbulent heating' and collisionless inter-species transfer of energy in drift wave turbulence. These issues are of interest for low collisionality, electron heated plasmas, such as ITER, where collisionless energy transfer from electrons to ions is likely to be significant. From the wave Poynting theorem at steady state, a volume integral over an annulus r{sub 1}=-S{sub r}|{sub r{sub 1}{sup r{sub 2}}}{ne}0. Here S{sub r} is the wave energy density flux in the radial direction. Thus, a wave energy flux differential across an annular region indeed gives rise to a net heating, in contrast to previous predictions. This heating is related to the Reynolds work by the zonal flow, since S{sub r} is directly linked to the zonal flow drive. In addition to net heating, there is inter-species heat transfer. For collisionless electron drift waves, the total turbulent energy source for collisionless heat transfer is due to quasilinear electron cooling. Subsequent quasilinear ion heating occurs through linear ion Landau damping. In addition, perpendicular heating via ion polarization currents contributes to ion heating. Since at steady state, Reynolds work of the turbulence on the zonal flow must balance zonal flow frictional damping ({approx}{nu}{sub ii}{sup 2}{approx}|(e{phi}(tilde sign)/T)|{sup 4}), it is no surprise that zonal flow friction appears as an important channel for ion heating. This process of energy transfer via zonal flow has not previously been accounted for in analyses of energy transfer. As an application, we compare the rate of turbulent energy transfer in a low collisionality plasma with the rate of the energy transfer by collisions. The result shows that the collisionless turbulent energy transfer is a significant energy coupling process for ITER plasma.

  8. Potential for Direct Interspecies Electron Transfer in Methanogenic Wastewater Digester Aggregates

    PubMed Central

    Morita, Masahiko; Malvankar, Nikhil S.; Franks, Ashley E.; Summers, Zarath M.; Giloteaux, Ludovic; Rotaru, Amelia E.; Rotaru, Camelia; Lovley, Derek R.

    2011-01-01

    ABSTRACT Mechanisms for electron transfer within microbial aggregates derived from an upflow anaerobic sludge blanket reactor converting brewery waste to methane were investigated in order to better understand the function of methanogenic consortia. The aggregates were electrically conductive, with conductivities 3-fold higher than the conductivities previously reported for dual-species aggregates of Geobacter species in which the two species appeared to exchange electrons via interspecies electron transfer. The temperature dependence response of the aggregate conductance was characteristic of the organic metallic-like conductance previously described for the conductive pili of Geobacter sulfurreducens and was inconsistent with electron conduction through minerals. Studies in which aggregates were incubated with high concentrations of potential electron donors demonstrated that the aggregates had no significant capacity for conversion of hydrogen to methane. The aggregates converted formate to methane but at rates too low to account for the rates at which that the aggregates syntrophically metabolized ethanol, an important component of the reactor influent. Geobacter species comprised 25% of 16S rRNA gene sequences recovered from the aggregates, suggesting that Geobacter species may have contributed to some but probably not all of the aggregate conductivity. Microorganisms most closely related to the acetate-utilizing Methanosaeta concilii accounted for more than 90% of the sequences that could be assigned to methane producers, consistent with the poor capacity for hydrogen and formate utilization. These results demonstrate for the first time that methanogenic wastewater aggregates can be electrically conductive and suggest that direct interspecies electron transfer could be an important mechanism for electron exchange in some methanogenic systems. PMID:21862629

  9. Analysis of nuclear export using photoactivatable GFP fusion proteins and interspecies heterokaryons.

    PubMed

    Nakrieko, Kerry-Ann; Ivanova, Iordanka A; Dagnino, Lina

    2010-01-01

    In this chapter, we review protocols for the analysis of nucleocytoplasmic shuttling of transcription factors and nuclear proteins, using two different approaches. The first involves the use of photoactivatable forms of the protein of interest by fusion to photoactivatable green fluorescent protein to follow its movement out of the nucleus by live-cell confocal microscopy. This methodology allows for the kinetic characterization of protein movements as well as measurement of steady-state levels. In a second procedure to assess the ability of a nuclear protein to move into and out of the nucleus, we describe the use of interspecies heterokaryon assays, which provide a measurement of steady-state distribution. These technologies are directly applicable to the analysis of nucleocytoplasmic movements not only of transcription factors, but also other nuclear proteins.

  10. Interspecies distances between propionic acid degraders and methanogens in syntrophic consortia for optimal hydrogen transfer.

    PubMed

    Felchner-Zwirello, Monika; Winter, Josef; Gallert, Claudia

    2013-10-01

    A mixed culture from an anaerobic biowaste digester was enriched on propionate and used to investigate interspecies hydrogen transfer in dependence of spatial distances between propionate degraders and methanogens. From 20.3 mM propionate, 20.8 mM acetate and 15.5 mM methane were formed. Maximum specific propionate oxidation and methane formation rates were 49 and 23 mmol mg(-1) day(-1), respectively. Propionate oxidation was inhibited by only 20 mM acetate by about 50 %. Intermediate formate formation during inhibited methanogensis was observed. The spatial distribution and the biovolume fraction of propionate degraders and of methanogens in relation to the total population during aggregate formation were determined. Measurements of interbacterial distances were conducted with fluorescence in situ hybridization by application of group-specific 16S rRNA-targeted probes and 3D image analyses. With increasing incubation time, floc formation and growth up to 54 μm were observed. Propionate degraders and methanogens were distributed randomly in the flocs. The methanogenic biovolume fraction was high at the beginning and remained constant over 42 days, whereas the fraction of propionate degraders increased with time during propionate feeding. Interbacterial distances between propionate degraders and methanogens decreased with time from 5.30 to 0.29 μm, causing an increase of the maximum possible hydrogen flux from 1.1 to 10.3 nmol ml(-1) min(-1). The maximum possible hydrogen flux was always higher than the hydrogen formation and consumption rate, indicating that reducing the interspecies distance by aggregation is advantageous in complex ecosystems.

  11. Intra- and interspecies transfer and expression of Rhizobium japonicum hydrogen uptake genes and autotrophic growth capability

    PubMed Central

    Lambert, Grant R.; Cantrell, Michael A.; Hanus, F. Joe; Russell, Sterling A.; Haddad, Karen R.; Evans, Harold J.

    1985-01-01

    Cosmids containing hydrogen uptake genes have previously been isolated in this laboratory. Four new cosmids that contain additional hup gene(s) have now been identified by conjugal transfer of a Rhizobium japonicum 122DES gene bank into a Tn5-generated Hup- mutant and screening for the acquisition of Hup activity. The newly isolated cosmids, pHU50-pHU53, contain part of the previously isolated pHU1 but extend as far as 20 kilobases beyond its border. pHU52 complements five of six Hup- mutants and confers activity on several Hup- wild-type R. japonicum strains in the free-living state and where tested in nodules. Transconjugants obtained from interspecies transfer of pHU52 to Rhizobium meliloti 102F28, 102F32, and 102F51 and Rhizobium leguminosarum 128C53 showed hydrogen-dependent methyleneblue reduction, performed the oxyhydrogen reaction, and showed hydrogen-dependent autotrophic growth by virtue of the introduced genes. The identity of the presumptive transconjugants was confirmed by antibiotic-resistance profiles and by plant nodulation tests. Images PMID:16578786

  12. Direct interspecies electron transfer accelerates syntrophic oxidation of butyrate in paddy soil enrichments.

    PubMed

    Li, Huijuan; Chang, Jiali; Liu, Pengfei; Fu, Li; Ding, Dewen; Lu, Yahai

    2015-05-01

    Syntrophic interaction occurs during anaerobic fermentation of organic substances forming methane as the final product. H2 and formate are known to serve as the electron carriers in this process. Recently, it has been shown that direct interspecies electron transfer (DIET) occurs for syntrophic CH4 production from ethanol and acetate. Here, we constructed paddy soil enrichments to determine the involvement of DIET in syntrophic butyrate oxidation and CH4 production. The results showed that CH4 production was significantly accelerated in the presence of nanoFe3 O4 in all continuous transfers. This acceleration increased with the increase of nanoFe3 O4 concentration but was dismissed when Fe3 O4 was coated with silica that insulated the mineral from electrical conduction. NanoFe3 O4 particles were found closely attached to the cell surfaces of different morphology, thus bridging cell connections. Molecular approaches, including DNA-based stable isotope probing, revealed that the bacterial Syntrophomonadaceae and Geobacteraceae, and the archaeal Methanosarcinaceae, Methanocellales and Methanobacteriales, were involved in the syntrophic butyrate oxidation and CH4 production. Among them, the growth of Geobacteraceae strictly relied on the presence of nanoFe3 O4 and its electrical conductivity in particular. Other organisms, except Methanobacteriales, were present in enrichments regardless of nanoFe3 O4 amendment. Collectively, our study demonstrated that the nanoFe3 O4 -facilitated DIET occurred in syntrophic CH4 production from butyrate, and Geobacter species played the key role in this process in the paddy soil enrichments.

  13. Communities stimulated with ethanol to perform direct interspecies electron transfer for syntrophic metabolism of propionate and butyrate.

    PubMed

    Zhao, Zhiqiang; Zhang, Yaobin; Yu, Qilin; Dang, Yan; Li, Yang; Quan, Xie

    2016-10-01

    Direct interspecies electron transfer (DIET) has been considered as an alternative to interspecies H2 transfer (IHT) for syntrophic metabolism, but the microorganisms capable of metabolizing the key intermediates, such as propionate and butyrate, via DIET have yet to be described. A strategy of culturing the enrichments with ethanol as a DIET substrate to stimulate the communities for the syntrophic metabolism of propionate and/or butyrate was proposed in this study. The results showed that the syntrophic propionate and/or butyrate degradation was significantly improved in the ethanol-stimulated reactor when propionate/butyrate was the sole carbon source. The conductivity of the ethanol-stimulated enrichments was as 5 folds (for propionate)/76 folds (for butyrate) as that of the traditional enrichments (never ethanol fed). Microbial community analysis revealed that Geobacter species known to proceed DIET were only detected in the ethanol-stimulated enrichments. Together with the significant increase of Methanosaeta and Methanosarcina species in these enrichments, the potential DIET between Geobacter and Methanosaeta or Methanosarcina species might be established to improve the syntrophic propionate and/or butyrate degradation. Further experiments demonstrated that granular activated carbon (GAC) could improve the syntrophic metabolism of propionate and/or butyrate of the ethanol-stimulated enrichments, while almost no effects on the traditional enrichments. Also, the high H2 partial pressure could inhibit the syntrophic propionate and/or butyrate degradation of the traditional enrichments, but its effect on that of the ethanol-stimulated enrichments was negligible. PMID:27403870

  14. Expanding the Diet for DIET: Electron Donors Supporting Direct Interspecies Electron Transfer (DIET) in Defined Co-Cultures

    PubMed Central

    Wang, Li-Ying; Nevin, Kelly P.; Woodard, Trevor L.; Mu, Bo-Zhong; Lovley, Derek R.

    2016-01-01

    Direct interspecies electron transfer (DIET) has been recognized as an alternative to interspecies H2 transfer as a mechanism for syntrophic growth, but previous studies on DIET with defined co-cultures have only documented DIET with ethanol as the electron donor in the absence of conductive materials. Co-cultures of Geobacter metallireducens and Geobacter sulfurreducens metabolized propanol, butanol, propionate, and butyrate with the reduction of fumarate to succinate. G. metallireducens utilized each of these substrates whereas only electrons available from DIET supported G. sulfurreducens respiration. A co-culture of G. metallireducens and a strain of G. sulfurreducens that could not metabolize acetate oxidized acetate with fumarate as the electron acceptor, demonstrating that acetate can also be syntrophically metabolized via DIET. A co-culture of G. metallireducens and Methanosaeta harundinacea previously shown to syntrophically convert ethanol to methane via DIET metabolized propanol or butanol as the sole electron donor, but not propionate or butyrate. The stoichiometric accumulation of propionate or butyrate in the propanol- or butanol-fed cultures demonstrated that M. harundinaceae could conserve energy to support growth solely from electrons derived from DIET. Co-cultures of G. metallireducens and Methanosarcina barkeri could also incompletely metabolize propanol and butanol and did not metabolize propionate or butyrate as sole electron donors. These results expand the range of substrates that are known to be syntrophically metabolized through DIET, but suggest that claims of propionate and butyrate metabolism via DIET in mixed microbial communities warrant further validation. PMID:26973614

  15. Potential for direct interspecies electron transfer in an electric-anaerobic system to increase methane production from sludge digestion

    PubMed Central

    Zhao, Zhiqiang; Zhang, Yaobin; Wang, Liying; Quan, Xie

    2015-01-01

    Direct interspecies electron transfer (DIET) between Geobacter species and Methanosaeta species is an alternative to interspecies hydrogen transfer (IHT) in anaerobic digester, which however has not been established in anaerobic sludge digestion as well as in bioelectrochemical systems yet. In this study, it was found that over 50% of methane production of an electric-anaerobic sludge digester was resulted from unknown pathway. Pyrosequencing analysis revealed that Geobacter species were significantly enriched with electrodes. Fluorescence in situ hybridization (FISH) further confirmed that the dominant Geobacter species enriched belonged to Geobacter metallireducens. Together with Methanosaeta species prevailing in the microbial communities, the direct electron exchange between Geobacter species and Methanosaeta species might be an important reason for the “unknown” increase of methane production. Conductivity of the sludge in this electric-anaerobic digester was about 30% higher than that of the sludge in a control digester without electrodes. This study not only revealed for the first time that DIET might be the important mechanism on the methanogenesis of bioelectrochemical system, but also provided a new method to enhance DIET by means of bioelectric enrichment of Geobacter species. PMID:26057581

  16. Expanding the Diet for DIET: Electron Donors Supporting Direct Interspecies Electron Transfer (DIET) in Defined Co-Cultures.

    PubMed

    Wang, Li-Ying; Nevin, Kelly P; Woodard, Trevor L; Mu, Bo-Zhong; Lovley, Derek R

    2016-01-01

    Direct interspecies electron transfer (DIET) has been recognized as an alternative to interspecies H2 transfer as a mechanism for syntrophic growth, but previous studies on DIET with defined co-cultures have only documented DIET with ethanol as the electron donor in the absence of conductive materials. Co-cultures of Geobacter metallireducens and Geobacter sulfurreducens metabolized propanol, butanol, propionate, and butyrate with the reduction of fumarate to succinate. G. metallireducens utilized each of these substrates whereas only electrons available from DIET supported G. sulfurreducens respiration. A co-culture of G. metallireducens and a strain of G. sulfurreducens that could not metabolize acetate oxidized acetate with fumarate as the electron acceptor, demonstrating that acetate can also be syntrophically metabolized via DIET. A co-culture of G. metallireducens and Methanosaeta harundinacea previously shown to syntrophically convert ethanol to methane via DIET metabolized propanol or butanol as the sole electron donor, but not propionate or butyrate. The stoichiometric accumulation of propionate or butyrate in the propanol- or butanol-fed cultures demonstrated that M. harundinaceae could conserve energy to support growth solely from electrons derived from DIET. Co-cultures of G. metallireducens and Methanosarcina barkeri could also incompletely metabolize propanol and butanol and did not metabolize propionate or butyrate as sole electron donors. These results expand the range of substrates that are known to be syntrophically metabolized through DIET, but suggest that claims of propionate and butyrate metabolism via DIET in mixed microbial communities warrant further validation. PMID:26973614

  17. Potential for direct interspecies electron transfer in an electric-anaerobic system to increase methane production from sludge digestion.

    PubMed

    Zhao, Zhiqiang; Zhang, Yaobin; Wang, Liying; Quan, Xie

    2015-06-09

    Direct interspecies electron transfer (DIET) between Geobacter species and Methanosaeta species is an alternative to interspecies hydrogen transfer (IHT) in anaerobic digester, which however has not been established in anaerobic sludge digestion as well as in bioelectrochemical systems yet. In this study, it was found that over 50% of methane production of an electric-anaerobic sludge digester was resulted from unknown pathway. Pyrosequencing analysis revealed that Geobacter species were significantly enriched with electrodes. Fluorescence in situ hybridization (FISH) further confirmed that the dominant Geobacter species enriched belonged to Geobacter metallireducens. Together with Methanosaeta species prevailing in the microbial communities, the direct electron exchange between Geobacter species and Methanosaeta species might be an important reason for the "unknown" increase of methane production. Conductivity of the sludge in this electric-anaerobic digester was about 30% higher than that of the sludge in a control digester without electrodes. This study not only revealed for the first time that DIET might be the important mechanism on the methanogenesis of bioelectrochemical system, but also provided a new method to enhance DIET by means of bioelectric enrichment of Geobacter species.

  18. Nuclear transfer in ruminants.

    PubMed

    Lee, Joon-Hee; Maalouf, Walid E

    2015-01-01

    Ruminants were the first mammalian species to be cloned successfully by nuclear transplantation. Those experiments were designed to multiply high merit animals (Willadsen, Nature 320(6057):63-65, 1986; Prather et al., Biol Reprod 37(4):859-866, 1987; Wilmut et al., Nature 385(6619):810-813, 1997). Since then, cloning has provided us with a vast amount of knowledge and information on the reprogramming ability of somatic cells to different cell types which became an important basis for stem cell research and human medicine. Nowadays, the goals of most nuclear transfer work vary widely but in most cases the micromanipulation procedures remain the same. However, differences between species require different technical considerations. In this chapter, we describe in detail somatic cell nuclear transfer which is the foremost method for cloning ruminants with specific reference to sheep and cattle.

  19. Interspecies Extrapolation

    EPA Science Inventory

    Interspecies extrapolation encompasses two related but distinct topic areas that are germane to quantitative extrapolation and hence computational toxicology-dose scaling and parameter scaling. Dose scaling is the process of converting a dose determined in an experimental animal ...

  20. Control of interspecies electron flow during anaerobic digestion: significance of formate transfer versus hydrogen transfer during syntrophic methanogenesis in flocs. [Methanobacterium formicicum; Desulfovibrio vulgaris

    SciTech Connect

    Thiele, J.H.; Zeikus, J.G.

    1988-01-01

    Microbial formate production and consumption during syntrophic conversion of ethanol or lactate to methane was examined in purified flocs and digestor contents obtained from a whey-processing digestor. Formate production by digestor contents or purified digestor flocs was dependent on CO/sub 2/ and either ethanol or lactate but not H/sub 2/ gas as an electron donor. Floc preparations accumulated fourfold-higher levels of formate (40 ..mu..M) than digestor contents, and the free flora was the primary site for formate cleavage to CO/sub 2/ and H/sub 2/ (90 ..mu..M formate per h). Inhibition of methanogenesis by CHCl/sub 3/ resulted in formate accumulation and suppression of syntrophic ethanol oxidation. H/sub 2/ gas was an insignificant intermediary metabolite of syntrophic ethanol conversion by flocs, and it exogenous addition neither stimulated methanogenes nor inhibited the initial rate of ethanol oxidation. These results demonstrated that >90% of the syntrophic ethanol conversion to methane by mixed cultures containing primarily Desulfovibrio vulgaris and Methanobacterium formicicum was mediated via interspecies formate transfer and the <10% was mediated via interspecies H/sub 2/ transfer. The results are discussed in relation to biochemical thermodynamics. A model is presented which describes the dynamics of a bicarbonate-formate electron shuttle mechanism for control of carbon and electron flow during syntrophic methanogenesis and provides a novel mechanism for energy conservation by syntrophic acetogens.

  1. Interspecies Gene Transfer as a Method for Understanding the Genetic Basis for Evolutionary Change: Progress, Pitfalls, and Prospects

    PubMed Central

    Nikolov, Lachezar A.; Tsiantis, Miltos

    2015-01-01

    The recent revolution in high throughput sequencing and associated applications provides excellent opportunities to catalog variation in DNA sequences and gene expression between species. However, understanding the astonishing diversity of the Tree of Life requires understanding the phenotypic consequences of such variation and identification of those rare genetic changes that are causal to diversity. One way to study the genetic basis for trait diversity is to apply a transgenic approach and introduce genes of interest from a donor into a recipient species. Such interspecies gene transfer (IGT) is based on the premise that if a gene is causal to the morphological divergence of the two species, the transfer will endow the recipient with properties of the donor. Extensions of this approach further allow identifying novel loci for the diversification of form and investigating cis- and trans-contributions to morphological evolution. Here we review recent examples from both plant and animal systems that have employed IGT to provide insight into the genetic basis of evolutionary change. We outline the practice of IGT, its methodological strengths and weaknesses, and consider guidelines for its application, emphasizing the importance of phylogenetic distance, character polarity, and life history. We also discuss future perspectives for exploiting IGT in the context of expanding genomic resources in emerging experimental systems and advances in genome editing. PMID:26734038

  2. [Nuclear transfer and therapeutic cloning].

    PubMed

    Xu, Xiao-Ming; Lei, An-Min; Hua, Jin-Lian; Dou, Zhong-Ying

    2005-03-01

    Nuclear transfer and therapeutic cloning have widespread and attractive prospects in animal agriculture and biomedical applications. We reviewed that the quality of oocytes and nuclear reprogramming of somatic donor cells were the main reasons of the common abnormalities in cloned animals and the low efficiency of cloning and showed the problems and outlets in therapeutic cloning, such as some basic problems in nuclear transfer affected clinical applications of therapeutic cloning. Study on isolation and culture of nuclear transfer embryonic stem (ntES) cells and specific differentiation of ntES cells into important functional cells should be emphasized and could enhance the efficiency. Adult stem cells could help to cure some great diseases, but could not replace therapeutic cloning. Ethics also impeded the development of therapeutic cloning. It is necessary to improve many techniques and reinforce the research of some basic theories, then somatic nuclear transfer and therapeutic cloning may apply to agriculture reproduction and benefit to human life better.

  3. Transfer reactions in nuclear astrophysics

    NASA Astrophysics Data System (ADS)

    Bardayan, D. W.

    2016-08-01

    To a high degree many aspects of the large-scale behavior of objects in the Universe are governed by the underlying nuclear physics. In fact the shell structure of nuclear physics is directly imprinted into the chemical abundances of the elements. The tranquility of the night sky is a direct result of the relatively slow rate of nuclear reactions that control and determines a star’s fate. Understanding the nuclear structure and reaction rates between nuclei is vital to understanding our Universe. Nuclear-transfer reactions make accessible a wealth of knowledge from which we can extract much of the required nuclear physics information. A review of transfer reactions for nuclear astrophysics is presented with an emphasis on the experimental challenges and opportunities for future development.

  4. N. elongata produces type IV pili that mediate interspecies gene transfer with N. gonorrhoeae.

    PubMed

    Higashi, Dustin L; Biais, Nicolas; Weyand, Nathan J; Agellon, Al; Sisko, Jennifer L; Brown, Lewis M; So, Magdalene

    2011-01-01

    The genus Neisseria contains at least eight commensal and two pathogenic species. According to the Neisseria phylogenetic tree, commensals are basal to the pathogens. N. elongata, which is at the opposite end of the tree from N. gonorrhoeae, has been observed to be fimbriated, and these fimbriae are correlated with genetic competence in this organism. We tested the hypothesis that the fimbriae of N. elongata are Type IV pili (Tfp), and that Tfp functions in genetic competence. We provide evidence that the N. elongata fimbriae are indeed Tfp. Tfp, as well as the DNA Uptake Sequence (DUS), greatly enhance N. elongata DNA transformation. Tfp allows N. elongata to make intimate contact with N. gonorrhoeae and to mediate the transfer of antibiotic resistance markers between these two species. We conclude that Tfp functional for genetic competence is a trait of a commensal member of the Neisseria genus. Our findings provide a mechanism for the horizontal gene transfer that has been observed among Neisseria species. PMID:21731720

  5. Interspecies Transfer and Regulation of Pseudomonas stutzeri A1501 Nitrogen Fixation Island in Escherichia coli.

    PubMed

    Han, Yunlei; Lu, Na; Chen, Qinghua; Zhan, Yuhua; Liu, Wei; Lu, Wei; Zhu, Baoli; Lin, Min; Yang, Zhirong; Yan, Yongliang

    2015-08-01

    Until now, considerable effort has been made to engineer novel nitrogen-fixing organisms through the transfer of nif genes from various diazotrophs to non-nitrogen fixers; however, regulatory coupling of the heterologous nif genes with the regulatory system of the new host is still not well understood. In this work, a 49 kb nitrogen fixation island from P. stutzeri A1501 was transferred into E. coli using a novel and efficient transformation strategy, and a series of recombinant nitrogen-fixing E. coli strains were obtained. We found that the nitrogenase activity of the recombinant E. coli strain EN-01, similar to the parent strain P. stutzeri A1501, was dependent on external ammonia concentration, oxygen tension, and temperature. We further found that there existed a regulatory coupling between the E. coli general nitrogen regulatory system and the heterologous P. stutzeri nif island in the recombinant E. coli strain. We also provided evidence that the E. coli general nitrogen regulator GlnG protein was involved in the activation of the nif-specific regulator NifA via a direct interaction with the NifA promoter. To the best of our knowledge, this work plays a groundbreaking role in increasing understanding of the regulatory coupling of the heterologous nitrogen fixation system with the regulatory system of the recipient host. Furthermore, it will shed light on the structure and functional integrity of the nif island and will be useful for the construction of novel and more robust nitrogen-fixing organisms through biosynthetic engineering.

  6. Development of a pheasant interspecies primordial germ cell transfer to chicken embryo: effect of donor cell sex on chimeric semen production.

    PubMed

    Kang, S J; Choi, J W; Park, K J; Lee, Y M; Kim, T M; Sohn, S H; Lim, J M; Han, J Y

    2009-09-01

    This study was conducted to evaluate whether the sex of donor primordial germ cells (PGCs) influences production of chimeric semen from recipient hatchlings produced by interspecies transfer between pheasant (Phasianus colchicus) and chicken (Gallus gallus). Pheasant PGCs were retrieved from 7-d-old embryos and subsequently transferred into circulatory blood of 2.5-d-old (Stage 17) embryos. The sex of embryos was discerned 3 to 6 days after laying, and in preliminary study, overall rate of embryo survival after sexing was 74.6% with male-to-female ratio of 0.49 to 0.51. In Experiment 1, magnetic-activated cell sorting (MACS) using QCR1 antibody was effective for enriching the population of male and female PGCs in gonadal cells (9.2- to 12.5-fold and 10.8- to 19.5-fold increase, respectively). In Experiment 2, an increase in the number of hatchlings producing chimeric semen was detected after the homosexual transfer of male-to-male compared with that after the heterosexual transfer of female-to-male (68% to 88%). Significant increase was found in the frequency of chimeric semen production (0.96 to 1.68 times); production of pheasant progenies by artificial insemination using chimeric semen was also increased in the homosexual transfer (0 to 3 cases). In conclusion, the homosexual PGC transfer of male-to-male yielded better rate of generating pheasant progenies after test cross-reproduction than that of the heterosexual transfer of female-to-male, which could improve the efficiency of interspecies germ cell transfer system.

  7. Advances in livestock nuclear transfer.

    PubMed

    Kühholzer, B; Prather, R S

    2000-09-01

    Cloning and transgenic animal production have been greatly enhanced by the development of nuclear transfer technology. In the past, genetic modification in domestic animals was not tightly controlled. With the nuclear transfer technology one can now create some domestic animals with specific genetic modifications. An ever-expanding variety of cell types have been successfully used as donors to create the clones. Both cell fusion and microinjection are successfully being used to create these animals. However, it is still not clear which stage(s) of the cell cycle for donor and recipient cells yield the greatest degree of development. While for the most part gene expression is reprogrammed in nuclear transfer embryos, all structural changes may not be corrected as evidenced by the length of the telomeres in sheep resulting from nuclear transfer. Even after these animals are created the question of "are they really clones?" arises due to mitochondrial inheritance from the donor cell versus the recipient oocyte. This review discusses these issues as they relate to livestock.

  8. Bovine somatic cell nuclear transfer.

    PubMed

    Ross, Pablo J; Cibelli, Jose B

    2010-01-01

    Somatic cell nuclear transfer (SCNT) is a technique by which the nucleus of a differentiated cell is introduced into an oocyte from which its genetic material has been removed by a process called enucleation. In mammals, the reconstructed embryo is artificially induced to initiate embryonic development (activation). The oocyte turns the somatic cell nucleus into an embryonic nucleus. This process is called nuclear reprogramming and involves an important change of cell fate, by which the somatic cell nucleus becomes capable of generating all the cell types required for the formation of a new individual, including extraembryonic tissues. Therefore, after transfer of a cloned embryo to a surrogate mother, an offspring genetically identical to the animal from which the somatic cells where isolated, is born. Cloning by nuclear transfer has potential applications in agriculture and biomedicine, but is limited by low efficiency. Cattle were the second mammalian species to be cloned after Dolly the sheep, and it is probably the most widely used species for SCNT experiments. This is, in part due to the high availability of bovine oocytes and the relatively higher efficiency levels usually obtained in cattle. Given the wide utilization of this species for cloning, several alternatives to this basic protocol can be found in the literature. Here we describe a basic protocol for bovine SCNT currently being used in our laboratory, which is amenable for the use of the nuclear transplantation technique for research or commercial purposes. PMID:20336522

  9. Potential enhancement of direct interspecies electron transfer for syntrophic metabolism of propionate and butyrate with biochar in up-flow anaerobic sludge blanket reactors.

    PubMed

    Zhao, Zhiqiang; Zhang, Yaobin; Holmes, Dawn E; Dang, Yan; Woodard, Trevor L; Nevin, Kelly P; Lovley, Derek R

    2016-06-01

    Promoting direct interspecies electron transfer (DIET) to enhance syntrophic metabolism may be a strategy for accelerating the conversion of organic wastes to methane, but microorganisms capable of metabolizing propionate and butyrate via DIET under methanogenic conditions have yet to be identified. In an attempt to establish methanogenic communities metabolizing propionate or butyrate with DIET, enrichments were initiated with up-flow anaerobic sludge blanket (UASB), similar to those that were previously reported to support communities that metabolized ethanol with DIET that relied on direct biological electrical connections. In the absence of any amendments, microbial communities enriched were dominated by microorganisms closely related to pure cultures that are known to metabolize propionate or butyrate to acetate with production of H2. When biochar was added to the reactors there was a substantial enrichment on the biochar surface of 16S rRNA gene sequences closely related to Geobacter and Methanosaeta species known to participate in DIET. PMID:26967338

  10. Nuclear fuel pellet transfer escalator

    SciTech Connect

    Huggins, T.B. Sr.; Roberts, E.; Edmunds, M.O.

    1991-09-17

    This patent describes a nuclear fuel pellet escalator for loading nuclear fuel pellets into a sintering boat. It comprises a generally horizontally-disposed pellet transfer conveyor for moving pellets in single file fashion from a receiving end to a discharge end thereof, the conveyor being mounted about an axis at its receiving end for pivotal movement to generally vertically move its discharge end toward and away from a sintering boat when placed below the discharge end of the conveyor, the conveyor including an elongated arm swingable vertically about the axis and having an elongated channel recessed below an upper side of the arm and extending between the receiving and discharge ends of the conveyor; a pellet dispensing chute mounted to the arm of the conveyor at the discharge end thereof and extending therebelow such that the chute is carried at the discharge end of the conveyor for generally vertical movement therewith toward and away from the sintering boat.

  11. High levels of mitochondrial heteroplasmy modify the development of ovine-bovine interspecies nuclear transferred embryos.

    PubMed

    Hua, Song; Lu, Chenglong; Song, Yakun; Li, Ruizhe; Liu, Xu; Quan, Fusheng; Wang, Yongsheng; Liu, Jun; Su, Feng; Zhang, Yong

    2012-01-01

    To investigate the effect of mitochondrial heteroplasmy on embryo development, cloned embryos produced using bovine oocytes as the recipient cytoplasm and ovine granulosa cells as the donor nuclei were complemented with 2pL mitochondrial suspension isolated from ovine (BOOMT embryos) or bovine (BOBMT embryos) granulosa cells; cloned embryos without mitochondrial injection served as the control group (BO embryos). Reverse transcription-quantitative polymerase chain reaction (RT-qPCR) and sodium bisulfite genomic sequencing were used to analyse mRNA and methylation levels of pluripotency genes (OCT4, SOX2) and mitochondrial genes (TFAM, POLRMT) in the early developmental stages of cloned embryos. The number of mitochondrial DNA copies in 2pL ovine-derived and bovine-derived mitochondrial suspensions was 960±110 and 1000±120, respectively. The blastocyst formation rates were similar in BOBMT and BO embryos (P>0.05), but significantly higher than in BOOMT embryos (P<0.01). Expression of OCT4 and SOX2, as detected by RT-qPCR, decreased significantly in BOOMT embryos (P<0.05), whereas the expression of TFAM and POLRMT increased significantly, compared with expression in BOOMT and BO embryos (P<0.05). In addition, methylation levels of OCT4 and SOX2 were significantly greater (P<0.05), whereas those of TFAM and POLRMT were significantly lower (P<0.01), in BOOMT embryos compared with BOBMT and BO embryos. Together, the results of the present study suggest that the degree of mitochondrial heteroplasmy may affect embryonic development.

  12. Nuclear reorganization barriers to electron transfer

    SciTech Connect

    Sutin, N.; Brunschwig, B.S.; Creutz, C.; Winkler, J.R.

    1988-01-01

    The nuclear barrier to electron transfer arises from the need for reorganization of intramolecular and solvent internuclear distances prior to electron transfer. For reactions with relatively small driving force (''normal'' free-energy region) the nuclear factors and rates increase as intrinsic inner-shell and outer-shell barriers decrease; this is illustrated by data for transition metal complexes in their ground electronic states. By contrast, in the inverted free-energy region, rates and nuclear factors decrease with decreasing ''intrinsic'' barriers; this is illustrated by data for the decay of charge-transfer excited states. Several approaches to the evaluation of the outer-shell barrier are explored in an investigation of the distance dependence of the nuclear factor in intramolecular electron-transfer processes. 39 refs., 14 figs., 3 tabs.

  13. Human somatic cell nuclear transfer and cloning.

    PubMed

    2012-10-01

    This document presents arguments that conclude that it is unethical to use somatic cell nuclear transfer (SCNT) for infertility treatment due to concerns about safety; the unknown impact of SCNT on children, families, and society; and the availability of other ethically acceptable means of assisted reproduction. This document replaces the ASRM Ethics Committee report titled, "Human somatic cell nuclear transfer (cloning)," last published in Fertil Steril 2000;74:873-6.

  14. Production of chicken progeny (Gallus gallus domesticus) from interspecies germline chimeric duck (Anas domesticus) by primordial germ cell transfer.

    PubMed

    Liu, Chunhai; Khazanehdari, Kamal A; Baskar, Vijaya; Saleem, Shazia; Kinne, Joerg; Wernery, Ulrich; Chang, Il-Kuk

    2012-04-01

    The present study aimed to investigate the differentiation of chicken (Gallus gallus domesticus) primordial germ cells (PGCs) in duck (Anas domesticus) gonads. Chimeric ducks were produced by transferring chicken PGCs into duck embryos. Transfer of 200 and 400 PGCs resulted in the detection of a total number of 63.0 ± 54.3 and 116.8 ± 47.1 chicken PGCs in the gonads of 7-day-old duck embryos, respectively. The chimeric rate of ducks prior to hatching was 52.9% and 90.9%, respectively. Chicken germ cells were assessed in the gonad of chimeric ducks with chicken-specific DNA probes. Chicken spermatogonia were detected in the seminiferous tubules of duck testis. Chicken oogonia, primitive and primary follicles, and chicken-derived oocytes were also found in the ovaries of chimeric ducks, indicating that chicken PGCs are able to migrate, proliferate, and differentiate in duck ovaries and participate in the progression of duck ovarian folliculogenesis. Chicken DNA was detected using PCR from the semen of chimeric ducks. A total number of 1057 chicken eggs were laid by Barred Rock hens after they were inseminated with chimeric duck semen, of which four chicken offspring hatched and one chicken embryo did not hatch. Female chimeric ducks were inseminated with chicken semen; however, no fertile eggs were obtained. In conclusion, these results demonstrated that chicken PGCs could interact with duck germinal epithelium and complete spermatogenesis and eventually give rise to functional sperm. The PGC-mediated germline chimera technology may provide a novel system for conserving endangered avian species.

  15. Dry Transfer Systems for Used Nuclear Fuel

    SciTech Connect

    Brett W. Carlsen; Michaele BradyRaap

    2012-05-01

    The potential need for a dry transfer system (DTS) to enable retrieval of used nuclear fuel (UNF) for inspection or repackaging will increase as the duration and quantity of fuel in dry storage increases. This report explores the uses for a DTS, identifies associated general functional requirements, and reviews existing and proposed systems that currently perform dry fuel transfers. The focus of this paper is on the need for a DTS to enable transfer of bare fuel assemblies. Dry transfer systems for UNF canisters are currently available and in use for transferring loaded canisters between the drying station and storage and transportation casks.

  16. Nuclear equivalence, nuclear transfer, and the cell cycle.

    PubMed

    Campbell, K H

    1999-01-01

    The last 20 years have seen the development of techniques for the production of mammals by nuclear transfer. Originally limited to the swapping of pronuclei and the use of early cleavage-stage embryos as nuclear donors, nuclear transfer came of age in 1995 with the birth of 2 Welsh Mountain lambs, Megan and Morag, that were produced using cultured differentiated cells as donors of genetic material. In 1996, Dolly was the first animal to be produced using the genetic material from an adult-derived somatic cell. The techniques used in the production of these animals have now been reproduced in both sheep and cattle, and as predicted, successful development has been obtained using donor cells taken directly ex vivo. This article reviews the current status of mammalian nuclear transfer and the biological background to these successes. PMID:16218826

  17. Interspecies Transfer of the Penicillin-Binding Protein 3-Encoding Gene ftsI between Haemophilus influenzae and Haemophilus haemolyticus Can Confer Reduced Susceptibility to β-Lactam Antimicrobial Agents

    PubMed Central

    Søndergaard, Annette; Witherden, Elizabeth A.

    2015-01-01

    Mutations in ftsI, encoding penicillin-binding protein 3, can cause decreased β-lactam susceptibility in Haemophilus influenzae. Sequencing of ftsI from clinical strains has indicated interspecies recombination of ftsI between H. influenzae and Haemophilus haemolyticus. This study documented apparently unrestricted homologous recombination of ftsI between H. influenzae and H. haemolyticus in vitro. Transfer of ftsI from resistant isolates conferred similar but not identical increases in the MICs of susceptible strains of H. influenzae and H. haemolyticus. PMID:25918135

  18. 10 CFR 70.42 - Transfer of special nuclear material.

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... 10 Energy 2 2012-01-01 2012-01-01 false Transfer of special nuclear material. 70.42 Section 70.42 Energy NUCLEAR REGULATORY COMMISSION (CONTINUED) DOMESTIC LICENSING OF SPECIAL NUCLEAR MATERIAL Acquisition, Use and Transfer of Special Nuclear Material, Creditors' Rights § 70.42 Transfer of...

  19. 10 CFR 70.42 - Transfer of special nuclear material.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... 10 Energy 2 2014-01-01 2014-01-01 false Transfer of special nuclear material. 70.42 Section 70.42 Energy NUCLEAR REGULATORY COMMISSION (CONTINUED) DOMESTIC LICENSING OF SPECIAL NUCLEAR MATERIAL Acquisition, Use and Transfer of Special Nuclear Material, Creditors' Rights § 70.42 Transfer of...

  20. 10 CFR 70.42 - Transfer of special nuclear material.

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ... 10 Energy 2 2013-01-01 2013-01-01 false Transfer of special nuclear material. 70.42 Section 70.42 Energy NUCLEAR REGULATORY COMMISSION (CONTINUED) DOMESTIC LICENSING OF SPECIAL NUCLEAR MATERIAL Acquisition, Use and Transfer of Special Nuclear Material, Creditors' Rights § 70.42 Transfer of...

  1. 10 CFR 70.42 - Transfer of special nuclear material.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... 10 Energy 2 2010-01-01 2010-01-01 false Transfer of special nuclear material. 70.42 Section 70.42 Energy NUCLEAR REGULATORY COMMISSION (CONTINUED) DOMESTIC LICENSING OF SPECIAL NUCLEAR MATERIAL Acquisition, Use and Transfer of Special Nuclear Material, Creditors' Rights § 70.42 Transfer of...

  2. 10 CFR 70.42 - Transfer of special nuclear material.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... 10 Energy 2 2011-01-01 2011-01-01 false Transfer of special nuclear material. 70.42 Section 70.42 Energy NUCLEAR REGULATORY COMMISSION (CONTINUED) DOMESTIC LICENSING OF SPECIAL NUCLEAR MATERIAL Acquisition, Use and Transfer of Special Nuclear Material, Creditors' Rights § 70.42 Transfer of...

  3. Heat transfer in a nuclear rocket engine

    SciTech Connect

    Konyukhov, G.V.; Petrov, A.I.

    1995-02-01

    Special features of heat transfer in the reactor of a nuclear rocket engine (NRE) are dealt with. It is shown that the design of the cooling system of the NRE reactor is governed by its stability to small deviations of the parameters from the corresponding calculated values and the possibility of compensating for effects due to nonuniformities and distrubances of various types and scales.

  4. Nuclear transfer and transgenesis in the pig.

    PubMed

    Kurome, Mayuko; Kessler, Barbara; Wuensch, Annegret; Nagashima, Hiroshi; Wolf, Eckhard

    2015-01-01

    Somatic cell nuclear transfer (SCNT) using genetically modified donor cells facilitates the generation of tailored pig models for biomedical research and for xenotransplantation. Up to now, SCNT is the main way to generate gene-targeted pigs, since germ line-competent pluripotent stem cells are not available for this species. In this chapter, we introduce our routine workflow for the production of genetically engineered pigs, especially focused on the genetic modification of somatic donor cells, SCNT using in vitro matured oocytes, and laparoscopic embryo transfer.

  5. Nuclear transfer and transgenesis in the pig.

    PubMed

    Kurome, Mayuko; Kessler, Barbara; Wuensch, Annegret; Nagashima, Hiroshi; Wolf, Eckhard

    2015-01-01

    Somatic cell nuclear transfer (SCNT) using genetically modified donor cells facilitates the generation of tailored pig models for biomedical research and for xenotransplantation. Up to now, SCNT is the main way to generate gene-targeted pigs, since germ line-competent pluripotent stem cells are not available for this species. In this chapter, we introduce our routine workflow for the production of genetically engineered pigs, especially focused on the genetic modification of somatic donor cells, SCNT using in vitro matured oocytes, and laparoscopic embryo transfer. PMID:25287337

  6. Sheep: The First Large Animal Model in Nuclear Transfer Research

    PubMed Central

    Czernik, Marta; Zacchini, Federica; Iuso, Domenico; Scapolo, Pier Augusto

    2013-01-01

    Abstract The scope of this article is not to provide an exhaustive review of nuclear transfer research, because many authoritative reviews exist on the biological issues related to somatic and embryonic cell nuclear transfer. We shall instead provide an overview on the work done specifically on sheep and the value of this work on the greater nuclear transfer landscape. PMID:24033140

  7. Culture, characteristics and chromosome complement of Siberian tiger fibroblasts for nuclear transfer.

    PubMed

    Song, Jimei; Hua, Song; Song, Kai; Zhang, Yong

    2007-01-01

    Tiger (Panthera tigris Linnaeus, 1758) is a characteristic species of Asia, which is in severe danger. Siberian tiger (Panthera tigris altaica) is the largest one of the five existent tiger subspecies. It is extremely endangered. One new way for tiger protection and rescue is to study interspecies cloning. But there is few research data about Siberian tiger. In this study, we cultured Siberian tiger fibroblasts in vitro, analyzed their biological characteristics, chromosomes, and cell cycles, to provide not only nuclear donors with good morphology, normal biological characteristics, and chromosome quantity for tiger interspecies cloning, but also reliable data for further studying Siberian tiger. The results indicated that Siberian tiger ear fibroblasts can be successfully obtained by tissue culture either with or without overnight cold digestion, the cultured cells were typical fibroblasts with normal morphology, growth curve, and chromosome quantity; G0/G1 percentage increased and S percentage decreased with the confluence of cells. G0/G1 and S stage rate was significantly different between 40-50% and 80-90%, 95-100% confluence; there is no distinct difference between 80-90% and 95-100% confluence. The cells at the same density (80-90% confluence) were treated with or without 0.5% serum starving, GO/G1 rate of the former was higher than the latter, but the difference was not significant. GO/G1 proportion of 95-100% confluence was slightly higher than serum starving (80-90% confluence), but no significant difference. Therefore, the Siberian tiger fibroblasts we cultured in vitro can be used as donor cells, and the donor cells do not need to be treated with normal serum starvation during nuclear transfer; if we will just consider the rate of the G0/G1 stage cells, serum starvation can be replaced by confluence inhibition when cultured cells were more than 80-90% confluence.

  8. 48 CFR 970.4402-4 - Nuclear material transfers.

    Code of Federal Regulations, 2010 CFR

    2010-10-01

    ... 48 Federal Acquisition Regulations System 5 2010-10-01 2010-10-01 false Nuclear material transfers... 970.4402-4 Nuclear material transfers. (a) Management and operating contractors, in preparing... nuclear material, shall be required to assure that each such subcontract or agreement contains a—...

  9. 48 CFR 970.4402-4 - Nuclear material transfers.

    Code of Federal Regulations, 2013 CFR

    2013-10-01

    ... 48 Federal Acquisition Regulations System 5 2013-10-01 2013-10-01 false Nuclear material transfers... 970.4402-4 Nuclear material transfers. (a) Management and operating contractors, in preparing... nuclear material, shall be required to assure that each such subcontract or agreement contains a—...

  10. 48 CFR 970.4402-4 - Nuclear material transfers.

    Code of Federal Regulations, 2014 CFR

    2014-10-01

    ... 48 Federal Acquisition Regulations System 5 2014-10-01 2014-10-01 false Nuclear material transfers... 970.4402-4 Nuclear material transfers. (a) Management and operating contractors, in preparing... nuclear material, shall be required to assure that each such subcontract or agreement contains a—...

  11. 48 CFR 970.4402-4 - Nuclear material transfers.

    Code of Federal Regulations, 2011 CFR

    2011-10-01

    ... 48 Federal Acquisition Regulations System 5 2011-10-01 2011-10-01 false Nuclear material transfers... 970.4402-4 Nuclear material transfers. (a) Management and operating contractors, in preparing... nuclear material, shall be required to assure that each such subcontract or agreement contains a—...

  12. 48 CFR 970.4402-4 - Nuclear material transfers.

    Code of Federal Regulations, 2012 CFR

    2012-10-01

    ... 48 Federal Acquisition Regulations System 5 2012-10-01 2012-10-01 false Nuclear material transfers... 970.4402-4 Nuclear material transfers. (a) Management and operating contractors, in preparing... nuclear material, shall be required to assure that each such subcontract or agreement contains a—...

  13. Bovine ooplasm partially remodels primate somatic nuclei following somatic cell nuclear transfer.

    PubMed

    Wang, Kai; Beyhan, Zeki; Rodriguez, Ramon M; Ross, Pablo J; Iager, Amy E; Kaiser, German G; Chen, Ying; Cibelli, Jose B

    2009-03-01

    Interspecies somatic cell nuclear transfer (iSCNT) has the potential to become a useful tool to address basic questions about the nucleus-cytoplasm interactions between species. It has also been proposed as an alternative for the preservation of endangered species and to derive autologous embryonic stem cells. Using chimpanzee/ bovine iSCNT as our experimental model we studied the early epigenetic events that take place soon after cell fusion until embryonic genome activation (EGA). Our analysis suggested partial EGA in iSCNT embryos at the eight-cell stage, as indicated by Br-UTP incorporation and expression of chimpanzee embryonic genes. Oct4, Stella, Crabp1, CCNE2, CXCL6, PTGER4, H2AFZ, c-MYC, KLF4, and GAPDH transcripts were expressed, while Nanog, Glut1, DSC2, USF2, Adrbk1, and Lin28 failed to be activated. Although development of iSCNT embryos did not progress beyond the 8- to 16-cell stage, chromatin remodeling events, monitored by H3K27 methylation, H4K5 acetylation, and global DNA methylation, were similar in both intra- and interspecies SCNT embryos. However, bisulfite sequencing indicated incomplete demethylation of Oct4 and Nanog promoters in eight-cell iSCNT embryos. ATP production levels were significantly higher in bovine SCNT embryos than in iSCNT embryos, TUNEL assays did not reveal any difference in the apoptotic status of the nuclei from both types of embryos. Collectively, our results suggest that bovine ooplasm can partially remodel chimpanzee somatic nuclei, and provides insight into some of the current barriers iSCNT must overcome if further embryonic development is to be expected. PMID:19196039

  14. Nuclear transfer technology in mammalian cloning.

    PubMed

    Wolf, D P; Mitalipov, S; Norgren, R B

    2001-01-01

    The past several years have witnessed remarkable progress in mammalian cloning using nuclear transfer (NT). Until 1997 and the announcement of the successful cloning of sheep from adult mammary gland or fetal fibroblast cells, our working assumption was that cloning by NT could only be accomplished with relatively undifferentiated embryonic cells. Indeed, live offspring were first produced by NT over 15 years ago from totipotent, embryonic blastomeres derived from early cleavage-stage embryos. However, once begun, the progression to somatic cell cloning or NT employing differentiated cells as the source of donor nuclei was meteoric, initially involving differentiated embryonic cell cultures in sheep in 1996 and quickly thereafter, fetal or adult somatic cells in sheep, cow, mouse, goat, and pig. Several recent reviews provide a background for and discussion of these successes. Here we will focus on the potential uses of reproductive cloning along with recent activities in the field and a discussion concerning current interests in human reproductive and therapeutic cloning.

  15. Passive heat transfer means for nuclear reactors

    DOEpatents

    Burelbach, James P.

    1984-01-01

    An improved passive cooling arrangement is disclosed for maintaining adjacent or related components of a nuclear reactor within specified temperature differences. Specifically, heat pipes are operatively interposed between the components, with the vaporizing section of the heat pipe proximate the hot component operable to cool it and the primary condensing section of the heat pipe proximate the other and cooler component operable to heat it. Each heat pipe further has a secondary condensing section that is located outwardly beyond the reactor confinement and in a secondary heat sink, such as air ambient the containment, that is cooler than the other reactor component. Means such as shrouding normally isolated the secondary condensing section from effective heat transfer with the heat sink, but a sensor responds to overheat conditions of the reactor to open the shrouding, which thereby increases the cooling capacity of the heat pipe. By having many such heat pipes, an emergency passive cooling system is defined that is operative without electrical power.

  16. Transgenic chicken, mice, cattle, and pig embryos by somatic cell nuclear transfer into pig oocytes.

    PubMed

    Gupta, Mukesh Kumar; Das, Ziban Chandra; Heo, Young Tae; Joo, Jin Young; Chung, Hak-Jae; Song, Hyuk; Kim, Jae-Hwan; Kim, Nam-Hyung; Lee, Hoon Taek; Ko, Dae Hwan; Uhm, Sang Jun

    2013-08-01

    This study explored the possibility of producing transgenic cloned embryos by interspecies somatic cell nuclear transfer (iSCNT) of cattle, mice, and chicken donor cells into enucleated pig oocytes. Enhanced green florescent protein (EGFP)-expressing donor cells were used for the nuclear transfer. Results showed that the occurrence of first cleavage did not differ significantly when pig, cattle, mice, or chicken cells were used as donor nuclei (p>0.05). However, the rate of blastocyst formation was significantly higher in pig (14.9±2.1%; p<0.05) SCNT embryos than in cattle (6.3±2.5%), mice (4.2±1.4%), or chicken (5.1±2.4%) iSCNT embryos. The iSCNT embryos also contained a significantly less number of cells per blastocyst than those of SCNT pig embryos (p<0.05). All (100%) iSCNT embryos expressed the EGFP gene, as evidenced by the green florescence under ultraviolet (UV) illumination. Microinjection of purified mitochondria from cattle somatic cells into pig oocytes did not have any adverse effect on their postfertilization in vitro development and embryo quality (p>0.05). Moreover, NCSU23 medium, which was designed for in vitro culture of pig embryos, was able to support the in vitro development of cattle, mice, and chicken iSCNT embryos up to the blastocyst stage. Taken together, these data suggest that enucleated pig oocytes may be used as a universal cytoplast for production of transgenic cattle, mice, and chicken embryos by iSCNT. Furthermore, xenogenic transfer of mitochondria to the recipient cytoplast may not be the cause for poor embryonic development of cattle-pig iSCNT embryos.

  17. Somatic Cell Nuclear Transfer in the Mouse

    NASA Astrophysics Data System (ADS)

    Kishigami, Satoshi; Wakayama, Teruhiko

    Somatic cell nuclear transfer (SCNT) has become a unique and powerful tool for epigenetic reprogramming research and gene manipulation in animals since “Dolly,” the first animal cloned from an adult cell was reported in 1997. Although the success rates of somatic cloning have been inefficient and the mechanism of reprogramming is still largely unknown, this technique has been proven to work in more than 10 mammalian species. Among them, the mouse provides the best model for both basic and applied research of somatic cloning because of its abounding genetic resources, rapid sexual maturity and propagation, minimal requirements for housing, etc. This chapter describes a basic protocol for mouse cloning using cumulus cells, the most popular cell type for NT, in which donor nuclei are directly injected into the oocyte using a piezo-actuated micromanipulator. In particular, we focus on a new, more efficient mouse cloning protocol using trichostatin A (TSA), a histone deacetylase (HDAC) inhibitor, which increases both in vitro and in vivo developmental rates from twofold to fivefold. This new method including TSA will be helpful to establish mouse cloning in many laboratories.

  18. Interspecies Dissemination of a Mobilizable Plasmid Harboring blaIMP-19 and the Possibility of Horizontal Gene Transfer in a Single Patient.

    PubMed

    Yamamoto, Masaki; Matsumura, Yasufumi; Gomi, Ryota; Matsuda, Tomonari; Tanaka, Michio; Nagao, Miki; Takakura, Shunji; Uemoto, Shinji; Ichiyama, Satoshi

    2016-09-01

    Carbapenemase-producing Gram-negative bacilli have been a global concern over the past 2 decades because these organisms can cause severe infections with high mortality rates. Carbapenemase genes are often carried by mobile genetic elements, and resistance plasmids can be transferred through conjugation. We conducted whole-genome sequencing (WGS) to demonstrate that the same plasmid harboring a metallo-β-lactamase gene was detected in two different species isolated from a single patient. Metallo-β-lactamase-producing Achromobacter xylosoxidans (KUN4507), non-metallo-β-lactamase-producing Klebsiella pneumoniae (KUN4843), and metallo-β-lactamase-producing K. pneumoniae (KUN5033) were sequentially isolated from a single patient and then analyzed in this study. Antimicrobial susceptibility testing, molecular typing (pulsed-field gel electrophoresis and multilocus sequence typing), and conjugation analyses were performed by conventional methods. Phylogenetic and molecular clock analysis of K. pneumoniae isolates were performed with WGS, and the nucleotide sequences of plasmids detected from these isolates were determined using WGS. Conventional molecular typing revealed that KUN4843 and KUN5033 were identical, whereas the phylogenetic tree analysis revealed a slight difference. These two isolates were separated from the most recent common ancestor 0.74 years before they were isolated. The same resistance plasmid harboring blaIMP-19 was detected in metallo-β-lactamase-producing A. xylosoxidans and K. pneumoniae Although this plasmid was not self-transferable, the conjugation of this plasmid from A. xylosoxidans to non-metallo-β-lactamase-producing K. pneumoniae was successfully performed. The susceptibility patterns for metallo-β-lactamase-producing K. pneumoniae and the transconjugant were similar. These findings supported the possibility of the horizontal transfer of plasmid-borne blaIMP-19 from A. xylosoxidans to K. pneumoniae in a single patient. PMID:27381397

  19. Electronic and Nuclear Factors in Charge and Excitation Transfer

    SciTech Connect

    Piotr Piotrowiak

    2004-09-28

    We report the and/or state of several subprojects of our DOE sponsored research on Electronic and Nuclear Factors in Electron and Excitation Transfer: (1) Construction of an ultrafast Ti:sapphire amplifier. (2) Mediation of electronic interactions in host-guest molecules. (3) Theoretical models of electrolytes in weakly polar media. (4) Symmetry effects in intramolecular excitation transfer.

  20. Handmade cloning: the future way of nuclear transfer?

    PubMed

    Vajta, Gábor

    2007-06-01

    The topic of this review is an alternative technique for somatic cell nuclear transfer. Removal of the zona pellucida facilitates manipulations of mammalian oocytes and early embryos, and problems related to their subsequent culture are commonly overestimated. This approach enables radical modifications to somatic cell nuclear transfer, and the handmade cloning (HMC) technique is now successfully applied to an increasing numbers of species. HMC radically decreases costs and the need for a skilled workforce; furthermore, it increases productivity, enables cryopreservation, and results in birth rates comparable, or even higher, than those achievable by micromanipulation-based traditional cloning (TC). The new technique can accelerate technology transfer and standardization and, eventually, might contribute to the widespread application of cloning. Additionally, HMC offers unique possibilities for the automation of somatic cell nuclear transfer.

  1. Cloning animals by somatic cell nuclear transfer--biological factors.

    PubMed

    Tian, X Cindy; Kubota, Chikara; Enright, Brian; Yang, Xiangzhong

    2003-11-13

    Cloning by nuclear transfer using mammalian somatic cells has enormous potential application. However, somatic cloning has been inefficient in all species in which live clones have been produced. High abortion and fetal mortality rates are commonly observed. These developmental defects have been attributed to incomplete reprogramming of the somatic nuclei by the cloning process. Various strategies have been used to improve the efficiency of nuclear transfer, however, significant breakthroughs are yet to happen. In this review we will discuss studies conducted, in our laboratories and those of others, to gain a better understanding of nuclear reprogramming. Because cattle are a species widely used for nuclear transfer studies, and more laboratories have succeeded in cloning cattle than any other species, this review will be focused on somatic cell cloning of cattle.

  2. Effects of A and B Wolbachia and host genotype on interspecies cytoplasmic incompatibility in Nasonia.

    PubMed

    Bordenstein, S R; Werren, J H

    1998-04-01

    Wolbachia endosymbionts cause postmating reproductive isolation between the sibling species Nasonia vitripennis and N. giraulti. Most Nasonia are doubly infected with a representative from each of the two major Wolbachia groups (A and B). This study investigates the role of single (A or B) and double (A and B) Wolbachia infections in interspecies cytoplasmic incompatibility (CI) and host genomic influences on the incompatibility phenotype. Results show that the single A Wolbachia harbored in N. vitripennis (wAv) is bidirectionally incompatible with the single A Wolbachia harbored in N. giraulti (wAg). Results also indirectly show that the N. vitripennis wBv is bidirectionally incompatible with the N. giraulti wBg. The findings support current phylogenetic evidence that suggests these single infections have independent origins and were acquired via horizontal transfer. The wAv Wolbachia expresses partial CI in the N. vitripennis nuclear background. However, following genomic replacement by introgression, wAv expresses complete CI in the N. giraulti background and remains bidirectionally incompatible with wAg. Results show that double infections can reinforce interspecies reproductive isolation through the addition of incompatibility types and indicate that the host genome can influence incompatibility levels. This study has implications for host-symbiont coevolution and the role of Wolbachia in speciation.

  3. Fibroblast cell line establishment, cryopreservation and interspecies embryos reconstruction in red panda ( Ailurus fulgens).

    PubMed

    Tao, Yong; Liu, Jianming; Zhang, Yunhai; Zhang, Meiling; Fang, Junshun; Han, Wei; Zhang, Zhizhong; Liu, Ya; Ding, Jianping; Zhang, Xiaorong

    2009-05-01

    In evolution, the red panda (Ailurus fulgens) plays a pivotal role in the higher level phylogeny of arctoides carnivore mammals. The red panda inhabits certain Asian countries only and its numbers are decreasing. Therefore, the development of feasible ways to preserve this species is necessary. Genetic resource cryopreservation and somatic cell nuclear transfer (SCNT) have been used extensively to rescue this endangered species. The present study describes the establishment, for the first time, of a red panda ear fibroblast cell line, which was then cryopreserved, thawed and cultured. Through micromanipulation, interspecies embryos were reconstructed using the cryopreserved-thawed fibroblasts of the red panda as the donor and rabbit oocytes as recipients. A total of 194 enucleated rabbit oocytes were reconstructed with red panda ear fibroblasts; enucleated oocytes were activated without fusion as the control. The results show that the fibroblast cell line was established successfully by tissue culture and then cryopreserved in liquid nitrogen. Supplementation with 20% fetal bovine serum and 8% dimethyl sulphoxide in basic medium facilitated the cryopreservation. The interspecies embryos were successfully reconstructed. The cleavage, morulae and blastocyst rates after in vitro culture were 71, 47 and 23% (31/194), respectively. This study indicated that a somatic cell line could be established and cryopreserved from red panda and that rabbit cytoplast supports mitotic cleavage of the red panda karyoplasts and is capable of reprogramming the nucleus to achieve blastocysts.

  4. Cloning in cattle: from embryo splitting to somatic nuclear transfer.

    PubMed

    Heyman, Y; Vignon, X; Chesné, P; Le Bourhis, D; Marchal, J; Renard, J P

    1998-01-01

    The ability to obtain genetically identical offspring in cattle (clones) is useful for research and for potential applications to breeding schemes. Experimental possibilities for generating such animals have evolved considerably in the last two decades. Embryo splitting has become a relatively simple technique but is limited to twinning. Embryonic nuclear transfer has improved and is associated with sexing to generate sets of clones despite a great variability of results between parent embryos. The factors of progress are reviewed here. Recently, somatic cells used as a source of nuclei in bovine nuclear transfer has been demonstrated. Here we present the results of the developmental potential of nuclei from skin and muscle cells.

  5. Cloning by somatic cell nuclear transfer.

    PubMed

    Fulka, J; First, N L; Loi, P; Moor, R M

    1998-10-01

    The birth of the first cloned mammals, produced by the introduction of somatic cell nuclei into enucleated oocytes, was an impressive and surprising development. Although the ethical debate has been intense, the important scientific questions raised by this work have been inadequately discussed and are still unresolved. In this essay we address three questions about nuclear transplantation in the eggs of mice and domestic animals. First, why were the recent experiments on somatic cell cloning successful, when so many others have failed? Second, were these exceptional cases, or is somatic cloning now open to all? Third, what are the future possibilities for increasing the efficiency and wider applicability of the cloning process?

  6. Somatic cell nuclear transfer in horses.

    PubMed

    Galli, Cesare; Lagutina, Irina; Duchi, Roberto; Colleoni, Silvia; Lazzari, Giovanna

    2008-07-01

    The cloning of equids was achieved in 2003, several years after the birth of Dolly the sheep and also after the cloning of numerous other laboratory and farm animal species. The delay was because of the limited development in the horse of more classical-assisted reproductive techniques required for successful cloning, such as oocyte maturation and in vitro embryo production. When these technologies were developed, the application of cloning also became possible and cloned horse offspring were obtained. This review summarizes the main technical procedures that are required for cloning equids and the present status of this technique. The first step is competent oocyte maturation, this is followed by oocyte enucleation and reconstruction, using either zona-enclosed or zona-free oocytes, by efficient activation to allow high cleavage rates and finally by a suitable in vitro embryo culture technique. Cloning of the first equid, a mule, was achieved using an in vivo-matured oocytes and immediate transfer of the reconstructed embryo, i.e. at the one cell stage, to the recipient oviduct. In contrast, the first horse offspring was obtained using a complete in vitro procedure from oocyte maturation to embryo culture to the blastocyst stage, followed by non-surgical transfer. Later studies on equine cloning report high efficiency relative to that for other species. Cloned equid offspring reported to date appear to be normal and those that have reached puberty have been confirmed to be fertile. In summary, horse cloning is now a reproducible technique that offers the opportunity to preserve valuable genetics and notably to generate copies of castrated champions and therefore, offspring from those champions that would be impossible to obtain otherwise. PMID:18638143

  7. Nuclear Waste Cross Site Transfer Pump Operational Resonance Resolution

    SciTech Connect

    HAUCK, F.M.

    1999-12-01

    Two single-volute, multi-stage centrifugal pumps are installed at a nuclear waste transfer station operated by the Department of Energy in Hanford, WA. The two parallel 100% pumps are Variable Frequency Drive operated and designed to transport waste etc.

  8. Nuclear electric power for multimegawatt orbit transfer vehicles

    NASA Technical Reports Server (NTRS)

    Casagrande, R. D.

    1987-01-01

    Multimegawatt nuclear propulsion is an attractive option for orbit transfer vehicles. The masses of these platforms are expected to exceed the capability of a single launch from Earth necessitating assembly in space in a parking orbit. The OTV would transfer the platform from the parking orbit to the operational orbit and then return for the next mission. Electric propulsion is advantageous because of the high specific impulse achieved by the technology, 1000 to 5000 s and beyond, to reduce the propellant required. Nuclear power is attractive as the power system because of the weight savings over solar systems in the multimegawatt regime, and multimegawatts of power are required. A conceptual diagram is shown of an OTV with a command control module using electric thrusters powered from an SP-100 class nuclear reactor power system.

  9. Commercial applications of nuclear transfer cloning: three examples.

    PubMed

    Forsberg, Erik J

    2005-01-01

    Potential applications of cloning go well beyond the popularly envisioned replication of valuable animals. This is because targeted genetic modifications can be made in donor cells before nuclear transfer. Applications that are currently being pursued include therapeutic protein production in the milk and blood of transgenic cloned animals, the use of cells, tissues and organs from gene-modified animals for transplantation into humans and genetically modified livestock that produce healthier and safer products in an environmentally friendly manner. Commercial and social acceptance of one or more of these early cloning applications will lead to yet unimagined applications of nuclear transfer technology. The present paper summarises progress on three additional applications of nuclear transfer, namely the development of male livestock that produce single-sex sperm, the transfer of immune responses from animals to their clones to permit the production of unlimited supplies of unique polyclonal antibodies, and the generation of genetically modified animals that accurately mimic human diseases for the purpose of developing new therapies. However, the myriad applications of cloning will require appropriate safeguards to ensure safe, humane and responsible outcomes of the technology.

  10. Somatic cell nuclear transfer: Past, present and future perspectives.

    PubMed

    Campbell, K H S; Fisher, P; Chen, W C; Choi, I; Kelly, R D W; Lee, J-H; Xhu, J

    2007-09-01

    It is now over a decade since the birth, in 1996, of Dolly the first animal to be produced by nuclear transfer using an adult derived somatic cell as nuclear donor. Since this time similar techniques have been successfully applied to a range of species producing live offspring and allowing the development of transgenic technologies for agricultural, biotechnological and medical uses. However, though applicable to a range of species, overall, the efficiencies of development of healthy offspring remain low. The low frequency of successful development has been attributed to incomplete or inappropriate reprogramming of the transferred nuclear genome. Many studies have demonstrated that such reprogramming occurs by epigenetic mechanisms not involving alterations in DNA sequence, however, at present the molecular mechanisms underlying reprogramming are poorly defined. Since the birth of Dolly many studies have attempted to improve the frequency of development, this review will discuss the process of animal production by nuclear transfer and in particular changes in the methodology which have increased development and survival, simplified or increased robustness of the technique. Although much of the discussion is applicable across species, for simplicity we will concentrate primarily on published data for cattle, sheep, pigs and mice. PMID:17610946

  11. Nuclear Effects in Neutrino Interactions at Low Momentum Transfer

    SciTech Connect

    Miltenberger, Ethan Ryan

    2015-05-01

    This is a study to identify predicted effects of the carbon nucleus environment on neutrino - nucleus interactions with low momentum transfer. A large sample of neutrino interaction data collected by the MINERvA experiment is analyzed to show the distribution of charged hadron energy in a region with low momentum transfer. These distributions reveal a major discrepancy between the data and a popular interaction model with only the simplest Fermi gas nuclear effects. Detailed analysis of systematic uncertainties due to energy scale and resolution can account for only a little of the discrepancy. Two additional nuclear model effects, a suppression/screening effect (RPA), and the addition of a meson exchange current process (MEC), are shown to improve the description of the data.

  12. The use of nuclear transfer to produce transgenic pigs.

    PubMed

    Macháty, Zoltán; Bondioli, Kenneth R; Ramsoondar, Jagdeece J; Fodor, William L

    2002-01-01

    Manipulation of the pig genome has the potential to improve pig production and offers powerful biomedical applications. Genetic manipulation of mammals has been possible for over two decades, but the technology available has proven both difficult and inefficient. The development of new techniques to enhance efficiency and overcome the complications of random insertion is of importance. Nuclear transfer combined with homologous recombination provides a possible solution: precise genetic modifications in the pig genome may be induced via homologous recombination, and viable offspring can be produced by nuclear transfer using cultured transfected cell lines. The technique is still ineffective, but it is believed to have immense potential. One area that would benefit from the technology is that of xenotransplantation: transgenic pigs are expected to be available as organ donors in the foreseeable future. PMID:12006153

  13. Intermediate filaments promote nuclear mechanical constraints during somatic cell nuclear transfer in the mouse.

    PubMed

    Gall, Laurence; Brochard, Vincent; Ruffini, Sylvie; Laffont, Ludivine; Fleurot, Renaud; Lavin, Tiphaine Aguirre; Jouneau, Alice; Beaujean, Nathalie

    2012-12-01

    The somatic cell nuclear transfer (SCNT) procedure requires nuclear remodeling to return differentiated somatic nuclei to the totipotent undifferentiated stage. We hypothesize that mechanical constraints might occur upon SCNT and thereby affect nuclear remodeling. Therefore, we analyzed the nuclear structures upon SCNT using as donors either wild-type fibroblasts with a dense vimentin network or vimentin-deprived cells [embryonic stem cells (ESCs) and fibroblasts invalidated for vimetin]. We demonstrated that following nuclear transfer of wild-type fibroblasts, vimentin intermediate filaments (IFs) persisted around the transplanted nuclei and 88% of them presented severe distortions. We also showed that the presence of vimentin filaments in the reconstructed embryos was correlated with DNA damage, as evidenced by γH2A.X foci. On the other hand, when ESCs or vimentin-null (Vim(-/-)) fibroblasts devoid of IFs were used as nuclear donors, no nuclear distortion and less DNA damage were observed. Altogether we believe that the introduction of vimentin into recipient oocytes during SCNT induces a mechanical constraint on the transplanted nucleus that is responsible for nuclear distortions and DNA damage. This could lead to incomplete reprogramming that would be detrimental to further embryonic development.

  14. Nuclear reactor fuel element having improved heat transfer

    DOEpatents

    Garnier, J.E.; Begej, S.; Williford, R.E.; Christensen, J.A.

    1982-03-03

    A nuclear reactor fuel element having improved heat transfer between fuel material and cladding is described. The element consists of an outer cladding tube divided into an upper fuel section containing a central core of fissionable or mixed fissionable and fertile fuel material, slightly smaller in diameter than the inner surface of the cladding tube and a small lower accumulator section, the cladding tube being which is filled with a low molecular weight gas to transfer heat from fuel material to cladding during irradiation. A plurality of essentially vertical grooves in the fuel section extend downward and communicate with the accumulator section. The radial depth of the grooves is sufficient to provide a thermal gradient between the hot fuel surface and the relatively cooler cladding surface to allow thermal segregation to take place between the low molecular weight heat transfer gas and high molecular weight fission product gases produced by the fuel material during irradiation.

  15. Nuclear reactor power for an electrically powered orbital transfer vehicle

    NASA Technical Reports Server (NTRS)

    Jaffe, L.; Beatty, R.; Bhandari, P.; Chow, E.; Deininger, W.; Ewell, R.; Fujita, T.; Grossman, M.; Kia, T.; Nesmith, B.

    1987-01-01

    To help determine the systems requirements for a 300-kWe space nuclear reactor power system, a mission and spacecraft have been examined which utilize electric propulsion and this nuclear reactor power for multiple transfers of cargo between low earth orbit (LEO) and geosynchronous earth orbit (GEO). A propulsion system employing ion thrusters and xenon propellant was selected. Propellant and thrusters are replaced after each sortie to GEO. The mass of the Orbital Transfer Vehicle (OTV), empty and dry, is 11,000 kg; nominal propellant load is 5000 kg. The OTV operates between a circular orbit at 925 km altitude, 28.5 deg inclination, and GEO. Cargo is brought to the OTV by Shuttle and an Orbital Maneuvering Vehicle (OMV); the OTV then takes it to GEO. The OTV can also bring cargo back from GEO, for transfer by OMV to the Shuttle. OTV propellant is resupplied and the ion thrusters are replaced by the OMV before each trip to GEO. At the end of mission life, the OTV's electric propulsion is used to place it in a heliocentric orbit so that the reactor will not return to earth. The nominal cargo capability to GEO is 6000 kg with a transit time of 120 days; 1350 kg can be transferred in 90 days, and 14,300 kg in 240 days. These capabilities can be considerably increased by using separate Shuttle launches to bring up propellant and cargo, or by changing to mercury propellant.

  16. Nuclear reactor power for an electrically powered orbital transfer vehicle

    SciTech Connect

    Jaffe, L.; Beatty, R.; Bhandari, P.; Chow, E.; Deininger, W.; Ewell, R.; Fujita, T.; Grossman, M.; Kia, T.; Nesmith, B.

    1987-05-01

    To help determine the systems requirements for a 300-kWe space nuclear reactor power system, a mission and spacecraft have been examined which utilize electric propulsion and this nuclear reactor power for multiple transfers of cargo between low earth orbit (LEO) and geosynchronous earth orbit (GEO). A propulsion system employing ion thrusters and xenon propellant was selected. Propellant and thrusters are replaced after each sortie to GEO. The mass of the Orbital Transfer Vehicle (OTV), empty and dry, is 11,000 kg; nominal propellant load is 5000 kg. The OTV operates between a circular orbit at 925 km altitude, 28.5 deg inclination, and GEO. Cargo is brought to the OTV by Shuttle and an Orbital Maneuvering Vehicle (OMV); the OTV then takes it to GEO. The OTV can also bring cargo back from GEO, for transfer by OMV to the Shuttle. OTV propellant is resupplied and the ion thrusters are replaced by the OMV before each trip to GEO. At the end of mission life, the OTV's electric propulsion is used to place it in a heliocentric orbit so that the reactor will not return to earth. The nominal cargo capability to GEO is 6000 kg with a transit time of 120 days; 1350 kg can be transferred in 90 days, and 14,300 kg in 240 days. These capabilities can be considerably increased by using separate Shuttle launches to bring up propellant and cargo, or by changing to mercury propellant.

  17. Design considerations for Mars transfer vehicles using nuclear thermal propulsion

    NASA Astrophysics Data System (ADS)

    Emrich, William J.

    1995-01-01

    The design of a Mars Transfer Vehicle (MTV) utilizing nuclear propulsion will require that careful consideration be given to the nuclear radiation environment in which it will operate. The extremely high neutron and gamma fluxes characteristic of nuclear thermal propulsion systems will cause significant heating of the fluid systems in close proximity to the reactor, especially in the lower propellant tanks. Crew radiation doses are also a concern particularly late in a mission when there is less shielding from the propellant tanks. In this study, various vehicle configuration and shielding strategies were examined and the resulting time dependent radiation fields evaluated. A common cluster of three particle bed reactor (PBR) engines were used in all configurations examined. In general, it appears that long, relatively narrow vehicles perform the best from a radiation standpoint, however, good shield optimization will be critical in maintaining a low radiation environment while minimizing the shield weight penalty.

  18. Technological Transfer from Research Nuclear Reactors to New Generation Nuclear Power Reactors

    NASA Astrophysics Data System (ADS)

    Radulescu, Laura; Pavelescu, Margarit

    2010-01-01

    The goal of this paper is the analysis of the technological transfer role in the nuclear field, with particular emphasis on nuclear reactors domain. The presentation is sustained by historical arguments. In this frame, it is very important to start with the achievements of the first nuclear systems, for instant those with natural uranium as fuel and heavy water as moderator, following in time through the history until the New Generation Nuclear Power Reactors. Starting with 1940, the accelerated development of the industry has implied the increase of the global demand for energy. In this respect, the nuclear energy could play an important role, being essentially an unlimited source of energy. However, the nuclear option faces the challenges of increasingly demanding safety requirements, economic competitiveness and public acceptance. Worldwide, a significant amount of experience has been accumulated during development, licensing, construction, and operation of nuclear power reactors. The experience gained is a strong basis for further improvements. Actually, the nuclear programs of many countries are addressing the development of advanced reactors, which are intended to have better economics, higher reliability, improved safety, and proliferation-resistant characteristics in order to overcome the current concerns about nuclear power. Advanced reactors, now under development, may help to meet the demand for energy power of both developed and developing countries as well as for district heating, desalination and for process heat. The paper gives historical examples that illustrate the steps pursued from first research nuclear reactors to present advanced power reactors. Emphasis was laid upon the fact that the progress is due to the great discoveries of the nuclear scientists using the technological transfer.

  19. Technological Transfer from Research Nuclear Reactors to New Generation Nuclear Power Reactors

    SciTech Connect

    Radulescu, Laura; Pavelescu, Margarit

    2010-01-21

    The goal of this paper is the analysis of the technological transfer role in the nuclear field, with particular emphasis on nuclear reactors domain. The presentation is sustained by historical arguments. In this frame, it is very important to start with the achievements of the first nuclear systems, for instant those with natural uranium as fuel and heavy water as moderator, following in time through the history until the New Generation Nuclear Power Reactors.Starting with 1940, the accelerated development of the industry has implied the increase of the global demand for energy. In this respect, the nuclear energy could play an important role, being essentially an unlimited source of energy. However, the nuclear option faces the challenges of increasingly demanding safety requirements, economic competitiveness and public acceptance. Worldwide, a significant amount of experience has been accumulated during development, licensing, construction, and operation of nuclear power reactors. The experience gained is a strong basis for further improvements. Actually, the nuclear programs of many countries are addressing the development of advanced reactors, which are intended to have better economics, higher reliability, improved safety, and proliferation-resistant characteristics in order to overcome the current concerns about nuclear power. Advanced reactors, now under development, may help to meet the demand for energy power of both developed and developing countries as well as for district heating, desalination and for process heat.The paper gives historical examples that illustrate the steps pursued from first research nuclear reactors to present advanced power reactors. Emphasis was laid upon the fact that the progress is due to the great discoveries of the nuclear scientists using the technological transfer.

  20. INTERSPECIES DOSIMETRY MODELS FOR PULMONARY PHARMACOLOGY

    EPA Science Inventory

    Interspecies Dosimetry Models for Pulmonary Pharmacology

    Ted B. Martonen, Jeffry D. Schroeter, and John S. Fleming

    Experimental Toxicology Division, National Health and Environmental Effects Research Laboratory, U.S. Environmental Protection Agency, Research Triangl...

  1. Heat transfer in nuclear fuels: Measurements of gap conductance

    NASA Astrophysics Data System (ADS)

    Cho, Chun Hyung

    Heat transfer in the fuel-clad gap in a nuclear reactor impacts the overall temperature distribution, stored energy and the mechanical properties of a nuclear fuel rod. Therefore, an accurate estimation of the gap conductance between the fuel and the clad is critically important for reactor design and operations. To obtain the requisite accuracy in the gap conductance estimation, it is important to understand the effects of the convective heat transfer coefficient, the gas composition, pressure and temperature, and so forth. The objectives of this study are to build a bench-scale experimental apparatus for the measurement of thermal gap conductances and to develop a better understanding of the differences that have been previously observed between such measured values and those predicted theoretically. This is accomplished by employing improved analyses of the experiments and improved theoretical models. Using laser heating of slightly separated stainless-steel plates, the gap conductance was measured using a technique that compares the theoretical and experimental time dependent temperatures at the back surface of the second plate. To consider the effects of surface temperature and gas pressure, the theoretical temperatures were calculated using a convective heat transfer coefficient that was dependent upon both the temperature and the gas pressure.

  2. Somatic cell nuclear transfer in mammals: progress and applications.

    PubMed

    Colman, A

    Somatic nuclear transfer has been performed with frogs since the early 1960s, yet it has proved impossible to generate an adult frog using an adult cell as nuclear donor. After some initial skepticism, the birth of sheep, cows, goats, and mice using this technique with fetal or adult cell donors is now established fact. The success with adult mammalian cell donors extends the historic work in frogs by attesting to the totipotency of nuclei in at least some adult, differentiated cell types. Because the technique offers a developmental read out of the totality of genetic and molecular lifetime changes accumulated by the nucleus of a single somatic cell, basic research applications are seen in the fields of ageing, cancer, X chromosome inactivation, and imprinting. The prospect of a method for gene targeting in livestock holds particular promise for commercial applications; whilst for humans, the use of nuclear transfer to provide diverse populations of customized stem cells for therapeutic purposes presents a tantalizing future goal.

  3. Method for somatic cell nuclear transfer in zebrafish.

    PubMed

    Siripattarapravat, K; Prukudom, S; Cibelli, J

    2016-01-01

    This chapter presents a detailed methodology for somatic cell nuclear transfer-cloning of zebrafish. We aim to place the reader in a virtual lab experience to assist acquisition of the technical skills required for reproducing the published protocol. All materials, including catalog numbers for reagents and techniques for their preparation, are provided. Our protocols describe laser inactivation of egg chromosomes, the transfer of a cell through the oocyte micropyle, and spontaneous activation of the reconstructed embryo. High-quality eggs are the key to cloning success, and Chinook salmon ovarian fluid is indispensable for keeping eggs arrested at the metaphase of meiosis II. This protocol continues to be refined by our laboratory. However, naive investigators should be able to apply it in its present form to generate cloned zebrafish. PMID:27443929

  4. Nuclear transfer: a new tool for reproductive biotechnology in cattle.

    PubMed

    Heyman, Yvan

    2005-01-01

    Recent evolutions of somatic cloning by nuclear transfer are reported, especially in the bovine species where potential applications are underway for biomedicine in association with transgenesis, or for agriculture by improving livestock. The overall efficiency of this biotechnology remains low in terms of viable offspring, but significant progress has been achieved on the different steps of the technique. However, the in vivo development of bovine blastocysts derived from somatic nuclear transfer is characterised by some important features that lead to the "cloning syndrome". Important losses occur during the peri-implantation period and further late foetal loss is observed in association with the Large Offspring Syndrome. About 60-70% of the cloned calves born survive normally to the adult stage and present an apparently normal physiology. Recent data already available on bovine somatic clones of both sexes indicate that they have a zootechnical performance similar to non cloned animals and they are able to reproduce normally without the pathologies associated to cloning thus confirming that the deviations observed in clones are of epigenetic origin and not transmitted to the progeny.

  5. Reshaping the Transcriptional Frontier: Epigenetics and Somatic Cell Nuclear Transfer

    PubMed Central

    LONG, CHARLES R.; WESTHUSIN, MARK E.; GOLDING, MICHAEL C.

    2014-01-01

    SUMMARY Somatic-cell nuclear transfer (SCNT) experiments have paved the way to the field of cellular reprogramming. The demonstrated ability to clone over 20 different species to date has proven that the technology is robust but very inefficient, and is prone to developmental anomalies. Yet, the offspring from cloned animals exhibit none of the abnormalities of their parents, suggesting the low efficiency and high developmental mortality are epigenetic in origin. The epigenetic barriers to reprogramming somatic cells into a totipotent embryo capable of developing into a viable offspring are significant and varied. Despite their intimate relationship, chromatin structure and transcription are often not uniformly reprogramed after nuclear transfer, and many cloned embryos develop gene expression profiles that are hybrids between the donor cell and an embryonic blastomere. Recent advances in cellular reprogramming suggest that alteration of donor-cell chromatin structure towards that found in an normal embryo is actually the rate-limiting step in successful development of SCNT embryos. Here we review the literature relevant to the transformation of a somatic-cell nucleus into an embryo capable of full-term development. Interestingly, while resetting somatic transcription and associated epigenetic marks are absolutely required for development of SCNT embryos, life does not demand perfection. PMID:24167064

  6. Production of Cloned Mice by Nuclear Transfer of Cumulus Cells

    PubMed Central

    Kurd, Soleiman; Zarei, Mohammad Ali; Fathi, Fardin; Ghadimi, Tayyeb; Hakhamaneshi, Mohammad Saeed; Jalili, Ali

    2013-01-01

    Background Over the past several years, mammals have been successfully cloned by either the splitting of an early stage embryo or nuclear transfer of adult somatic cells (NT) into oocytes. Although it has been 15 years since the generation of the first cloned mammals from somatic cells by NT, the success rate for producing live offspring by this technique is low regardless of the cell type and animal species used. However, these techniques have the potential to be important tools for future research in basic biology. In the present study, we described our experiences in producing successfully cloned mouse using NT method and piezo-actuated micromanipulator. Methods B6D2F1 mice, 8-12 weeks old, were superovulated with injections of 5 IU of pregnant mare serum gonadotropin and 5 IU of human chorionic gonadotropin administered 48 hr apart. Enucleation and donor nuclei cumulus cell injection were performed with a piezo-actuated micromanipulator after which activation and trichostatin A treatment were used for reconstructed oocytes. Two-cell stage cloned embryos that developed in the mWM medium were transferred into the oviducts of pseudopregnant NMRI mice. Results Of 367 oocytes collected, 131 (69%) developed into 2-cell stage embryos. Of these, 5 (1%) live pups were successfully delivered. We used NMRI foster mother to raise the pups by lactation. One adult cloned mouse was mated, after which she delivered and raised normal offspring. Conclusion For mouse cloning, the present study also successfully tested the capability of somatic cell nuclear transfer SCNT using a piezo unit. PMID:23919122

  7. The effect of the number of transferred embryos, the interval between nuclear transfer and embryo transfer, and the transfer pattern on pig cloning efficiency.

    PubMed

    Rim, Chol Ho; Fu, Zhixin; Bao, Lei; Chen, Haide; Zhang, Dan; Luo, Qiong; Ri, Hak Chol; Huang, Hefeng; Luan, Zhidong; Zhang, Yan; Cui, Chun; Xiao, Lei; Jong, Ui Myong

    2013-12-01

    To improve the efficiency of producing cloned pigs, we investigated the influence of the number of transferred embryos, the culturing interval between nuclear transfer (NT) and embryo transfer, and the transfer pattern (single oviduct or double oviduct) on cloning efficiency. The results demonstrated that transfer of either 150-200 or more than 200NT embryos compared to transfer of 100-150 embryos resulted in a significantly higher pregnancy rate (48 ± 16, 50 ± 16 vs. 29 ± 5%, p<0.05) and average litter size (4.1 ± 2.3, 7 ± 3.6 vs. 2.5 ± 0.5). In vitro culture of reconstructed embryos for a longer time (40 h vs. 20 h) resulted in higher (p<0.05) pregnancy rate (44 ± 9 vs. 31 ± 3%) and delivery rate (44 ± 9 vs. 25 ± 9%). Furthermore, double oviductal transfer dramatically increased pregnancy rate (83 ± 6 vs. 27+8%, p<0.05), delivery rate (75 ± 2 vs. 27+8%, p<0.05) and average litter size (6.5 ± 2.8 vs. 2.6 ± 1.2) compared to single oviductal transfer. Our study demonstrated that an improvement in pig cloning efficiency is achieved by adjusting the number and in vitro culture time of reconstructed embryos as well as the embryo transfer pattern.

  8. Factors affecting the development of somatic cell nuclear transfer embryos in Cattle.

    PubMed

    Akagi, Satoshi; Matsukawa, Kazutsugu; Takahashi, Seiya

    2014-01-01

    Nuclear transfer is a complex multistep procedure that includes oocyte maturation, cell cycle synchronization of donor cells, enucleation, cell fusion, oocyte activation and embryo culture. Therefore, many factors are believed to contribute to the success of embryo development following nuclear transfer. Numerous attempts to improve cloning efficiency have been conducted since the birth of the first sheep by somatic cell nuclear transfer. However, the efficiency of somatic cell cloning has remained low, and applications have been limited. In this review, we discuss some of the factors that affect the developmental ability of somatic cell nuclear transfer embryos in cattle.

  9. The developmental competence of bovine nuclear transfer embryos derived from cow versus heifer cytoplasts.

    PubMed

    Aston, Kenneth I; Li, Guang-Peng; Hicks, Brady A; Sessions, Benjamin R; Pate, Barry J; Hammon, Douglas S; Bunch, Thomas D; White, Kenneth L

    2006-10-01

    Due to its economic importance, the production of cattle by nuclear transfer has been a primary research focus for many researchers during the past few years. While many groups have successfully produced cattle by nuclear transfer, and progress in this area continues, nuclear transfer remains a very inefficient technology. This study evaluates the effect of the oocyte source (cow and heifer) on the developmental competence of nuclear transfer embryos. In order for nuclear transfer to be successful, a differentiated donor cell must be reprogrammed and restored to a totipotent state. This reprogramming is probably accomplished by factors within the oocyte cytoplasm. This study indicates that oocytes derived from cows have a greater capacity to reprogram donor cell DNA following nuclear transfer as compared to heifer oocytes based on in vitro development to the 2-cell stage and to the compacted morula/blastocyst stages. Nuclear transfer embryos derived from cow oocytes resulted in significantly higher rates of pregnancy establishment than embryos derived from heifer oocytes and resulted in higher pregnancy retention at 90 and 180 days and a greater number of term deliveries. Following delivery more calves derived from cow oocytes tended to be healthy and normal than those derived from heifer oocytes. The differences in developmental efficiency between nuclear transfer embryos derived from cow and heifer cytoplasts demonstrate that subtle differences in oocyte biology can have significant effects on subsequent development of nuclear transfer embryos. PMID:16324805

  10. Recent advancements in cloning by somatic cell nuclear transfer

    PubMed Central

    Ogura, Atsuo; Inoue, Kimiko; Wakayama, Teruhiko

    2013-01-01

    Somatic cell nuclear transfer (SCNT) cloning is the sole reproductive engineering technology that endows the somatic cell genome with totipotency. Since the first report on the birth of a cloned sheep from adult somatic cells in 1997, many technical improvements in SCNT have been made by using different epigenetic approaches, including enhancement of the levels of histone acetylation in the chromatin of the reconstructed embryos. Although it will take a considerable time before we fully understand the nature of genomic programming and totipotency, we may expect that somatic cell cloning technology will soon become broadly applicable to practical purposes, including medicine, pharmaceutical manufacturing and agriculture. Here we review recent progress in somatic cell cloning, with a special emphasis on epigenetic studies using the laboratory mouse as a model. PMID:23166393

  11. Cloning of ES cells and mice by nuclear transfer.

    PubMed

    Wakayama, Sayaka; Kishigami, Satoshi; Wakayama, Teruhiko

    2009-01-01

    We have been able to develop a stable nuclear transfer (NT) method in the mouse, in which donor nuclei are directly injected into the oocyte using a piezo-actuated micromanipulator. Although the piezo unit is a complex tool, once mastered it is of great help not only in NT experiments, but also in almost all other forms of micromanipulation. Using this technique, embryonic stem (ntES) cell lines established from somatic cell nuclei can be generated relatively easily from a variety of mouse genotypes and cell types. Such ntES cells can be used not only for experimental models of human therapeutic cloning but also as a means of preserving mouse genomes instead of preserving germ cells. Here, we describe our most recent protocols for mouse cloning.

  12. Recent advancements in cloning by somatic cell nuclear transfer.

    PubMed

    Ogura, Atsuo; Inoue, Kimiko; Wakayama, Teruhiko

    2013-01-01

    Somatic cell nuclear transfer (SCNT) cloning is the sole reproductive engineering technology that endows the somatic cell genome with totipotency. Since the first report on the birth of a cloned sheep from adult somatic cells in 1997, many technical improvements in SCNT have been made by using different epigenetic approaches, including enhancement of the levels of histone acetylation in the chromatin of the reconstructed embryos. Although it will take a considerable time before we fully understand the nature of genomic programming and totipotency, we may expect that somatic cell cloning technology will soon become broadly applicable to practical purposes, including medicine, pharmaceutical manufacturing and agriculture. Here we review recent progress in somatic cell cloning, with a special emphasis on epigenetic studies using the laboratory mouse as a model.

  13. A New, Dynamic Era for Somatic Cell Nuclear Transfer?

    PubMed

    Loi, Pasqualino; Iuso, Domenico; Czernik, Marta; Ogura, Atsuo

    2016-10-01

    Cloning animals by somatic cell nuclear transfer (SCNT) has remained an uncontrollable process for many years. High rates of embryonic losses, stillbirths, and postnatal mortality have been typical outcomes. These developmental problems arise from abnormal genomic reprogramming: the capacity of the oocyte to reset the differentiated memory of a somatic cell. However, effective reprogramming strategies are now available. These target the whole genome or single domains such as the Xist gene, and their effectiveness has been validated with the ability of experimental animals to develop to term. Thus, SCNT has become a controllable process that can be used to 'rescue' endangered species, and for biomedical research such as therapeutic cloning and the isolation of induced pluripotent stem cells (iPSCs). PMID:27118511

  14. Using somatic-cell nuclear transfer to study aging.

    PubMed

    Kishigami, Satoshi; Lee, Ah Reum; Wakayama, Teruhiko

    2013-01-01

    In mammals, a diploid genome following fertilization of haploid cells, an egg, and a spermatozoon is unique and irreproducible. This implies that the generated unique diploid genome is doomed with the individual's inevitable demise. Since it was first reported in 1997 that Dolly the sheep had been cloned, many mammalian species have been cloned successfully using somatic-cell nuclear transfer (SCNT). The success of SCNT in mammals enables us not only to reproduce offspring without germ cells, that is, to "passage" a unique diploid genome, but also to address valuable biological questions on development, nuclear reprogramming, and epigenetic memory. Successful cloning can also support epigenetic reprogramming where the aging clock is reset or reversed. Recent work using iPS cell technology has explored the practicality and led to the recapitulation of premature aging with iPSCs from progeroid laminopathies. As a result, reprogramming tools are also expected to contribute to studying biological age. However, the efficiency of animal cloning is still low in most cases and the mechanism of reprogramming in cloned embryos is still largely unclear. Here, based on recent advances, we describe an improved, more efficient mouse cloning protocol using histone deacetylase inhibitors (HDACis) and latrunculin A, which increases the success rates of producing cloned mice or establishing ES cells fivefold. This improved method of cloning will provide a strong tool to address many issues including biological aging more easily and with lower cost.

  15. Using somatic-cell nuclear transfer to study aging.

    PubMed

    Kishigami, Satoshi; Lee, Ah Reum; Wakayama, Teruhiko

    2013-01-01

    In mammals, a diploid genome following fertilization of haploid cells, an egg, and a spermatozoon is unique and irreproducible. This implies that the generated unique diploid genome is doomed with the individual's inevitable demise. Since it was first reported in 1997 that Dolly the sheep had been cloned, many mammalian species have been cloned successfully using somatic-cell nuclear transfer (SCNT). The success of SCNT in mammals enables us not only to reproduce offspring without germ cells, that is, to "passage" a unique diploid genome, but also to address valuable biological questions on development, nuclear reprogramming, and epigenetic memory. Successful cloning can also support epigenetic reprogramming where the aging clock is reset or reversed. Recent work using iPS cell technology has explored the practicality and led to the recapitulation of premature aging with iPSCs from progeroid laminopathies. As a result, reprogramming tools are also expected to contribute to studying biological age. However, the efficiency of animal cloning is still low in most cases and the mechanism of reprogramming in cloned embryos is still largely unclear. Here, based on recent advances, we describe an improved, more efficient mouse cloning protocol using histone deacetylase inhibitors (HDACis) and latrunculin A, which increases the success rates of producing cloned mice or establishing ES cells fivefold. This improved method of cloning will provide a strong tool to address many issues including biological aging more easily and with lower cost. PMID:23929101

  16. HEAT TRANSFER ANALYSIS FOR NUCLEAR WASTE SOLIDIFICATION CONTAINER

    SciTech Connect

    Lee, S.

    2009-06-01

    The Nuclear Nonproliferation Programs Design Authority is in the design stage of the Waste Solidification Building (WSB) for the treatment and solidification of the radioactive liquid waste streams generated by the Pit Disassembly and Conversion Facility (PDCF) and Mixed Oxide (MOX) Fuel Fabrication Facility (MFFF). The waste streams will be mixed with a cementitious dry mix in a 55-gallon waste container. Savannah River National Laboratory (SRNL) has been performing the testing and evaluations to support technical decisions for the WSB. Engineering Modeling & Simulation Group was requested to evaluate the thermal performance of the 55-gallon drum containing hydration heat source associated with the current baseline cement waste form. A transient axi-symmetric heat transfer model for the drum partially filled with waste form cement has been developed and heat transfer calculations performed for the baseline design configurations. For this case, 65 percent of the drum volume was assumed to be filled with the waste form, which has transient hydration heat source, as one of the baseline conditions. A series of modeling calculations has been performed using a computational heat transfer approach. The baseline modeling results show that the time to reach the maximum temperature of the 65 percent filled drum is about 32 hours when a 43 C initial cement temperature is assumed to be cooled by natural convection with 27 C external air. In addition, the results computed by the present model were compared with analytical solutions. The modeling results will be benchmarked against the prototypic test results. The verified model will be used for the evaluation of the thermal performance for the WSB drum. Detailed results and the cases considered in the calculations will be discussed here.

  17. Gnotobiotic Miniature Pig Interbreed Somatic Cell Nuclear Transfer for Xenotransplantation.

    PubMed

    Hwang, Jeong Ho; Kim, Sang Eun; Gupta, Mukesh Kumar; Lee, HoonTaek

    2016-08-01

    Transgenic animal producing technology has improved consistently over the last couple of decades. Among the available methods, somatic cell nuclear transfer (SCNT) technology was officially the most popular. However, SCNT has low efficiency and requires a highly skilled individual. Additionally, the allo-SCNT nuclear reprogramming mechanism is poorly understood in the gnotobiotic miniature pig, which is a candidate for xenotransplantation, making sampling in oocytes very difficult compared to commercial hybrid pigs. Therefore, interbreed SCNT (ibSCNT), which is a combination of miniature pig and commercial pig (Landrace based), was analyzed and was found to be similar to SCNT in terms of the rate of blastocyst formation (12.6% ± 2.9% vs. 15.5% ± 2.2%; p > 0.05). However, a significantly lower fusion rate was observed in the ibSCNT compared to normal SCNT with Landrace pig somatic cells (29.6% ± 0.8% vs. 65.0% ± 4.9%). Thus, the optimization of fusion parameters was necessary for efficient SCNT. Our results further revealed that ibSCNT by the whole-cell intracytoplasmic injection (WCICI) method had a significantly higher blastocyst forming efficiency than the electrofusion method (31.1 ± 8.5 vs. 15.5% ± 2.2%). The nuclear remodeling and the pattern of changes in acetylation at H3K9 residue were similar in both SCNT and ibSCNT embryos. PMID:27459580

  18. Cloned ferrets produced by somatic cell nuclear transfer

    PubMed Central

    Li, Ziyi; Sun, Xingshen; Chen, Juan; Liu, Xiaoming; Wisely, Samantha M.; Zhou, Qi; Renard, Jean-Paul; Leno, Gregory H.; Engelhardt, John F.

    2007-01-01

    Somatic cell nuclear transfer (SCNT) offers great potential for developing better animal models of human disease. The domestic ferret (Mustela putorius furo) is an ideal animal model for influenza infections and potentially other human respiratory diseases such as cystic fibrosis, where mouse models have failed to reproduce the human disease phenotype. Here, we report the successful production of live cloned, reproductively competent, ferrets using species-specific SCNT methodologies. Critical to developing a successful SCNT protocol for the ferret was the finding that hormonal treatment, normally used for superovulation, adversely affected the developmental potential of recipient oocytes. The onset of Oct4 expression was delayed and incomplete in parthenogenetically activated oocytes collected from hormone-treated females relative to oocytes collected from females naturally mated with vasectomized males. Stimulation induced by mating and in vitro oocyte maturation produced the optimal oocyte recipient for SCNT. Although nuclear injection and cell fusion produced mid-term fetuses at equivalent rates (~3–4%), only cell fusion gave rise to healthy surviving clones. Single cell fusion rates and the efficiency of SCNT were also enhanced by placing two somatic cells into the perivitelline space. These species-specific modifications facilitated the birth of live, healthy, and fertile cloned ferrets. The development of microsatellite genotyping for domestic ferrets confirmed that ferret clones were genetically derived from their respective somatic cells and unrelated to their surrogate mother. With this technology, it is now feasible to begin generating genetically defined ferrets for studying transmissible and inherited human lung diseases. Cloning of the domestic ferret may also aid in recovery and conservation of the endangered black-footed ferret and European mink. PMID:16584722

  19. 10 CFR 73.28 - Security background checks for secure transfer of nuclear materials.

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... 10 Energy 2 2012-01-01 2012-01-01 false Security background checks for secure transfer of nuclear materials. 73.28 Section 73.28 Energy NUCLEAR REGULATORY COMMISSION (CONTINUED) PHYSICAL PROTECTION OF PLANTS AND MATERIALS Physical Protection of Special Nuclear Material in Transit § 73.28...

  20. 10 CFR 73.28 - Security background checks for secure transfer of nuclear materials.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... 10 Energy 2 2011-01-01 2011-01-01 false Security background checks for secure transfer of nuclear materials. 73.28 Section 73.28 Energy NUCLEAR REGULATORY COMMISSION (CONTINUED) PHYSICAL PROTECTION OF PLANTS AND MATERIALS Physical Protection of Special Nuclear Material in Transit § 73.28...

  1. 10 CFR 73.28 - Security background checks for secure transfer of nuclear materials.

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ... 10 Energy 2 2013-01-01 2013-01-01 false Security background checks for secure transfer of nuclear materials. 73.28 Section 73.28 Energy NUCLEAR REGULATORY COMMISSION (CONTINUED) PHYSICAL PROTECTION OF PLANTS AND MATERIALS Physical Protection of Special Nuclear Material in Transit § 73.28...

  2. 10 CFR 73.28 - Security background checks for secure transfer of nuclear materials.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... 10 Energy 2 2010-01-01 2010-01-01 false Security background checks for secure transfer of nuclear materials. 73.28 Section 73.28 Energy NUCLEAR REGULATORY COMMISSION (CONTINUED) PHYSICAL PROTECTION OF PLANTS AND MATERIALS Physical Protection of Special Nuclear Material in Transit § 73.28...

  3. 10 CFR 73.28 - Security background checks for secure transfer of nuclear materials.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... 10 Energy 2 2014-01-01 2014-01-01 false Security background checks for secure transfer of nuclear materials. 73.28 Section 73.28 Energy NUCLEAR REGULATORY COMMISSION (CONTINUED) PHYSICAL PROTECTION OF PLANTS AND MATERIALS Physical Protection of Special Nuclear Material in Transit § 73.28...

  4. Somatic cell nuclear transfer cloning: practical applications and current legislation.

    PubMed

    Niemann, H; Lucas-Hahn, A

    2012-08-01

    Somatic cloning is emerging as a new biotechnology by which the opportunities arising from the advances in molecular genetics and genome analysis can be implemented in animal breeding. Significant improvements have been made in SCNT protocols in the past years which now allow to embarking on practical applications. The main areas of application of SCNT are: Reproductive cloning, therapeutic cloning and basic research. A great application potential of SCNT based cloning is the production of genetically modified (transgenic) animals. Somatic cell nuclear transfer based transgenic animal production has significant advances over the previously employed microinjection of foreign DNA into pronuclei of zygotes. This cell based transgenesis is compatible with gene targeting and allows both, the addition of a specific gene and the deletion of an endogenous gene. Efficient transgenic animal production provides numerous opportunities for agriculture and biomedicine. Regulatory agencies around the world have agreed that food derived from cloned animals and their offspring is safe and there is no scientific basis for questioning this. Commercial application of somatic cloning within the EU is via the Novel Food regulation EC No. 258/97. Somatic cloning raises novel questions regarding the ethical and moral status of animals and their welfare which has prompted a controversial discussion in Europe which has not yet been resolved.

  5. Developmental kinetics of bovine nuclear transfer and parthenogenetic embryos.

    PubMed

    Holm, P; Booth, P J; Callesen, H

    2003-01-01

    Early developmental kinetics of nuclear transfer (NT) embryos reconstituted with blastomeres and parthenogenones produced by ionophore activation followed by either dimethylaminopurine (DMAP) or cycloheximide (CHX) treatment was studied. In vitro produced (IVP) embryos served as controls. Embryos were cultured to the hatched blastocyst stage, and images were recorded every 0.5 h throughout the culture period. The longest cell cycle shifted from 4th to 5th cycle (26 +/- 4 and 44 +/- 5 h) in NT-embryos compared to IVP-embryos (41 +/- 2 and 20 +/- 3 h) and showed greater asynchrony between blastomeres than any other embryo category. Compared to DMAP, CHX prolonged the 1(st) (23 +/- 1 vs. 33 +/- 1 h) and shortened the 3(rd) cell cycle (17 +/- 2 vs. 13 +/- 1 h). Moreover, though cytoskeleton activity was initialised, a larger proportion of CHX embryos was unable to accomplish first cleavage. The parthegenones differed from IVP embryos with respect to the lengths of the 1st, 3rd, and 4th cell cycles and time of hatching. The findings are discussed in relation to known ultrastructural, chromosomal and genomic aberrations found in NT embryos and parthenogenones. We hypothesize that the shift of the longest cell cycle in NT embryos is associated with a shift in the time of major genomic transition.

  6. Natural convection heat transfer within horizontal spent nuclear fuel assemblies

    SciTech Connect

    Canaan, R.E.

    1995-12-01

    Natural convection heat transfer is experimentally investigated in an enclosed horizontal rod bundle, which characterizes a spent nuclear fuel assembly during dry storage and/or transport conditions. The basic test section consists of a square array of sixty-four stainless steel tubular heaters enclosed within a water-cooled rectangular copper heat exchanger. The heaters are supplied with a uniform power generation per unit length while the surrounding enclosure is maintained at a uniform temperature. The test section resides within a vacuum/pressure chamber in order to subject the assembly to a range of pressure statepoints and various backfill gases. The objective of this experimental study is to obtain convection correlations which can be used in order to easily incorporate convective effects into analytical models of horizontal spent fuel systems, and also to investigate the physical nature of natural convection in enclosed horizontal rod bundles in general. The resulting data consist of: (1) measured temperatures within the assembly as a function of power, pressure, and backfill gas; (2) the relative radiative contribution for the range of observed temperatures; (3) correlations of convective Nusselt number and Rayleigh number for the rod bundle as a whole; and (4) correlations of convective Nusselt number as a function of Rayleigh number for individual rods within the array.

  7. Nuclear Technology Series. Course 4: Heat Transfer and Fluid Flow.

    ERIC Educational Resources Information Center

    Center for Occupational Research and Development, Inc., Waco, TX.

    This technical specialty course is one of thirty-five courses designed for use by two-year postsecondary institutions in five nuclear technician curriculum areas: (1) radiation protection technician, (2) nuclear instrumentation and control technician, (3) nuclear materials processing technician, (4) nuclear quality-assurance/quality-control…

  8. A comparison of established and new approaches in ovine and bovine nuclear transfer.

    PubMed

    Peura, Teija T; Vajta, Gábor

    2003-01-01

    Several breakthroughs in nuclear transfer research were first achieved in sheep, although cattle soon became the main livestock species of interest. However, sheep still offer significant advantages both in basic and applied research. With increased interest in cloning of livestock, new approaches have been developed for both sheep and cattle nuclear transfer technology. These include methods for zona-free nuclear transfer that can be performed with or without the use of micromanipulator. Here we describe four different nuclear transfer methods including the traditional micromanipulation-assisted method in sheep, zona-free method in sheep in which the order of enucleation and nucleus delivery have been reversed ("reverse-order" cloning) and zona free manual cloning methods ("hand-made cloning") for embryonic and somatic cloning in cattle. The purpose of this paper is to encourage people to familiarize themselves with these different methods available and to help them choose and test the method most suitable for their particular circumstances.

  9. Heat Transfer Phenomena in Supercritical Water Nuclear Reactors

    SciTech Connect

    Mark H. Anderson; MichaelL. Corradini; Riccardo Bonazza; Jeremy R. Licht

    2007-10-03

    A supercritical water heat transfer facility has been built at the University of Wisconsin to study heat transfer in ancircular and square annular flow channel. A series of integral heat transfer measurements has been carried out over a wide range of heat flux, mas velocity and bulk water temperatures at a pressure of 25 MPa. The circular annular test section geometry is a 1.07 cm diameter heater rod within a 4.29 diameter flow channel.

  10. Molten Chloride Salts for Heat Transfer in Nuclear Systems

    NASA Astrophysics Data System (ADS)

    Ambrosek, James Wallace

    2011-12-01

    A forced convection loop was designed and constructed to examine the thermal-hydraulic performance of molten KCl-MgCl2 (68-32 at %) salt for use in nuclear co-generation facilities. As part of this research, methods for prediction of the thermo-physical properties of salt mixtures for selection of the coolant salt were studied. In addition, corrosion studies of 10 different alloys were exposed to the KCl-MgCl2 to determine a suitable construction material for the loop. Using experimental data found in literature for unary and binary salt systems, models were found, or developed to extrapolate the available experimental data to unstudied salt systems. These property models were then used to investigate the thermo-physical properties of the LINO3-NaNO3-KNO 3-Ca(NO3), system used in solar energy applications. Using these models, the density, viscosity, adiabatic compressibility, thermal conductivity, heat capacity, and melting temperatures of higher order systems can be approximated. These models may be applied to other molten salt systems. Coupons of 10 different alloys were exposed to the chloride salt for 100 hours at 850°C was undertaken to help determine with which alloy to construct the loop. Of the alloys exposed, Haynes 230 had the least amount of weight loss per area. Nickel and Hastelloy N performed best based on maximum depth of attack. Inconel 625 and 718 had a nearly uniform depletion of Cr from the surface of the sample. All other alloys tested had depletion of Cr along the grain boundaries. The Nb in Inconel 625 and 718 changed the way the Cr is depleted in these alloys. Grain-boundary engineering (GBE) of Incoloy 800H improved the corrosion resistance (weight loss and maximum depth of attack) by nearly 50% as compared to the as-received Incoloy 800H sample. A high temperature pump, thermal flow meter, and pressure differential device was designed, constructed and tested for use in the loop, The heat transfer of the molten chloride salt was found to

  11. Human somatic cell nuclear transfer and reproductive cloning: an Ethics Committee opinion.

    PubMed

    2016-04-01

    This document presents arguments that conclude that it is unethical to use somatic cell nuclear transfer (SCNT) for infertility treatment due to concerns about safety; the unknown impact of SCNT on children, families, and society; and the availability of other ethically acceptable means of assisted reproduction. This document replaces the ASRM Ethics Committee report titled, "Human somatic cell nuclear transfer and cloning," last published in Fertil Steril 2012;98:804-7. PMID:26746137

  12. Human somatic cell nuclear transfer and reproductive cloning: an Ethics Committee opinion.

    PubMed

    2016-04-01

    This document presents arguments that conclude that it is unethical to use somatic cell nuclear transfer (SCNT) for infertility treatment due to concerns about safety; the unknown impact of SCNT on children, families, and society; and the availability of other ethically acceptable means of assisted reproduction. This document replaces the ASRM Ethics Committee report titled, "Human somatic cell nuclear transfer and cloning," last published in Fertil Steril 2012;98:804-7.

  13. Nuclear transfer to prevent mitochondrial DNA disorders: revisiting the debate on reproductive cloning.

    PubMed

    Bredenoord, A L; Dondorp, W; Pennings, G; De Wert, G

    2011-02-01

    Preclinical experiments are currently performed to examine the feasibility of several types of nuclear transfer to prevent mitochondrial DNA (mtDNA) disorders. Whereas the two most promising types of nuclear transfer to prevent mtDNA disorders, spindle transfer and pronuclear transfer, do not amount to reproductive cloning, one theoretical variant, blastomere transfer does. This seems the most challenging both technically and ethically. It is prohibited by many jurisdictions and also the scientific community seems to avoid it. Nevertheless, this paper examines the moral acceptability of blastomere transfer as a method to prevent mtDNA disorders. The reason for doing so is that most objections against reproductive cloning refer to reproductive adult cloning, while blastomere transfer would amount to reproductive embryo cloning. After clarifying this conceptual difference, this paper examines whether the main non-safety objections brought forward against reproductive cloning also apply in the context of blastomere transfer. The conclusion is that if this variant were to become safe and effective, dismissing it because it would involve reproductive cloning is unjustified. Nevertheless, as it may lead to more complex ethical appraisals than the other variants, researchers should initially focus on the development of the other types of nuclear transfer to prevent mtDNA disorders.

  14. Cloned embryos from semen. Part 2: Intergeneric nuclear transfer of semen-derived eland (Taurotragus oryx) epithelial cells into bovine oocytes

    USGS Publications Warehouse

    Nel-Themaat, L.; Gomez, M.C.; Pope, C.E.; Lopez, M.; Wirtu, G.; Jenkins, J.A.; Cole, A.; Dresser, B.L.; Bondioli, K.R.; Godke, R.A.

    2008-01-01

    The production of cloned offspring by nuclear transfer (NT) of semen-derived somatic cells holds considerable potential for the incorporation of novel genes into endangered species populations. Because oocytes from endangered species are scarce, domestic species oocytes are often used as cytoplasts for interspecies NT. In the present study, epithelial cells isolated from eland semen were used for intergeneric transfer (IgNT) into enucleated bovine oocytes and compared with bovine NT embryos. Cleavage rates of bovine NT and eland IgNT embryos were similar (80 vs. 83%, respectively; p > 0.05); however, development to the morula and blastocyst stage was higher for bovine NT embryos (38 and 21%, respectively; p < 0.0001), than for eland IgNT embryos (0.5 and 0%, respectively). DNA synthesis was not observed in either bovine NT or eland IgNT cybrids before activation, but in 75 and 70% of bovine NT and eland igNT embryos, respectively, cell-cycle resumption was observed at 16 h postactivation (hpa). For eland IgNT embryos, 13% had ???8 cells at 84 hpa, while 32% of the bovine NT embryos had ???8 cells at the same interval. However, 100 and 66% of bovine NT and eland IgNT embryos, respectively, that had ???8 cells synthesized DNA. From these results we concluded that (1) semen-derived epithelial cell nuclei can interact and be transcriptionally controlled by bovine cytoplast, (2) the first cell-cycle occurred in IgNT embryos, (3) a high frequency of developmental arrest occurs before the eight-cell stage in IgNT embryos, and (4) IgNT embryos that progress through the early cleavage stage arrest can (a) synthesize DNA, (b) progress through subsequent cell cycles, and (c) may have the potential to develop further. ?? 2008 Mary Ann Liebert, Inc.

  15. Cloned embryos from semen. Part 2: intergeneric nuclear transfer of semen-derived eland (Taurotragus oryx) epithelial cells into bovine oocytes.

    PubMed

    Nel-Themaat, Liesl; Gómez, Martha C; Pope, C Earle; Lopez, Monica; Wirtu, Gemechu; Jenkins, Jill A; Cole, Alex; Dresser, Betsy L; Bondioli, Kenneth R; Godke, Robert A

    2008-03-01

    The production of cloned offspring by nuclear transfer (NT) of semen-derived somatic cells holds considerable potential for the incorporation of novel genes into endangered species populations. Because oocytes from endangered species are scarce, domestic species oocytes are often used as cytoplasts for interspecies NT. In the present study, epithelial cells isolated from eland semen were used for intergeneric transfer (IgNT) into enucleated bovine oocytes and compared with bovine NT embryos. Cleavage rates of bovine NT and eland IgNT embryos were similar (80 vs. 83%, respectively; p > 0.05); however, development to the morula and blastocyst stage was higher for bovine NT embryos (38 and 21%, respectively; p < 0.0001), than for eland IgNT embryos (0.5 and 0%, respectively). DNA synthesis was not observed in either bovine NT or eland IgNT cybrids before activation, but in 75 and 70% of bovine NT and eland igNT embryos, respectively, cell-cycle resumption was observed at 16 h postactivation (hpa). For eland IgNT embryos, 13% had > or = 8 cells at 84 hpa, while 32% of the bovine NT embryos had > or = 8 cells at the same interval. However, 100 and 66% of bovine NT and eland IgNT embryos, respectively, that had > or = 8 cells synthesized DNA. From these results we concluded that (1) semen-derived epithelial cell nuclei can interact and be transcriptionally controlled by bovine cytoplast, (2) the first cell-cycle occurred in IgNT embryos, (3) a high frequency of developmental arrest occurs before the eight-cell stage in IgNT embryos, and (4) IgNT embryos that progress through the early cleavage stage arrest can (a) synthesize DNA, (b) progress through subsequent cell cycles, and (c) may have the potential to develop further. PMID:18241126

  16. Generation of cloned mice and nuclear transfer embryonic stem cell lines from urine-derived cells

    PubMed Central

    Mizutani, Eiji; Torikai, Kohei; Wakayama, Sayaka; Nagatomo, Hiroaki; Ohinata, Yasuhide; Kishigami, Satoshi; Wakayama, Teruhiko

    2016-01-01

    Cloning animals by nuclear transfer provides the opportunity to preserve endangered mammalian species. However, there are risks associated with the collection of donor cells from the body such as accidental injury to or death of the animal. Here, we report the production of cloned mice from urine-derived cells collected noninvasively. Most of the urine-derived cells survived and were available as donors for nuclear transfer without any pretreatment. After nuclear transfer, 38–77% of the reconstructed embryos developed to the morula/blastocyst, in which the cell numbers in the inner cell mass and trophectoderm were similar to those of controls. Male and female cloned mice were delivered from cloned embryos transferred to recipient females, and these cloned animals grew to adulthood and delivered pups naturally when mated with each other. The results suggest that these cloned mice had normal fertility. In additional experiments, 26 nuclear transfer embryonic stem cell lines were established from 108 cloned blastocysts derived from four mouse strains including inbreds and F1 hybrids with relatively high success rates. Thus, cells derived from urine, which can be collected noninvasively, may be used in the rescue of endangered mammalian species by using nuclear transfer without causing injury to the animal. PMID:27033801

  17. Generation of cloned mice and nuclear transfer embryonic stem cell lines from urine-derived cells.

    PubMed

    Mizutani, Eiji; Torikai, Kohei; Wakayama, Sayaka; Nagatomo, Hiroaki; Ohinata, Yasuhide; Kishigami, Satoshi; Wakayama, Teruhiko

    2016-01-01

    Cloning animals by nuclear transfer provides the opportunity to preserve endangered mammalian species. However, there are risks associated with the collection of donor cells from the body such as accidental injury to or death of the animal. Here, we report the production of cloned mice from urine-derived cells collected noninvasively. Most of the urine-derived cells survived and were available as donors for nuclear transfer without any pretreatment. After nuclear transfer, 38-77% of the reconstructed embryos developed to the morula/blastocyst, in which the cell numbers in the inner cell mass and trophectoderm were similar to those of controls. Male and female cloned mice were delivered from cloned embryos transferred to recipient females, and these cloned animals grew to adulthood and delivered pups naturally when mated with each other. The results suggest that these cloned mice had normal fertility. In additional experiments, 26 nuclear transfer embryonic stem cell lines were established from 108 cloned blastocysts derived from four mouse strains including inbreds and F1 hybrids with relatively high success rates. Thus, cells derived from urine, which can be collected noninvasively, may be used in the rescue of endangered mammalian species by using nuclear transfer without causing injury to the animal. PMID:27033801

  18. A background to nuclear transfer and its applications in agriculture and human therapeutic medicine.

    PubMed

    Campbell, Keith H S

    2002-03-01

    The development of a single celled fertilized zygote to an animal capable of reproduction involves not only cell division but the differentiation or specialization to numerous cell types forming each tissue and organ of the adult animal. The technique of nuclear transfer allows the reconstruction of an embryo by the transfer of genetic material from a single donor cell, to an unfertilized egg from which the genetic material has been removed. Successful development of live offspring from such embryos demonstrates that the differentiated state of the donor nucleus is not fixed and can be reprogrammed by the egg cytoplasm to control embryo and fetal development. Nuclear transfer has many applications in agriculture and human medicine. This article will review some of the factors associated with the success of embryo development following nuclear transfer and outline the potential uses of the technology.

  19. A background to nuclear transfer and its applications in agriculture and human therapeutic medicine*

    PubMed Central

    Campbell, Keith HS

    2002-01-01

    The development of a single celled fertilized zygote to an animal capable of reproduction involves not only cell division but the differentiation or specialization to numerous cell types forming each tissue and organ of the adult animal. The technique of nuclear transfer allows the reconstruction of an embryo by the transfer of genetic material from a single donor cell, to an unfertilized egg from which the genetic material has been removed. Successful development of live offspring from such embryos demonstrates that the differentiated state of the donor nucleus is not fixed and can be reprogrammed by the egg cytoplasm to control embryo and fetal development. Nuclear transfer has many applications in agriculture and human medicine. This article will review some of the factors associated with the success of embryo development following nuclear transfer and outline the potential uses of the technology. PMID:12033731

  20. An improved heat transfer configuration for a solid-core nuclear thermal rocket engine

    NASA Technical Reports Server (NTRS)

    Clark, John S.; Walton, James T.; Mcguire, Melissa L.

    1992-01-01

    Interrupted flow, impingement cooling, and axial power distribution are employed to enhance the heat-transfer configuration of a solid-core nuclear thermal rocket engine. Impingement cooling is introduced to increase the local heat-transfer coefficients between the reactor material and the coolants. Increased fuel loading is used at the inlet end of the reactor to enhance heat-transfer capability where the temperature differences are the greatest. A thermal-hydraulics computer program for an unfueled NERVA reactor core is employed to analyze the proposed configuration with attention given to uniform fuel loading, number of channels through the impingement wafers, fuel-element length, mass-flow rate, and wafer gap. The impingement wafer concept (IWC) is shown to have heat-transfer characteristics that are better than those of the NERVA-derived reactor at 2500 K. The IWC concept is argued to be an effective heat-transfer configuration for solid-core nuclear thermal rocket engines.

  1. Thermoacoustic sensor for nuclear fuel temperaturemonitoring and heat transfer enhancement

    SciTech Connect

    James A. Smith; Dale K. Kotter; Randall A. Alli; Steven L. Garrett

    2013-05-01

    A new acoustical sensing system for the nuclear power industry has been developed at The Pennsylvania State University in collaboration with Idaho National Laboratories. This sensor uses the high temperatures of nuclear fuel to convert a nuclear fuel rod into a standing-wave thermoacoustic engine. When a standing wave is generated, the sound wave within the fuel rod will be propagated, by acoustic radiation, through the cooling fluid within the reactor or spent fuel pool and can be monitored a remote location external to the reactor. The frequency of the sound can be correlated to an effective temperature of either the fuel or the surrounding coolant. We will present results for a thermoacoustic resonator built into a Nitonic-60 (stainless steel) fuel rod that requires only one passive component and no heat exchangers.

  2. Development of interspecies cloned embryos reconstructed with rabbit (Oryctolagus cuniculus) oocytes and cynomolgus monkey (Macaca fascicularis) fibroblast cell nuclei.

    PubMed

    Yamochi, Takayuki; Kida, Yuta; Oh, Noriyoshi; Ohta, Sei; Amano, Tomoko; Anzai, Masayuki; Kato, Hiromi; Kishigami, Satoshi; Mitani, Tasuku; Matsumoto, Kazuya; Saeki, Kazuhiro; Takenoshita, Makoto; Iritani, Akira; Hosoi, Yoshihiko

    2013-11-01

    Interspecies somatic cell nuclear transfer (ISCNT) has been proposed as a technique to produce cloned offspring of endangered species as well as to investigate nucleus-cytoplasm interactions in mammalian embryo. However, it is still not known which embryo culture medium is optimal for ISCNT embryos for the nuclear donor or the oocyte recipient. We assessed the effects of the culture medium on the developmental competence of the ISCNT embryos by introducing cynomolgus monkey (Macaca fascicularis) fibroblast nuclei into enucleated rabbit (Oryctolagus cuniculus) oocytes (monkey-rabbit embryo). The monkey-rabbit ISCNT embryos that were cultured in mCMRL-1066 developed to the blastocyst stage, although all monkey-rabbit ISCNT embryos cultured in M199 were arrested by the 4-cell stage. When monkey-rabbit ISCNT and rabbit-rabbit somatic cell nuclear transfer (SCNT) embryos were cultured in mCMRL-1066, the blastocyst cell numbers of the monkey-rabbit ISCNT embryos corresponded to the cell numbers of the control rabbit-rabbit SCNT embryos, which were produced from a rabbit fibroblast nucleus and an enucleated rabbit oocyte. In addition, the presence of mitochondria, which were introduced with monkey fibroblasts into rabbit recipient cytoplasm, was confirmed up to the blastocyst stage by polymerase chain reaction (PCR). This study demonstrated that: (1) rabbit oocytes can reprogramme cynomolgus monkey somatic cell nuclei, and support preimplantation development; (2) monkey-rabbit ISCNT embryos developed well in monkey culture medium at early embryonic developmental stages; (3) the cell number of monkey-rabbit ISCNT embryos is similar to that of rabbit-rabbit SCNT embryos; and (4) the mitochondrial fate of monkey-rabbit ISCNT embryos is heteroplasmic from the time just after injection to the blastocyst stage that has roots in both rabbit oocytes and monkey fibroblasts.

  3. Ultrafast Charge Transfer of a Valence Double Hole in Glycine Driven Exclusively by Nuclear Motion.

    PubMed

    Li, Zheng; Vendrell, Oriol; Santra, Robin

    2015-10-01

    We explore theoretically the ultrafast transfer of a double electron hole between the functional groups of glycine after K-shell ionization and subsequent Auger decay. Although a large energy gap of about 15 eV initially exists between the two electronic states involved and coherent electronic dynamics play no role in the hole transfer, we find that the double hole is transferred within 3 to 4 fs between both functional ends of the glycine molecule driven solely by specific nuclear displacements and non-Born-Oppenheimer effects. The nuclear displacements along specific vibrational modes are of the order of 15% of a typical chemical bond between carbon, oxygen, and nitrogen atoms and about 30% for bonds involving hydrogen atoms. The time required for the hole transfer corresponds to less than half a vibrational period of the involved nuclear modes. This finding challenges the common wisdom that nuclear dynamics of the molecular skeleton are unimportant for charge transfer processes at the few-femtosecond time scale and shows that they can even play a prominent role. It also indicates that in x-ray imaging experiments, in which ionization is unavoidable, valence electron redistribution caused by nuclear dynamics might be much faster than previously anticipated. Thus, non-Born-Oppenheimer effects may affect the apparent electron densities extracted from such measurements.

  4. Ultrafast Charge Transfer of a Valence Double Hole in Glycine Driven Exclusively by Nuclear Motion

    NASA Astrophysics Data System (ADS)

    Li, Zheng; Vendrell, Oriol; Santra, Robin

    2015-10-01

    We explore theoretically the ultrafast transfer of a double electron hole between the functional groups of glycine after K -shell ionization and subsequent Auger decay. Although a large energy gap of about 15 eV initially exists between the two electronic states involved and coherent electronic dynamics play no role in the hole transfer, we find that the double hole is transferred within 3 to 4 fs between both functional ends of the glycine molecule driven solely by specific nuclear displacements and non-Born-Oppenheimer effects. The nuclear displacements along specific vibrational modes are of the order of 15% of a typical chemical bond between carbon, oxygen, and nitrogen atoms and about 30% for bonds involving hydrogen atoms. The time required for the hole transfer corresponds to less than half a vibrational period of the involved nuclear modes. This finding challenges the common wisdom that nuclear dynamics of the molecular skeleton are unimportant for charge transfer processes at the few-femtosecond time scale and shows that they can even play a prominent role. It also indicates that in x-ray imaging experiments, in which ionization is unavoidable, valence electron redistribution caused by nuclear dynamics might be much faster than previously anticipated. Thus, non-Born-Oppenheimer effects may affect the apparent electron densities extracted from such measurements.

  5. Functional full-term placentas formed from parthenogenetic embryos using serial nuclear transfer.

    PubMed

    Hikichi, Takafusa; Ohta, Hiroshi; Wakayama, Sayaka; Wakayama, Teruhiko

    2010-09-01

    Mammalian parthenogenetic embryos invariably die in mid-gestation from imprinted gene defects and placental hypoplasia. Based on chimera experiments, trophoblastic proliferation is supposed to be inhibited in the absence of a male genome. Here, we show that parthenogenetic mouse embryonic cell nuclei can be reprogrammed by serial rounds of nuclear transfer without using any genetic modification. The durations of survival in uteri of cloned foetuses derived from green fluorescent protein (GFP)-labelled parthenogenetic cell nuclei were extended with repeated nuclear transfers. After five repeats, live cloned foetuses were obtained up to day 14.5 of gestation; however, they did not survive longer even when we repeated nuclear transfer up to nine times. All foetuses showed intestinal herniation and possessed well-expanded large placentas. When embryonic stem (ES) cells derived from fertilised embryos were aggregated with the cloned embryos, full-term offspring with large placentas were obtained from the chimeric embryos. Those placentas were derived from parthenogenetic cell nuclei, judging from GFP expression. The patterns of imprinted gene expression and methylation status were similar to their parthenogenetic origin, except for Peg10, which showed the same level as in the normal placenta. These results suggest that there is a limitation for foetal development in the ability to reprogramme imprinted genes by repeated rounds of nuclear transfer. However, the placentas of parthenogenetic embryos can escape epigenetic regulation when developed using nuclear transfer techniques and can support foetal development to full gestation.

  6. Interspecies interactions and potential Influenza A virus risk in small swine farms in Peru

    PubMed Central

    2012-01-01

    Background The recent avian influenza epidemic in Asia and the H1N1 pandemic demonstrated that influenza A viruses pose a threat to global public health. The animal origins of the viruses confirmed the potential for interspecies transmission. Swine are hypothesized to be prime "mixing vessels" due to the dual receptivity of their trachea to human and avian strains. Additionally, avian and human influenza viruses have previously been isolated in swine. Therefore, understanding interspecies contact on smallholder swine farms and its potential role in the transmission of pathogens such as influenza virus is very important. Methods This qualitative study aimed to determine swine-associated interspecies contacts in two coastal areas of Peru. Direct observations were conducted at both small-scale confined and low-investment swine farms (n = 36) and in open areas where swine freely range during the day (n = 4). Interviews were also conducted with key stakeholders in swine farming. Results In both locations, the intermingling of swine and domestic birds was common. An unexpected contact with avian species was that swine were fed poultry mortality in 6/20 of the farms in Chancay. Human-swine contacts were common, with a higher frequency on the confined farms. Mixed farming of swine with chickens or ducks was observed in 36% of all farms. Human-avian interactions were less frequent overall. Use of adequate biosecurity and hygiene practices by farmers was suboptimal at both locations. Conclusions Close human-animal interaction, frequent interspecies contacts and suboptimal biosecurity and hygiene practices pose significant risks of interspecies influenza virus transmission. Farmers in small-scale swine production systems constitute a high-risk population and need to be recognized as key in preventing interspecies pathogen transfer. A two-pronged prevention approach, which offers educational activities for swine farmers about sound hygiene and biosecurity practices and

  7. Fatal attraction: Explaining Russia's sensitive nuclear transfers to Iran

    NASA Astrophysics Data System (ADS)

    Kuchinsky, Leah R.

    This paper explores Russia's sensitive nuclear assistance to Iran in an effort to determine why a supplier state might proliferate against its own apparent security interests. The goal is to help readers understand the supply-side dynamics of nuclear proliferation. Through careful reconstruction of the historical narrative, using open source data, this study tests the plausibility of a "fatalistic calculus" explanation, identified by Stephen Sestanovich as a possible driver for Russia's behavior. According to the hypothesis, Russia has cooperated with Iran as a way both to stay in the good graces of a neighbor that is suspected of developing nuclear weapons and to win short-term influence and profits. The paper also examines the role of other factors advanced in the existing supply-side literature, such as economic motives identified by physicist and nonproliferation scholar David Albright. The findings show that bureaucratic, economic and fatalistic factors have each played a role in motivating Russia's cooperation with Iran, with their relative importance shifting over time. Fatalism begets a strategy of Russian "minimaxing," in the lexicon of Russia scholar Robert Freedman, wherein Russia attempts to minimize damage to its relationship with the U.S. while maximizing influence in Iran via nuclear cooperation. Fatalism, as actualized by minimaxing, best explains Russia's behavior after former Russian president Vladmir Putin came to power, when the bureaucratic and economic arguments become less cogent.

  8. Modeling transient heat transfer in nuclear waste repositories.

    PubMed

    Yang, Shaw-Yang; Yeh, Hund-Der

    2009-09-30

    The heat of high-level nuclear waste may be generated and released from a canister at final disposal sites. The waste heat may affect the engineering properties of waste canisters, buffers, and backfill material in the emplacement tunnel and the host rock. This study addresses the problem of the heat generated from the waste canister and analyzes the heat distribution between the buffer and the host rock, which is considered as a radial two-layer heat flux problem. A conceptual model is first constructed for the heat conduction in a nuclear waste repository and then mathematical equations are formulated for modeling heat flow distribution at repository sites. The Laplace transforms are employed to develop a solution for the temperature distributions in the buffer and the host rock in the Laplace domain, which is numerically inverted to the time-domain solution using the modified Crump method. The transient temperature distributions for both the single- and multi-borehole cases are simulated in the hypothetical geological repositories of nuclear waste. The results show that the temperature distributions in the thermal field are significantly affected by the decay heat of the waste canister, the thermal properties of the buffer and the host rock, the disposal spacing, and the thickness of the host rock at a nuclear waste repository.

  9. Modeling transient heat transfer in nuclear waste repositories.

    PubMed

    Yang, Shaw-Yang; Yeh, Hund-Der

    2009-09-30

    The heat of high-level nuclear waste may be generated and released from a canister at final disposal sites. The waste heat may affect the engineering properties of waste canisters, buffers, and backfill material in the emplacement tunnel and the host rock. This study addresses the problem of the heat generated from the waste canister and analyzes the heat distribution between the buffer and the host rock, which is considered as a radial two-layer heat flux problem. A conceptual model is first constructed for the heat conduction in a nuclear waste repository and then mathematical equations are formulated for modeling heat flow distribution at repository sites. The Laplace transforms are employed to develop a solution for the temperature distributions in the buffer and the host rock in the Laplace domain, which is numerically inverted to the time-domain solution using the modified Crump method. The transient temperature distributions for both the single- and multi-borehole cases are simulated in the hypothetical geological repositories of nuclear waste. The results show that the temperature distributions in the thermal field are significantly affected by the decay heat of the waste canister, the thermal properties of the buffer and the host rock, the disposal spacing, and the thickness of the host rock at a nuclear waste repository. PMID:19376651

  10. Interspecies scaling of regional drug delivery.

    PubMed

    Dedrick, R L

    1986-11-01

    Calculation of the pharmacokinetic advantage of regional drug administration requires knowledge of the relevant intercompartment transport parameter. In a lumped model this is the blood (or plasma) flow rate for intra-arterial drug infusion or the permeability-area product for intraperitoneal or intrathecal administration. It is suggested that the perfusion of many tissues and the intrinsic permeability of the peritoneal surface or the brain-cerebrospinal fluid interface are similar among mammals. This provides a clear allometric basis for interspecies scaling based on organ size or surface areas. Intra-arterial or intrathecal treatment of the brain or meninges is a particularly interesting problem because of the relatively large brain of humans and because increased folding results in a cortical surface area that is almost proportional to brain size. Major unresolved issues remain concerning the distributed character of the processes such as streaming of drug infused into an artery and nonuniform mixing of cerebrospinal fluid.

  11. Interspecies modeling of inhaled particle deposition patterns

    SciTech Connect

    Martonen, T.B.; Zhang, Z.; Yang, Y.

    1992-01-01

    To evaluate the potential toxic effects of ambient contaminants or therapeutic effects of airborne drugs, inhalation exposure experiments can be performed with surrogate laboratory animals. Herein, an interspecies particle deposition theory is presented for physiologically based pharmacokinetic modeling. It is derived to improve animal testing protocols. The computer code describes the behavior and fate of particles in the lungs of human subjects and a selected surrogate, the laboratory rat. In the simulations CO2 is integrated with exposure chamber atmospheres, and its concentrations regulated to produce rat breathing profiles corresponding to selected levels of human physical activity. The dosimetric model is used to calculate total, compartmental (i.e., tracheobronchial and pulmonary), and localized distribution patterns of inhaled particles in rats and humans for comparable ventilatory conditions. It is demonstrated that the model can be used to predetermine the exposure conditions necessary to produce deposition patterns in rats that are equivalent to those in humans at prescribed physical activities.

  12. Histone deacetylase inhibitor improves the development and acetylation levels of cat-cow interspecies cloned embryos.

    PubMed

    Wittayarat, Manita; Sato, Yoko; Do, Lanh Thi Kim; Morita, Yasuhiro; Chatdarong, Kaywalee; Techakumphu, Mongkol; Taniguchi, Masayasu; Otoi, Takeshige

    2013-08-01

    Abnormal epigenetic reprogramming, such as histone acetylation, might cause low efficiency of interspecies somatic cell nuclear transfer (iSCNT). This study was conducted to evaluate the effects of trichostatin A (TSA) on the developmental competence and histone acetylation of iSCNT embryos reconstructed from cat somatic cells and bovine cytoplasm. The iSCNT cat and parthenogenetic bovine embryos were treated with various concentrations of TSA (0, 25, 50, or 100 nM) for 24 h, respectively, following fusion and activation. Treatment with 50 nM TSA produced significantly higher rates of cleavage and blastocyst formation (84.3% and 4.6%, respectively) of iSCNT embryos than the rates of non-TSA-treated iSCNT embryos (63.8% and 0%, respectively). Similarly, the treatment of 50 nM TSA increased the blastocyst formation rate of parthenogenetic bovine embryos. The acetylation levels of histone H3 lysine 9 (H3K9) in the iSCNT embryos with the treatment of 50 nM TSA were similar to those of in vitro-fertilized embryos and significantly higher (p<0.05) than those of non-TSA-treated iSCNT embryos (control), irrespective of the embryonic development stage (two-cell, four-cell, and eight-cell stages). These results indicated that the treatment of 50 nM TSA postfusion was beneficial for development to the blastocyst stage of iSCNT cat embryos and correlated with the increasing levels of acetylation at H3K9. PMID:23790014

  13. Histone deacetylase inhibitor improves the development and acetylation levels of cat-cow interspecies cloned embryos.

    PubMed

    Wittayarat, Manita; Sato, Yoko; Do, Lanh Thi Kim; Morita, Yasuhiro; Chatdarong, Kaywalee; Techakumphu, Mongkol; Taniguchi, Masayasu; Otoi, Takeshige

    2013-08-01

    Abnormal epigenetic reprogramming, such as histone acetylation, might cause low efficiency of interspecies somatic cell nuclear transfer (iSCNT). This study was conducted to evaluate the effects of trichostatin A (TSA) on the developmental competence and histone acetylation of iSCNT embryos reconstructed from cat somatic cells and bovine cytoplasm. The iSCNT cat and parthenogenetic bovine embryos were treated with various concentrations of TSA (0, 25, 50, or 100 nM) for 24 h, respectively, following fusion and activation. Treatment with 50 nM TSA produced significantly higher rates of cleavage and blastocyst formation (84.3% and 4.6%, respectively) of iSCNT embryos than the rates of non-TSA-treated iSCNT embryos (63.8% and 0%, respectively). Similarly, the treatment of 50 nM TSA increased the blastocyst formation rate of parthenogenetic bovine embryos. The acetylation levels of histone H3 lysine 9 (H3K9) in the iSCNT embryos with the treatment of 50 nM TSA were similar to those of in vitro-fertilized embryos and significantly higher (p<0.05) than those of non-TSA-treated iSCNT embryos (control), irrespective of the embryonic development stage (two-cell, four-cell, and eight-cell stages). These results indicated that the treatment of 50 nM TSA postfusion was beneficial for development to the blastocyst stage of iSCNT cat embryos and correlated with the increasing levels of acetylation at H3K9.

  14. Single and Multi-Nucleon Transfer Reactions for Nuclear Moment Studies Toward Radioactive-Ion Beams

    SciTech Connect

    Lozeva, R. L.; Georgiev, G. P.; Audi, G.; Cabaret, S.; Fiori, E.; Gaulard, C.; Hauschilda, K.; Lopez-Martens, A.; Risegari, L.; Blazhev, A.; Jolie, J.; Moschner, K.; Zell, K.-O.; Daugas, J.-M.; Faul, T.; Morel, P.; Roig, O.; Ferraton, M.; Ibrahim, F.

    2010-04-30

    This study is a part of an experimental program to measure nuclear moments in transfer reactions. It aims to probe for a first time the nuclear -spin orientation in multi-nucleon transfer. Fist experiments were performed to measure the quadrupole moment of an isomeric state in {sup 66}Cu (I{sup p}i 6{sup -}, E{sub x} = 1154 keV, T{sub 1/2} = 595(20) ns) in single nucleon transfer and the population of mus isomers in {sup 66}Cu and {sup 63}Ni in multi-nucleon transfer. The experimentally tested methodology allows broad applications toward more exotic species and feasibility of these reactions to produce species away from stability.

  15. Pig transgenesis by piggyBac transposition in combination with somatic cell nuclear transfer.

    PubMed

    Wu, Zhenfang; Xu, Zhiqian; Zou, Xian; Zeng, Fang; Shi, Junsong; Liu, Dewu; Urschitz, Johann; Moisyadi, Stefan; Li, Zicong

    2013-12-01

    The production of animals by somatic cell nuclear transfer (SCNT) is inefficient, with approximately 2% of micromanipulated oocytes going to term and resulting in live births. However, it is the most commonly used method for the generation of cloned transgenic livestock as it facilitates the attainment of transgenic animals once the nuclear donor cells are stably transfected and more importantly as alternatives methods of transgenesis in farm animals have proven even less efficient. Here we describe piggyBac-mediated transposition of a transgene into porcine primary cells and use of these genetically modified cells as nuclear donors for the generation of transgenic pigs by SCNT. Gene transfer by piggyBac transposition serves to provide an alternative approach for the transfection of nuclear donor cells used in SCNT.

  16. Treating humanity as an inviolable end: an analysis of contraception and altered nuclear transfer.

    PubMed

    Masek, Lawrence

    2008-04-01

    I argue that contraception is morally wrong but that periodic abstinence (or natural family planning) is not. Further, I argue that altered nuclear transfer -- a proposed technique for creating human stem cells without destroying human embryos -- is morally wrong for the same reason that contraception is. Contrary to what readers might expect, my argument assumes nothing about the morality of cloning or abortion and requires no premises about God or natural teleology. Instead, I argue that contraception and altered nuclear transfer are morally wrong because they fail to treat humanity as an inviolable end. PMID:18480499

  17. Cell-mediated transgenesis in rabbits: chimeric and nuclear transfer animals.

    PubMed

    Zakhartchenko, V; Flisikowska, T; Li, S; Richter, T; Wieland, H; Durkovic, M; Rottmann, O; Kessler, B; Gungor, T; Brem, G; Kind, A; Wolf, E; Schnieke, A

    2011-02-01

    The ability to perform precise genetic engineering such as gene targeting in rabbits would benefit biomedical research by enabling, for example, the generation of genetically defined rabbit models of human diseases. This has so far not been possible because of the lack of functional rabbit embryonic stem cells and the high fetal and perinatal mortality associated with rabbit somatic cell nuclear transfer. We examined cultured pluripotent and multipotent cells for their ability to support the production of viable animals. Rabbit putative embryonic stem (ES) cells were derived and shown capable of in vitro and in vivo pluripotent differentiation. We report the first live born ES-derived rabbit chimera. Rabbit mesenchymal stem cells (MSCs) were derived from bone marrow, and multipotent differentiation was demonstrated in vitro. Nuclear transfer was carried out with both cell types, and embryo development was assessed in vitro and in vivo. Rabbit MSCs were markedly more successful than ES cells as nuclear donors. MSCs were transfected with fluorescent reporter gene constructs and assessed for nuclear transfer competence. Transfected MSCs supported development with similar efficiency as normal MSCs and resulted in the first live cloned rabbits from genetically manipulated MSCs. Reactivation of fluorescence reporter gene expression in reconstructed embryos was investigated as a means of identifying viable embryos in vitro but was not a reliable predictor. We also examined serial nuclear transfer as a means of rescuing dead animals.

  18. Genome Sequence of the Lager Brewing Yeast, an Interspecies Hybrid

    PubMed Central

    Nakao, Yoshihiro; Kanamori, Takeshi; Itoh, Takehiko; Kodama, Yukiko; Rainieri, Sandra; Nakamura, Norihisa; Shimonaga, Tomoko; Hattori, Masahira; Ashikari, Toshihiko

    2009-01-01

    This work presents the genome sequencing of the lager brewing yeast (Saccharomyces pastorianus) Weihenstephan 34/70, a strain widely used in lager beer brewing. The 25 Mb genome comprises two nuclear sub-genomes originating from Saccharomyces cerevisiae and Saccharomyces bayanus and one circular mitochondrial genome originating from S. bayanus. Thirty-six different types of chromosomes were found including eight chromosomes with translocations between the two sub-genomes, whose breakpoints are within the orthologous open reading frames. Several gene loci responsible for typical lager brewing yeast characteristics such as maltotriose uptake and sulfite production have been increased in number by chromosomal rearrangements. Despite an overall high degree of conservation of the synteny with S. cerevisiae and S. bayanus, the syntenies were not well conserved in the sub-telomeric regions that contain lager brewing yeast characteristic and specific genes. Deletion of larger chromosomal regions, a massive unilateral decrease of the ribosomal DNA cluster and bilateral truncations of over 60 genes reflect a post-hybridization evolution process. Truncations and deletions of less efficient maltose and maltotriose uptake genes may indicate the result of adaptation to brewing. The genome sequence of this interspecies hybrid yeast provides a new tool for better understanding of lager brewing yeast behavior in industrial beer production. PMID:19261625

  19. Genome sequence of the lager brewing yeast, an interspecies hybrid.

    PubMed

    Nakao, Yoshihiro; Kanamori, Takeshi; Itoh, Takehiko; Kodama, Yukiko; Rainieri, Sandra; Nakamura, Norihisa; Shimonaga, Tomoko; Hattori, Masahira; Ashikari, Toshihiko

    2009-04-01

    This work presents the genome sequencing of the lager brewing yeast (Saccharomyces pastorianus) Weihenstephan 34/70, a strain widely used in lager beer brewing. The 25 Mb genome comprises two nuclear sub-genomes originating from Saccharomyces cerevisiae and Saccharomyces bayanus and one circular mitochondrial genome originating from S. bayanus. Thirty-six different types of chromosomes were found including eight chromosomes with translocations between the two sub-genomes, whose breakpoints are within the orthologous open reading frames. Several gene loci responsible for typical lager brewing yeast characteristics such as maltotriose uptake and sulfite production have been increased in number by chromosomal rearrangements. Despite an overall high degree of conservation of the synteny with S. cerevisiae and S. bayanus, the syntenies were not well conserved in the sub-telomeric regions that contain lager brewing yeast characteristic and specific genes. Deletion of larger chromosomal regions, a massive unilateral decrease of the ribosomal DNA cluster and bilateral truncations of over 60 genes reflect a post-hybridization evolution process. Truncations and deletions of less efficient maltose and maltotriose uptake genes may indicate the result of adaptation to brewing. The genome sequence of this interspecies hybrid yeast provides a new tool for better understanding of lager brewing yeast behavior in industrial beer production.

  20. Assessing Uncertainty of Interspecies Correlation Estimation Models for Aromatic Compounds

    EPA Science Inventory

    We developed Interspecies Correlation Estimation (ICE) models for aromatic compounds containing 1 to 4 benzene rings to assess uncertainty in toxicity extrapolation in two data compilation approaches. ICE models are mathematical relationships between surrogate and predicted test ...

  1. Significant improvement of pig cloning efficiency by treatment with LBH589 after somatic cell nuclear transfer.

    PubMed

    Jin, Jun-Xue; Li, Suo; Gao, Qing-Shan; Hong, Yu; Jin, Long; Zhu, Hai-Ying; Yan, Chang-Guo; Kang, Jin-Dan; Yin, Xi-Jun

    2013-10-01

    The low success rate of animal cloning by somatic cell nuclear transfer (SCNT) associates with epigenetic aberrancy, including the abnormal acetylation of histones. Altering the epigenetic status by histone deacetylase inhibitors (HDACi) enhances the developmental potential of SCNT embryos. In the current study, we examined the effects of LBH589 (panobinostat), a novel broad-spectrum HDACi, on the nuclear reprogramming and development of pig SCNT embryos in vitro. In experiment 1, we compared the in vitro developmental competence of nuclear transfer embryos treated with different concentrations of LBH589. Embryos treated with 50 nM LBH589 for 24 hours showed a significant increase in the rate of blastocyst formation compared with the control or embryos treated with 5 or 500 nM LBH589 (32.4% vs. 11.8%, 12.1%, and 10.0%, respectively, P < 0.05). In experiment 2, we examined the in vitro developmental competence of nuclear transfer embryos treated with 50 nM LBH589 for various intervals after activation and 6-dimethylaminopurine. Embryos treated for 24 hours had higher rates of blastocyst formation than the other groups. In experiment 3, when the acetylation of H4K12 was examined in SCNT embryos treated for 6 hours with 50 nM LBH589 by immunohistochemistry, the staining intensities of these proteins in LBH589-treated SCNT embryos were significantly higher than in the control. In experiment 4, LBH589-treated nuclear transfer and control embryos were transferred into surrogate mothers, resulting in three (100%) and two (66.7%) pregnancies, respectively. In conclusion, LBH589 enhances the nuclear reprogramming and developmental potential of SCNT embryos by altering the epigenetic status and expression, and increasing blastocyst quality.

  2. Quantum state transfer between an optomechanical cavity and a diamond nuclear spin ensemble

    NASA Astrophysics Data System (ADS)

    Feng, Zhi-Bo; Wang, Hong-Ling; Yan, Run-Ying

    2016-08-01

    We explore an efficient scheme for transferring quantum state between an optomechanical cavity and nuclear spins of nitrogen-vacancy centers in diamond, where quantum information can be efficiently stored (retrieved) into (from) the nuclear spin ensemble assisted by a mechanical resonator in a dispersive regime. Our scheme works for a broad range of cavity frequencies and might have potential applications in employing the nuclear spin ensemble as a memory in quantum information processing. The feasibility of our protocol is analyzed using currently available parameters.

  3. Passive heat-transfer means for nuclear reactors. [LMFBR

    DOEpatents

    Burelbach, J.P.

    1982-06-10

    An improved passive cooling arrangement is disclosed for maintaining adjacent or related components of a nuclear reactor within specified temperature differences. Specifically, heat pipes are operatively interposed between the components, with the vaporizing section of the heat pipe proximate the hot component operable to cool it and the primary condensing section of the heat pipe proximate the other and cooler component operable to heat it. Each heat pipe further has a secondary condensing section that is located outwardly beyond the reactor confinement and in a secondary heat sink, such as air ambient the containment, that is cooler than the other reactor component. By having many such heat pipes, an emergency passive cooling system is defined that is operative without electrical power.

  4. Comparison of two approaches to nuclear transfer in the bovine: hand-made cloning with modifications and the conventional nuclear transfer technique.

    PubMed

    Tecirlioglu, R Tayfur; Cooney, Melissa A; Lewis, Ian M; Korfiatis, Natasha A; Hodgson, Renee; Ruddock, Nancy T; Vajta, Gábor; Downie, Shara; Trounson, Alan O; Holland, Michael K; French, Andrew J

    2005-01-01

    The aim of the present study was to compare the in vitro and in vivo developmental competence of hand-made cloning (HMC) embryos with the conventional nuclear transfer (NT) method using five somatic cell lines and in vitro-fertilised (IVF; control) embryos. Modifications to the HMC procedure included fusion efficiency optimisation, effect of cytoplasmic volume and cloned embryo aggregation. The developmental competence of blastocysts from each of the treatment groups and cell lines used was assessed following transfer to 345 recipients. Vitrification was also used to enable management of recipient resources and to assess the susceptibility of membranes to cryopreservation following zona removal. Increasing cytoplasmic volume to 150% or aggregating two embryos improved the blastocyst development rate and increased the total cell number. Although HMC embryo transfers established a significantly higher pregnancy rate on Day 30 than fresh IVF or NT embryo transfers, the overall outcome in terms of cloned live births derived from either fresh or vitrified/thawed HMC or NT embryo transfers across the five cell lines did not differ. The birth and continued survival of clones produced with HMC technology with equivalent efficiency to NT shows that it can be used as an alternative method for the generation of cloned offspring in the bovine.

  5. One-two step transfer observed in 16O+11B nuclear system

    NASA Astrophysics Data System (ADS)

    Hamada, Sh.; Burtebayev, N.

    2015-06-01

    The angular distribution measurements for 16O ion beam elastically scattered from 11B target of thickness 32.9μg/cm2 at energy 22.4 MeV had been performed in the cyclotron DC-60 INP NNC RK. The previous measurements for 16O+11B nuclear system at energies 27, 30, 32.5 and 35 MeV showed an increase in the differential cross-section at backward angles due to the contribution of cluster transfer. Such transfer process could not be described in terms of optical model (OM); it could be described within the framework of distorted wave Born approximation method implemented in FRESCO code. Both one (5Li) and two-step transfer (proton transfer followed by Alpha transfer) were taken into considerations. We have extracted the spectroscopic amplitude (SA) for the configuration 16O→11B+5Li.

  6. Methanosarcina spp. drive vinyl chloride dechlorination via interspecies hydrogen transfer.

    PubMed

    Heimann, Axel C; Batstone, Damien J; Jakobsen, Rasmus

    2006-04-01

    Two highly enriched cultures containing Dehalococcoides spp. were used to study the effect of aceticlastic methanogens on reductive vinyl chloride (VC) dechlorination. In terms of aceticlastic methanogens, one culture was dominated by Methanosaeta, while the other culture was dominated by Methanosarcina, as determined by fluorescence in situ hybridization. Cultures amended with 2-bromoethanesulfonate (BES), an efficient inhibitor of methanogens, exhibited slow VC dechlorination when grown on acetate and VC. Methanogenic cultures dominated by Methanosaeta had no impact on dechlorination rates, compared to BES-amended controls. In contrast, methanogenic cultures dominated by Methanosarcina displayed up to sevenfold-higher rates of VC dechlorination than their BES-amended counterparts. Methanosarcina-dominated cultures converted a higher percentage of [2-(14)C]acetate to (14)CO(2) when concomitant VC dechlorination took place, compared to nondechlorinating controls. Respiratory indices increased from 0.12 in nondechlorinating cultures to 0.51 in actively dechlorinating cultures. During VC dechlorination, aqueous hydrogen (H(2)) concentrations dropped to 0.3 to 0.5 nM. However, upon complete VC consumption, H(2) levels increased by a factor of 10 to 100, indicating active hydrogen production from acetate oxidation. This process was thermodynamically favorable by means of the extremely low H(2) levels during dechlorination. VC degradation in nonmethanogenic cultures was not inhibited by BES but was limited by the availability of H(2) as electron donor, in cultures both with and without BES. These findings all indicate that Methanosarcina (but not Methanosaeta), while cleaving acetate to methane, simultaneously oxidizes acetate to CO(2) plus H(2), driving hydrogenotrophic dehalorespiration of VC to ethene by Dehalococcoides.

  7. Efficiency of porcine somatic cell nuclear transfer - a retrospective study of factors related to embryo recipient and embryos transferred.

    PubMed

    Huang, Yongye; Ouyang, Hongsheng; Yu, Hao; Lai, Liangxue; Pang, Daxin; Li, Zhanjun

    2013-01-01

    The successful generation of pigs via somatic cell nuclear transfer depends on reducing risk factors in several aspects. To provide an overview of some influencing factors related to embryo transfer, the follow-up data related to cloned pig production collected in our laboratory was examined. (i) Spring showed a higher full-term pregnancy rate compared with winter (33.6% vs 18.6%, P = 0.006). Furthermore, a regression equation can be drawn between full-term pregnancy numbers and pregnancy numbers in different months (y = 0.692x-3.326). (ii) There were no significant differences detected in the number of transferred embryos between surrogate sows exhibiting full-term development compared to those that did not. (iii) Non-ovulating surrogate sows presented a higher percentage of full-term pregnancies compared with ovulating sows (32.0% vs 17.5%, P = 0.004; respectively). (iv) Abortion was most likely to take place between Day 27 to Day 34. (v) Based on Life Table Survival Analysis, delivery in normally fertilized and surrogate sows is expected to be completed before Day 117 or Day 125, respectively. Additionally, the length of pregnancy in surrogate sows was negatively correlated with the average litter size, which was not found for normally fertilized sows. In conclusion, performing embryo transfer in appropriate seasons, improving the quality of embryos transferred, optimizing the timing of embryo transfer, limiting the occurrence of abortion, combined with ameliorating the management of delivery, is expected to result in the harvest of a great number of surviving cloned piglets. PMID:24244859

  8. The heat transfer analysis of a microinsulation for nuclear microbatteries

    NASA Astrophysics Data System (ADS)

    Yao, Rui

    Microelectromechanical Systems (MEMS) have not gained wide use because they lack the on-device power required by many important applications. Numerous studies have been carried out regarding technologies for providing electrical power to MEMS devices. Compared with solar or chemical energies, radioisotope sources have the advantage of large energy density and long lifetime. The radioisotope decay heat could be directly converted to electricity using thermoelectric or thermionic techniques. As a crucial part of this process, a microinsulation concept has been designed to maintain the radioisotope source temperature high in order to obtain good conversion efficiency. This paper reviews thermionic energy conversion technology and creates heat transfer models for a Micro Heat Barrier (MHB) and radioisotope powered thermionic microbattery developed by Sandia National Laboratory. The computed results indicate that the MHB has apparent thermal conductivity on the order of 10-4 W/mK at vacuum and is capable of providing sufficient thermal resistance to produce a high conversion efficiency for a thermionic microbattery. A new microinsulation concept using photo resist SU-8 is designed and the photolithography fabrication process is described. The microinsulation sample's apparent thermal conductivity is measured and compared with theoretical results. The experimental results show that a typical design has apparent thermal conductivity on the order of 10-4 W/mK at vacuum and have a good agreement with what the model predicts.

  9. Somatic cell nuclear transfer (cloning): implications for the medical practitioner.

    PubMed

    Tong, W F; Ng, Y F; Ng, S C

    2002-07-01

    The current century will bring tremendous changes to the science and the practice of medicine. This century will be acknowledged as the century of Biology as the fusion of molecular genetics and experimental embryology pushes the barriers of science beyond perimeters that we have thought existed, as much as the past century was the century of Physics, with all the exact scientific calculations and predictions, resulting in electricity, nuclear power and quantum physics. The first major breakthrough has been the pioneering work of Wilmut and Campbell, first with the birth of Megan and Moran in 1995 (1), followed by the birth of Dolly the sheep, the first reported mammalian clone from a fully differentiated adult cell, reported in July 1996 (2). However, current cloning techniques are an extension of over 40 years of research using nuclei derived from non-human embryonic and fetal cells. However, following the birth of Dolly, the prospects of cloning technology have extended to ethically hazier areas of human cloning and embryonic stem cell research. This review hopes to bring the reader closer to the science and the ethics of this new technology, and what the implications are for the medical practitioner.

  10. Tie Tube Heat Transfer Modeling for Bimodal Nuclear Thermal Rockets

    NASA Astrophysics Data System (ADS)

    Clough, Joshua A.; Starkey, Ryan P.; Lewis, Mark J.; Lavelle, Thomas M.

    2007-01-01

    Bimodal nuclear thermal rocket systems have been shown to reduce the weight and cost of space vehicles to Mars and beyond by utilizing the reactor for power generation in the relatively long duration between burns in an interplanetary trajectory. No information, however, is available regarding engine and reactor-level operation of such bimodal systems. The purpose of this project is to generate engine and reactor models with sufficient fidelity and flexibility to accurately study the component-level effects of operating a propulsion-designed reactor at power generation levels. Previous development of a 1-D reactor and tie tube model found that ignoring heat generation inside of the tie tube leads to under-prediction of the temperature change and over-prediction of pressure change across the tie tube. This paper will present the development and results of a tie tube model that has been extended to account for heat generation, specifically in the moderator layer. This model is based on a 1-D distribution of power in the fuel elements and tie tubes, as a precursor to an eventual neutron-driven reactor model.

  11. Tie Tube Heat Transfer Modeling for Bimodal Nuclear Thermal Rockets

    SciTech Connect

    Clough, Joshua A.; Starkey, Ryan P.; Lewis, Mark J.; Lavelle, Thomas M.

    2007-01-30

    Bimodal nuclear thermal rocket systems have been shown to reduce the weight and cost of space vehicles to Mars and beyond by utilizing the reactor for power generation in the relatively long duration between burns in an interplanetary trajectory. No information, however, is available regarding engine and reactor-level operation of such bimodal systems. The purpose of this project is to generate engine and reactor models with sufficient fidelity and flexibility to accurately study the component-level effects of operating a propulsion-designed reactor at power generation levels. Previous development of a 1-D reactor and tie tube model found that ignoring heat generation inside of the tie tube leads to under-prediction of the temperature change and over-prediction of pressure change across the tie tube. This paper will present the development and results of a tie tube model that has been extended to account for heat generation, specifically in the moderator layer. This model is based on a 1-D distribution of power in the fuel elements and tie tubes, as a precursor to an eventual neutron-driven reactor model.

  12. Somatic cell nuclear transfer (cloning): implications for the medical practitioner.

    PubMed

    Tong, W F; Ng, Y F; Ng, S C

    2002-07-01

    The current century will bring tremendous changes to the science and the practice of medicine. This century will be acknowledged as the century of Biology as the fusion of molecular genetics and experimental embryology pushes the barriers of science beyond perimeters that we have thought existed, as much as the past century was the century of Physics, with all the exact scientific calculations and predictions, resulting in electricity, nuclear power and quantum physics. The first major breakthrough has been the pioneering work of Wilmut and Campbell, first with the birth of Megan and Moran in 1995 (1), followed by the birth of Dolly the sheep, the first reported mammalian clone from a fully differentiated adult cell, reported in July 1996 (2). However, current cloning techniques are an extension of over 40 years of research using nuclei derived from non-human embryonic and fetal cells. However, following the birth of Dolly, the prospects of cloning technology have extended to ethically hazier areas of human cloning and embryonic stem cell research. This review hopes to bring the reader closer to the science and the ethics of this new technology, and what the implications are for the medical practitioner. PMID:12437047

  13. Trichostatin A modified histone covalent pattern and enhanced expression of pluripotent genes in interspecies black-footed cat cloned embryos but did not improve in vitro and in vivo viability.

    PubMed

    Gómez, Martha C; Pope, C Earle; Biancardi, Monica N; Dumas, Cherie; Galiguis, Jason; Morris, Anna Claire; Wang, Guoshun; Dresser, Betsy L

    2011-08-01

    Abstract The black-footed cat (BFC; Felis nigripes), one of the smallest wild cats, is listed as threatened. Interspecies somatic cell nuclear transfer (Is-SCNT) offers the possibility of preserving endangered species. Development to term of interspecies BFC (Is-BFC) cloned embryos has not been obtained, possibly due to abnormal epigenetic reprogramming. Treatment of intraspecies cloned embryos with TSA improves nuclear reprogramming and in vitro and in vivo viability. In this study, we evaluated (1) whether covalent histone modifications differ between Is-BFC cloned embryos and their IVF counterparts, (2) the optimal TSA concentration and exposure times to modify the covalent histone patterns, (3) if TSA enhances in vitro and in vivo developmental competence of cloned embryos, and (4) expression of pluripotent genes. Results indicated that the covalent histone modifications of Is-BFC cloned embryos aberrantly differ from their DSH-IVF counterpart embryos. Aberrant epigenetic events may be due partially to the inability of the DSH cytoplasm to modify the restrictive epigenetic marks of the BFC nuclei after somatic cell nuclear transfer (SCNT). Incomplete remodeling of the histone H3K9me2 in Is-BFC cloned embryos possibly contributes to abnormal expression of pluripotent genes and low embryonic development. Treatment of Is-BFC cloned embryos with TSA remodeled the covalent pattern in H3K9ac and H3K9me2, resembling epigenetic patterns in IVF counterpart embryos, and resulted in activation of some pluripotent genes. However, genomic reprogramming of Is-BFC cloned blastocysts did not follow the same reprogramming pattern observed in DSH-IVF embryos, and in vitro and in vivo developmental competence was not enhanced.

  14. Analysis of telomere length in Dolly, a sheep derived by nuclear transfer.

    PubMed

    Shiels, P G; Kind, A J; Campbell, K H; Wilmut, I; Waddington, D; Colman, A; Schnieke, A E

    1999-01-01

    We have used a (TTAGGG) oligonucleotide probe to demonstrate that ovine telomeres are composed of (TTAGGG) repeat arrays and to compare the terminal restriction fragment lengths of sheep derived by natural mating and nuclear transfer. Here we show that ovine somatic telomeres decrease in length with age, and that Dolly, derived by the transfer of 6-year-old adult somatic nucleus, exhibits diminished terminal restriction fragment lengths. The decrease is consistent with the age of the donor tissue and telomere erosion during in vitro culture. Nuclear transfer does not restore telomere lengths. Dolly otherwise appears physiologically and phenotypically normal for her breed and age. We further report on apparent telomere lengthening in sheep, occurring during the first year in naturally derived lambs.

  15. Analysis of telomere length in Dolly, a sheep derived by nuclear transfer.

    PubMed

    Shiels, P G; Kind, A J; Campbell, K H; Wilmut, I; Waddington, D; Colman, A; Schnieke, A E

    1999-01-01

    We have used a (TTAGGG) oligonucleotide probe to demonstrate that ovine telomeres are composed of (TTAGGG) repeat arrays and to compare the terminal restriction fragment lengths of sheep derived by natural mating and nuclear transfer. Here we show that ovine somatic telomeres decrease in length with age, and that Dolly, derived by the transfer of 6-year-old adult somatic nucleus, exhibits diminished terminal restriction fragment lengths. The decrease is consistent with the age of the donor tissue and telomere erosion during in vitro culture. Nuclear transfer does not restore telomere lengths. Dolly otherwise appears physiologically and phenotypically normal for her breed and age. We further report on apparent telomere lengthening in sheep, occurring during the first year in naturally derived lambs. PMID:16218837

  16. High-fidelity transfer and storage of photon states in a single nuclear spin

    NASA Astrophysics Data System (ADS)

    Yang, Sen; Wang, Ya; Rao, D. D. Bhaktavatsala; Hien Tran, Thai; Momenzadeh, Ali S.; Markham, M.; Twitchen, D. J.; Wang, Ping; Yang, Wen; Stöhr, Rainer; Neumann, Philipp; Kosaka, Hideo; Wrachtrup, Jörg

    2016-08-01

    Long-distance quantum communication requires photons and quantum nodes that comprise qubits for interaction with light and good memory capabilities, as well as processing qubits for the storage and manipulation of photons. Owing to the unavoidable photon losses, robust quantum communication over lossy transmission channels requires quantum repeater networks. A necessary and highly demanding prerequisite for these networks is the existence of quantum memories with long coherence times to reliably store the incident photon states. Here we demonstrate the high-fidelity (˜98%) coherent transfer of a photon polarization state to a single solid-state nuclear spin that has a coherence time of over 10 s. The storage process is achieved by coherently transferring the polarization state of a photon to an entangled electron-nuclear spin state of a nitrogen-vacancy centre in diamond. The nuclear spin-based optical quantum memory demonstrated here paves the way towards an absorption-based quantum repeater network.

  17. High-fidelity transfer and storage of photon states in a single nuclear spin

    NASA Astrophysics Data System (ADS)

    Yang, Sen; Wang, Ya; Rao, D. D. Bhaktavatsala; Hien Tran, Thai; Momenzadeh, Ali S.; Markham, M.; Twitchen, D. J.; Wang, Ping; Yang, Wen; Stöhr, Rainer; Neumann, Philipp; Kosaka, Hideo; Wrachtrup, Jörg

    2016-08-01

    Long-distance quantum communication requires photons and quantum nodes that comprise qubits for interaction with light and good memory capabilities, as well as processing qubits for the storage and manipulation of photons. Owing to the unavoidable photon losses, robust quantum communication over lossy transmission channels requires quantum repeater networks. A necessary and highly demanding prerequisite for these networks is the existence of quantum memories with long coherence times to reliably store the incident photon states. Here we demonstrate the high-fidelity (∼98%) coherent transfer of a photon polarization state to a single solid-state nuclear spin that has a coherence time of over 10 s. The storage process is achieved by coherently transferring the polarization state of a photon to an entangled electron–nuclear spin state of a nitrogen–vacancy centre in diamond. The nuclear spin-based optical quantum memory demonstrated here paves the way towards an absorption-based quantum repeater network.

  18. Modularization and nuclear power. Report by the Technology Transfer Modularization Task Team

    SciTech Connect

    Not Available

    1985-06-01

    This report describes the results of the work performed by the Technology Transfer Task Team on Modularization. This work was performed as part of the Technology Transfer work being performed under Department of Energy Contract 54-7WM-335406, between December, 1984 and February, 1985. The purpose of this task team effort was to briefly survey the current use of modularization in the nuclear and non-nuclear industries and to assess and evaluate the techniques available for potential application to nuclear power. A key conclusion of the evaluation was that there was a need for a study to establish guidelines for the future development of Light Water Reactor, High Temperature Gas Reactor and Liquid Metal Reactor plants. The guidelines should identify how modularization can improve construction, maintenance, life extension and decommissioning.

  19. 10 CFR 770.7 - What procedures are to be used to transfer real property at defense nuclear facilities for...

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... 10 Energy 4 2011-01-01 2011-01-01 false What procedures are to be used to transfer real property at defense nuclear facilities for economic development? 770.7 Section 770.7 Energy DEPARTMENT OF ENERGY TRANSFER OF REAL PROPERTY AT DEFENSE NUCLEAR FACILITIES FOR ECONOMIC DEVELOPMENT § 770.7...

  20. 10 CFR 770.7 - What procedures are to be used to transfer real property at defense nuclear facilities for...

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... 10 Energy 4 2012-01-01 2012-01-01 false What procedures are to be used to transfer real property at defense nuclear facilities for economic development? 770.7 Section 770.7 Energy DEPARTMENT OF ENERGY TRANSFER OF REAL PROPERTY AT DEFENSE NUCLEAR FACILITIES FOR ECONOMIC DEVELOPMENT § 770.7...

  1. 10 CFR 770.8 - May DOE transfer real property at defense nuclear facilities for economic development at less...

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... 10 Energy 4 2011-01-01 2011-01-01 false May DOE transfer real property at defense nuclear facilities for economic development at less than fair market value? 770.8 Section 770.8 Energy DEPARTMENT OF ENERGY TRANSFER OF REAL PROPERTY AT DEFENSE NUCLEAR FACILITIES FOR ECONOMIC DEVELOPMENT § 770.8 May...

  2. 10 CFR 770.8 - May DOE transfer real property at defense nuclear facilities for economic development at less...

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... 10 Energy 4 2012-01-01 2012-01-01 false May DOE transfer real property at defense nuclear facilities for economic development at less than fair market value? 770.8 Section 770.8 Energy DEPARTMENT OF ENERGY TRANSFER OF REAL PROPERTY AT DEFENSE NUCLEAR FACILITIES FOR ECONOMIC DEVELOPMENT § 770.8 May...

  3. 10 CFR 770.7 - What procedures are to be used to transfer real property at defense nuclear facilities for...

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... 10 Energy 4 2014-01-01 2014-01-01 false What procedures are to be used to transfer real property at defense nuclear facilities for economic development? 770.7 Section 770.7 Energy DEPARTMENT OF ENERGY TRANSFER OF REAL PROPERTY AT DEFENSE NUCLEAR FACILITIES FOR ECONOMIC DEVELOPMENT § 770.7...

  4. 10 CFR 770.8 - May DOE transfer real property at defense nuclear facilities for economic development at less...

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... 10 Energy 4 2014-01-01 2014-01-01 false May DOE transfer real property at defense nuclear facilities for economic development at less than fair market value? 770.8 Section 770.8 Energy DEPARTMENT OF ENERGY TRANSFER OF REAL PROPERTY AT DEFENSE NUCLEAR FACILITIES FOR ECONOMIC DEVELOPMENT § 770.8 May...

  5. 10 CFR 770.8 - May DOE transfer real property at defense nuclear facilities for economic development at less...

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ... 10 Energy 4 2013-01-01 2013-01-01 false May DOE transfer real property at defense nuclear facilities for economic development at less than fair market value? 770.8 Section 770.8 Energy DEPARTMENT OF ENERGY TRANSFER OF REAL PROPERTY AT DEFENSE NUCLEAR FACILITIES FOR ECONOMIC DEVELOPMENT § 770.8 May...

  6. 10 CFR 770.8 - May DOE transfer real property at defense nuclear facilities for economic development at less...

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... 10 Energy 4 2010-01-01 2010-01-01 false May DOE transfer real property at defense nuclear facilities for economic development at less than fair market value? 770.8 Section 770.8 Energy DEPARTMENT OF ENERGY TRANSFER OF REAL PROPERTY AT DEFENSE NUCLEAR FACILITIES FOR ECONOMIC DEVELOPMENT § 770.8 May...

  7. Model ecosystems with random nonlinear interspecies interactions.

    PubMed

    Santos, Danielle O C; Fontanari, José F

    2004-12-01

    The principle of competitive exclusion in ecology establishes that two species living together cannot occupy the same ecological niche. Here we present a model ecosystem in which the species are described by a series of phenotypic characters and the strength of the competition between two species is given by a nondecreasing (modulating) function of the number of common characters. Using analytical tools of statistical mechanics we find that the ecosystem diversity, defined as the fraction of species that coexist at equilibrium, decreases as the complexity (i.e., number of characters) of the species increases, regardless of the modulating function. By considering both selective and random elimination of the links in the community web, we show that ecosystems composed of simple species are more robust than those composed of complex species. In addition, we show that the puzzling result that there exists either rich or poor ecosystems for a linear modulating function is not typical of communities in which the interspecies interactions are determined by a complementarity rule.

  8. Nuclear and microtubule remodeling and in vitro development of nuclear transferred cat oocytes with skin fibroblasts of the domestic cat (Felis silvestris catus) and leopard cat (Prionailurus bengalensis).

    PubMed

    Yin, X J; Lee, Y H; Jin, J Y; Kim, N H; Kong, I K

    2006-10-01

    The leopard cat (Prionailurus bengalensis), a member of the felidae family, is a threatened animal in South Korea. In terms of protecting endangered felids, nuclear transfer (NT) is a potentially valuable technique for assuring the continuation of species with dwindling numbers. In the present experiment, nuclear and microtubule remodeling and the in vitro developmental potential of enucleated domestic cat oocytes reconstructed with nuclei of somatic cells from either domestic cat fibroblast (DCF) or leopard cat fibroblast (LCF) were evaluated. Microtubule aster is allocated to de-condensed chromatin following nuclear transfer (3h after activation) of fibroblast cells from both domestic and leopard cats, suggesting the introduction of a somatic cell centrosome. The transferred fibroblast nuclei formed a large, swollen, pronuclear-like structure in most reconstructed oocytes, in the cat or leopard cat. At 18h following nuclear transfer, mitosis occurred, and according to the photo (F) it appears that spindle microtubules and two asters were observed. The percentages of blastocyst formation from nuclear transfer embryos derived from domestic cat fibroblasts (4/46, 8.6%) were not significantly different than those for nuclear transfer embryos constructed with leopard cat fibroblasts (4/52, 7.6%). These results indicate that nuclear and microtubule remodeling processes and in vitro developmental ability are similar in reconstructed cat oocytes following transfer of nuclei from either domestic or leopard cats. PMID:16310987

  9. Interspecies correlations of toxicity to eight aquatic organisms: theoretical considerations.

    PubMed

    Zhang, Xu J; Qin, Hong W; Su, Li M; Qin, Wei C; Zou, Ming Y; Sheng, Lian X; Zhao, Yuan H; Abraham, Michael H

    2010-09-15

    Interspecies correlations allow the prediction of toxicity to a number of other species. However, little attention has been paid to the theoretical considerations of the interspecies relationship based on the differences of bio-uptake and toxic mechanism between species. This study examines the interspecies correlations of toxicity between species of Vibrio fischeri, river bacteria, algae, Daphnia magna, carp, Tetrahymena pyriformis, fathead minnow and guppy based on the theoretical background. The results show that there are good interspecies correlations between marine bacterium and fresh water bacteria or fish and fish. It is suggested that compounds share the same bio-uptake and toxic mechanism of action between the species. On the other hand, poor interspecies relationships were found between toxicities to algae and T. pyriformis or D. magna. It is suggested that compounds have different toxic mechanisms of action between these species. Interspecies relationships can be improved by inclusion of the octanol/water partition coefficient or the energy of the lowest unoccupied molecular orbital. They reflect the difference of bio-uptake or toxic mechanism of action between species for organic compounds. Benzoic acids show very different toxicity contributions to the three species, V. fischeri, D. magna and carp. They can be easily absorbed into the unicellular bacteria, V. fischeri. On the contrary, the skin and lipid content of multicellular organisms, such as D. magna and fish, can strongly inhibit the bio-uptake for ionizable compounds, which results in the different toxic effect between V. fischeri and D. magna or carp. Good correlation coefficients were observed between toxicities to V. fischeri and D. magna or fishes by inclusion of hydrophobic and ionization parameters. V. fischeri or D. magna can serve as a surrogate of fish toxicity for hydrophobic and ionizable compounds studied. Toxic mechanisms of action are discussed based on the theoretical background

  10. 10 CFR 770.7 - What procedures are to be used to transfer real property at defense nuclear facilities for...

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ... 10 Energy 4 2013-01-01 2013-01-01 false What procedures are to be used to transfer real property... ENERGY TRANSFER OF REAL PROPERTY AT DEFENSE NUCLEAR FACILITIES FOR ECONOMIC DEVELOPMENT § 770.7 What... congressional defense committees through the Secretary of Energy. (d) Transfer. After the...

  11. 10 CFR 770.7 - What procedures are to be used to transfer real property at defense nuclear facilities for...

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... 10 Energy 4 2010-01-01 2010-01-01 false What procedures are to be used to transfer real property... ENERGY TRANSFER OF REAL PROPERTY AT DEFENSE NUCLEAR FACILITIES FOR ECONOMIC DEVELOPMENT § 770.7 What... congressional defense committees through the Secretary of Energy. (d) Transfer. After the...

  12. Significant improvement of mouse cloning technique by treatment with trichostatin A after somatic nuclear transfer

    SciTech Connect

    Kishigami, Satoshi . E-mail: kishigami@cdb.riken.jp; Mizutani, Eiji; Ohta, Hiroshi; Hikichi, Takafusa; Thuan, Nguyen Van; Wakayama, Sayaka; Bui, Hong-Thuy; Wakayama, Teruhiko

    2006-02-03

    The low success rate of animal cloning by somatic cell nuclear transfer (SCNT) is believed to be associated with epigenetic errors including abnormal DNA hypermethylation. Recently, we elucidated by using round spermatids that, after nuclear transfer, treatment of zygotes with trichostatin A (TSA), an inhibitor of histone deacetylase, can remarkably reduce abnormal DNA hypermethylation depending on the origins of transferred nuclei and their genomic regions [S. Kishigami, N. Van Thuan, T. Hikichi, H. Ohta, S. Wakayama. E. Mizutani, T. Wakayama, Epigenetic abnormalities of the mouse paternal zygotic genome associated with microinsemination of round spermatids, Dev. Biol. (2005) in press]. Here, we found that 5-50 nM TSA-treatment for 10 h following oocyte activation resulted in more efficient in vitro development of somatic cloned embryos to the blastocyst stage from 2- to 5-fold depending on the donor cells including tail tip cells, spleen cells, neural stem cells, and cumulus cells. This TSA-treatment also led to more than 5-fold increase in success rate of mouse cloning from cumulus cells without obvious abnormality but failed to improve ES cloning success. Further, we succeeded in establishment of nuclear transfer-embryonic stem (NT-ES) cells from TSA-treated cloned blastocyst at a rate three times higher than those from untreated cloned blastocysts. Thus, our data indicate that TSA-treatment after SCNT in mice can dramatically improve the practical application of current cloning techniques.

  13. Coronavirus diversity, phylogeny and interspecies jumping.

    PubMed

    Woo, Patrick C Y; Lau, Susanna K P; Huang, Yi; Yuen, Kwok-Yung

    2009-10-01

    The SARS epidemic has boosted interest in research on coronavirus biodiversity and genomics. Before 2003, there were only 10 coronaviruses with complete genomes available. After the SARS epidemic, up to December 2008, there was an addition of 16 coronaviruses with complete genomes sequenced. These include two human coronaviruses (human coronavirus NL63 and human coronavirus HKU1), 10 other mammalian coronaviruses [bat SARS coronavirus, bat coronavirus (bat-CoV) HKU2, bat-CoV HKU4, bat-CoV HKU5, bat-CoV HKU8, bat-CoV HKU9, bat-CoV 512/2005, bat-CoV 1A, equine coronavirus, and beluga whale coronavirus] and four avian coronaviruses (turkey coronavirus, bulbul coronavirus HKU11, thrush coronavirus HKU12, and munia coronavirus HKU13). Two novel subgroups in group 2 coronavirus (groups 2c and 2d) and two novel subgroups in group 3 coronavirus (groups 3b and 3c) have been proposed. The diversity of coronaviruses is a result of the infidelity of RNA-dependent RNA polymerase, high frequency of homologous RNA recombination, and the large genomes of coronaviruses. Among all hosts, the diversity of coronaviruses is most evidenced in bats and birds, which may be a result of their species diversity, ability to fly, environmental pressures, and habits of roosting and flocking. The present evidence supports that bat coronaviruses are the gene pools of group 1 and 2 coronaviruses, whereas bird coronaviruses are the gene pools of group 3 coronaviruses. With the increasing number of coronaviruses, more and more closely related coronaviruses from distantly related animals have been observed, which were results of recent interspecies jumping and may be the cause of disastrous outbreaks of zoonotic diseases. PMID:19546349

  14. An improved heat transfer configuration for a solid-core nuclear thermal rocket engine

    SciTech Connect

    Clark, J.S.; Walton, J.T.; Mcguire, M.L. Cincinnati, University, OH )

    1992-07-01

    Interrupted flow, impingement cooling, and axial power distribution are employed to enhance the heat-transfer configuration of a solid-core nuclear thermal rocket engine. Impingement cooling is introduced to increase the local heat-transfer coefficients between the reactor material and the coolants. Increased fuel loading is used at the inlet end of the reactor to enhance heat-transfer capability where the temperature differences are the greatest. A thermal-hydraulics computer program for an unfueled NERVA reactor core is employed to analyze the proposed configuration with attention given to uniform fuel loading, number of channels through the impingement wafers, fuel-element length, mass-flow rate, and wafer gap. The impingement wafer concept (IWC) is shown to have heat-transfer characteristics that are better than those of the NERVA-derived reactor at 2500 K. The IWC concept is argued to be an effective heat-transfer configuration for solid-core nuclear thermal rocket engines. 11 refs.

  15. Production of nuclear transfer-derived swine that express the enhanced green fluorescent protein.

    PubMed

    Park, K W; Cheong, H T; Lai, L; Im, G S; Kühholzer, B; Bonk, A; Samuel, M; Rieke, A; Day, B N; Murphy, C N; Carter, D B; Prather, R S

    2001-11-01

    The ability to add or delete specific genes in swine will likely provide considerable benefits not just to agriculture but also to medicine, where pigs have potential as models for human disease and as organ donors. Here we have transferred nuclei from a genetically modified fibroblast cell line to porcine oocytes, matured in vitro under defined culture conditions, to create piglets expressing enhanced green fluorescent protein. The nuclear transfer-derived piglets were of normal size, although some mild symptoms of "large offspring syndrome" were evident. These experiments represent a next step towards creating swine with more useful genetic modifications. PMID:11808633

  16. Nuclear-mitochondrial incompatibility in interorder rhesus monkey-cow embryos derived from somatic cell nuclear transfer.

    PubMed

    Kwon, Daekee; Koo, Ok-Jae; Kim, Min-Jung; Jang, Goo; Lee, Byeong Chun

    2016-10-01

    Monkey interorder somatic cell nuclear transfer (iSCNT) using enucleated cow oocytes yielded poor blastocysts development and contradictory results among research groups. Determining the reason for this low blastocyst development is a prerequisite for optimizing iSCNT in rhesus monkeys. The aim of this study was to elucidate nuclear-mitochondrial incompatibility of rhesus monkey-cow iSCNT embryos and its relationship to low blastocyst development. Cytochrome b is a protein of complex III of the electron transport chain (ETC). According to meta-analysis of amino acid sequences, the homology of cytochrome b is 75 % between rhesus monkeys and cattle. To maintain the function of ETC after iSCNT, 4n iSCNT embryos were produced by fusion of non-enucleated cow oocytes and rhesus monkey somatic cells. The blastocyst development rate of 4n iSCNT embryos was higher than that of 2n embryos (P < 0.01). Formation of reactive oxygen species (ROS) is an indirect indicator of ETC activity of cells. The ROS levels of 4n iSCNT embryos was higher than that of 2n embryos (P < 0.01). Collectively, rhesus monkey iSCNT embryos reconstructed with cow oocytes have nuclear-mitochondrial incompatibility due to fundamental species differences between rhesus monkeys and cattle. Nuclear-mitochondrial incompatibility seems to correlate with low ETC activity and extremely low blastocyst development of rhesus monkey-cow iSCNT embryos.

  17. Cloning mice and ES cells by nuclear transfer from somatic stem cells and fully differentiated cells.

    PubMed

    Wang, Zhongde

    2011-01-01

    Cloning animals by nuclear transfer (NT) has been successful in several mammalian species. In addition to cloning live animals (reproductive cloning), this technique has also been used in several species to establish cloned embryonic stem (ntES) cell lines from somatic cells. It is the latter application of this technique that has been heralded as being the potential means to produce isogenic embryonic stem cells from patients for cell therapy (therapeutic cloning). These two types of cloning differ only in the steps after cloned embryos are produced: for reproductive cloning the cloned embryos are transferred to surrogate mothers to allow them to develop to full term and for therapeutic cloning the cloned embryos are used to derive ntES cells. In this chapter, a detailed NT protocol in mouse by using somatic stem cells (neuron and skin stem cells) and fully differentiated somatic cells (cumulus cells and fibroblast cells) as nuclear donors is described.

  18. Treatment of donor cell/embryo with different approaches to improve development after nuclear transfer.

    PubMed

    Mizutani, Eiji; Wakayama, Sayaka; Wakayama, Teruhiko

    2015-01-01

    The successful production of cloned animals by somatic cell nuclear transfer (SCNT) is a promising technology with many potential applications in basic research, medicine, and agriculture. However, the low efficiency and the difficulty of cloning are major obstacles to the widespread use of this technology. Since the first mammal cloned from an adult donor cell was born, many attempts have been made to improve animal cloning techniques, and some approaches have successfully improved its efficiency. Nuclear transfer itself is still difficult because it requires an accomplished operator with a practiced technique. Thus, it is very important to find simple and reproducible methods for improving the success rate of SCNT. In this chapter, we will review our recent protocols, which seem to be the simplest and most reliable method to date to improve development of SCNT embryos.

  19. Effects of donor cells' sex on nuclear transfer efficiency and telomere lengths of cloned goats.

    PubMed

    Liu, H-J; Peng, H; Hu, C-C; Li, X-Y; Zhang, J-L; Zheng, Z; Zhang, W-C

    2016-10-01

    The aim of this study was to investigate the effects of donor cells' sex on nuclear transfer efficiency and telomere length of cloned goats from adult skin fibroblast cells. The telomere length of somatic cell cloned goats and their offspring was determined by measuring their mean terminal restriction fragment (TRF) length. The result showed that (i) reconstructed embryos with fibroblast cells from males Boer goats obtained significantly higher kids rate and rate of live kids than those of female embryos and (ii) the telomere lengths of four female cloned goats were shorter compared to their donor cells, but five male cloned goats had the same telomere length with their donor cells, mainly due to great variation existed among them. The offspring from female cloned goats had the same telomere length with their age-matched counterparts. In conclusion, the donor cells' sex had significant effects on nuclear transfer efficiency and telomere lengths of cloned goats.

  20. Effects of donor cells' sex on nuclear transfer efficiency and telomere lengths of cloned goats.

    PubMed

    Liu, H-J; Peng, H; Hu, C-C; Li, X-Y; Zhang, J-L; Zheng, Z; Zhang, W-C

    2016-10-01

    The aim of this study was to investigate the effects of donor cells' sex on nuclear transfer efficiency and telomere length of cloned goats from adult skin fibroblast cells. The telomere length of somatic cell cloned goats and their offspring was determined by measuring their mean terminal restriction fragment (TRF) length. The result showed that (i) reconstructed embryos with fibroblast cells from males Boer goats obtained significantly higher kids rate and rate of live kids than those of female embryos and (ii) the telomere lengths of four female cloned goats were shorter compared to their donor cells, but five male cloned goats had the same telomere length with their donor cells, mainly due to great variation existed among them. The offspring from female cloned goats had the same telomere length with their age-matched counterparts. In conclusion, the donor cells' sex had significant effects on nuclear transfer efficiency and telomere lengths of cloned goats. PMID:27558653

  1. Neutron Transfer Reactions: Surrogates for Neutron Capture for Basic and Applied Nuclear Science

    NASA Astrophysics Data System (ADS)

    Cizewski, J. A.; Jones, K. L.; Kozub, R. L.; Pain, S. D.; Peters, W. A.; Adekola, A.; Allen, J.; Bardayan, D. W.; Becker, J. A.; Blackmon, J. C.; Chae, K. Y.; Chipps, K. A.; Erikson, L.; Gaddis, A.; Harlin, C.; Hatarik, R.; Howard, J.; Jandel, M.; Johnson, M. S.; Kapler, R.; Krolas, W.; Liang, F.; Livesay, R. J.; Ma, Z.; Matei, C.; Matthews, C.; Moazen, B.; Nesaraja, C. D.; O'Malley, P.; Patterson, N.; Paulauskas, S. V.; Pelham, T.; Pittman, S. T.; Radford, D.; Rogers, J.; Schmitt, K.; Shapira, D.; Shriner, J. F.; Sissom, D. J.; Smith, M. S.; Swan, T.; Thomas, J. S.; Vieira, D. J.; Wilhelmy, J. B.; Wilson, G. L.

    2009-03-01

    Neutron capture reactions on unstable nuclei are important for both basic and applied nuclear science. A program has been developed at the Holifield Radioactive Ion Beam Facility at Oak Ridge National Laboratory to study single-neutron transfer (d,p) reactions with rare isotope beams to provide information on neutron-induced reactions on unstable nuclei. Results from (d,p) studies on 130,132Sn, 134Te and 75As are discussed.

  2. Ion engine propelled Earth-Mars cycler with nuclear thermal propelled transfer vehicle, volume 2

    NASA Technical Reports Server (NTRS)

    Meyer, Rudolf X.; Baker, Myles; Melko, Joseph

    1994-01-01

    The goal of this project was to perform a preliminary design of a long term, reusable transportation system between earth and Mars which would be capable of providing both artificial gravity and shelter from solar flare radiation. The heart of this system was assumed to be a Cycler spacecraft propelled by an ion propulsion system. The crew transfer vehicle was designed to be propelled by a nuclear-thermal propulsion system. Several Mars transportation system architectures and their associated space vehicles were designed.

  3. Neutron transfer reactions: Surrogates for neutron capture for basic and applied nuclear science

    SciTech Connect

    Cizewski, J. A.; Jones, K. L.; Kozub, R. L.; Pain, Steven D; Peters, W. A.; Adekola, Aderemi S; Allen, J.; Bardayan, Daniel W; Becker, J.; Blackmon, Jeff C; Chae, K. Y.; Chipps, K.; Erikson, Luke; Gaddis, A. L.; Harlin, Christopher W; Hatarik, Robert; Howard, Joshua A; Jandel, M.; Johnson, Micah; Kapler, R.; Krolas, W.; Liang, J Felix; Livesay, Jake; Ma, Zhanwen; Matei, Catalin; Matthews, C.; Moazen, Brian; Nesaraja, Caroline D; O'Malley, Patrick; Patterson, N. P.; Paulauskas, Stanley; Pelham, T.; Pittman, S. T.; Radford, David C; Rogers, J.; Schmitt, Kyle; Shapira, Dan; ShrinerJr., J. F.; Sissom, D. J.; Smith, Michael Scott; Swan, T. P.; Thomas, J. S.; Vieira, D. J.; Wilhelmy, J. B.; Wilson, Gemma L

    2009-04-01

    Neutron capture reactions on unstable nuclei are important for both basic and applied nuclear science. A program has been developed at the Holifield Radioactive Ion Beam Facility at Oak Ridge National Laboratory to study single-neutron transfer (d,p) reactions with rare isotope beams to provide information on neutron-induced reactions on unstable nuclei. Results from (d,p) studies on {sup 130,132}Sn, {sup 134}Te and {sup 75}As are discussed.

  4. Effects of nuclear transfer procedures on ES cell cloning efficiency in the mouse.

    PubMed

    Yabuuchi, Akiko; Yasuda, Yoshiko; Kato, Yoko; Tsunoda, Yukio

    2004-04-01

    Enucleated oocytes receiving mouse embryonic stem (ES) cells develop into fertile young. The developmental potential to young is low, however, and the rate of postnatal death is high. We examined the effect of various nuclear transfer procedures on the in vitro and in vivo developmental potential of nuclear-transferred oocytes. The potential of oocytes receiving ES cells at M phase to develop into blastocysts after fusion by Sendai virus was high compared with that after direct injection (67% vs. 30%). The developmental potential of oocytes receiving ES cells at the M phase is higher than that of oocytes receiving ES cells at the G(1) phase (30-67% vs. 2-5%). Developmental ability to live young was low in all groups (0-4%). Different activation protocols affected the potential to develop into blastocysts to a different extent (27-62%), but did not affect the potential to develop into live young (0-3%). The present study demonstrated that the various conditions examined did not affect the potential of nuclear-transferred oocytes receiving ES cells to develop into live young or the incidence of postnatal death.

  5. Somatic cell nuclear transfer: origins, the present position and future opportunities.

    PubMed

    Wilmut, Ian; Bai, Yu; Taylor, Jane

    2015-10-19

    Nuclear transfer that involves the transfer of the nucleus from a donor cell into an oocyte or early embryo from which the chromosomes have been removed was considered first as a means of assessing changes during development in the ability of the nucleus to control development. In mammals, development of embryos produced by nuclear transfer depends upon coordination of the cell cycles of donor and recipient cells. Our analysis of nuclear potential was completed in 1996 when a nucleus from an adult ewe mammary gland cell controlled development to term of Dolly the sheep. The new procedure has been used to target the first precise genetic modification into livestock; however, the greatest inheritance of the Dolly experiment was to make biologists think differently. If unknown factors in the recipient oocyte could reprogramme the nucleus to a stage very early in development then there must be other ways of making that change. Within 10 years, two laboratories working independently established protocols by which the introduction of selected transcription factors changes a small proportion of the treated cells to pluripotent stem cells. This ability to produce 'induced pluripotent stem cells' is providing revolutionary new opportunities in research and cell therapy.

  6. Production of human CD59-transgenic pigs by embryonic germ cell nuclear transfer

    SciTech Connect

    Ahn, Kwang Sung; Won, Ji Young; Park, Jin-Ki; Sorrell, Alice M.; Heo, Soon Young; Kang, Jee Hyun; Woo, Jae-Seok; Choi, Bong-Hwan; Chang, Won-Kyong; Shim, Hosup

    2010-10-01

    Research highlights: {yields} Human CD59 (hCD59) gene was introduced into porcine embryonic germ (EG) cells. {yields} hCD59-transgenic EG cells were resistant to hyperacute rejection in cytolytic assay. {yields} hCD59-transgenic pigs were produced by EG cell nuclear transfer. -- Abstract: This study was performed to produce transgenic pigs expressing the human complement regulatory protein CD59 (hCD59) using the nuclear transfer (NT) of embryonic germ (EG) cells, which are undifferentiated stem cells derived from primordial germ cells. Because EG cells can be cultured indefinitely in an undifferentiated state, they may provide an inexhaustible source of nuclear donor cells for NT to produce transgenic pigs. A total of 1980 NT embryos derived from hCD59-transgenic EG cells were transferred to ten recipients, resulting in the birth of fifteen piglets from three pregnancies. Among these offspring, ten were alive without overt health problems. Based on PCR analysis, all fifteen piglets were confirmed as hCD59 transgenic. The expression of the hCD59 transgene in the ten living piglets was verified by RT-PCR. Western analysis showed the expression of the hCD59 protein in four of the ten RT-PCR-positive piglets. These results demonstrate that hCD59-transgenic pigs could effectively be produced by EG cell NT and that such transgenic pigs may be used as organ donors in pig-to-human xenotransplantation.

  7. Somatic cell nuclear transfer: origins, the present position and future opportunities

    PubMed Central

    Wilmut, Ian; Bai, Yu; Taylor, Jane

    2015-01-01

    Nuclear transfer that involves the transfer of the nucleus from a donor cell into an oocyte or early embryo from which the chromosomes have been removed was considered first as a means of assessing changes during development in the ability of the nucleus to control development. In mammals, development of embryos produced by nuclear transfer depends upon coordination of the cell cycles of donor and recipient cells. Our analysis of nuclear potential was completed in 1996 when a nucleus from an adult ewe mammary gland cell controlled development to term of Dolly the sheep. The new procedure has been used to target the first precise genetic modification into livestock; however, the greatest inheritance of the Dolly experiment was to make biologists think differently. If unknown factors in the recipient oocyte could reprogramme the nucleus to a stage very early in development then there must be other ways of making that change. Within 10 years, two laboratories working independently established protocols by which the introduction of selected transcription factors changes a small proportion of the treated cells to pluripotent stem cells. This ability to produce ‘induced pluripotent stem cells’ is providing revolutionary new opportunities in research and cell therapy. PMID:26416677

  8. Somatic cell nuclear transfer: origins, the present position and future opportunities.

    PubMed

    Wilmut, Ian; Bai, Yu; Taylor, Jane

    2015-10-19

    Nuclear transfer that involves the transfer of the nucleus from a donor cell into an oocyte or early embryo from which the chromosomes have been removed was considered first as a means of assessing changes during development in the ability of the nucleus to control development. In mammals, development of embryos produced by nuclear transfer depends upon coordination of the cell cycles of donor and recipient cells. Our analysis of nuclear potential was completed in 1996 when a nucleus from an adult ewe mammary gland cell controlled development to term of Dolly the sheep. The new procedure has been used to target the first precise genetic modification into livestock; however, the greatest inheritance of the Dolly experiment was to make biologists think differently. If unknown factors in the recipient oocyte could reprogramme the nucleus to a stage very early in development then there must be other ways of making that change. Within 10 years, two laboratories working independently established protocols by which the introduction of selected transcription factors changes a small proportion of the treated cells to pluripotent stem cells. This ability to produce 'induced pluripotent stem cells' is providing revolutionary new opportunities in research and cell therapy. PMID:26416677

  9. Modeling of the heat transfer performance of plate-type dispersion nuclear fuel elements

    NASA Astrophysics Data System (ADS)

    Ding, Shurong; Huo, Yongzhong; Yan, XiaoQing

    2009-08-01

    Considering the mutual actions between fuel particles and the metal matrix, the three-dimensional finite element models are developed to simulate the heat transfer behaviors of dispersion nuclear fuel plates. The research results indicate that the temperatures of the fuel plate might rise more distinctly with considering the particle swelling and the degraded surface heat transfer coefficients with increasing burnup; the local heating phenomenon within the particles appears when their thermal conductivities are too low. With rise of the surface heat transfer coefficients, the temperatures within the fuel plate decrease; the temperatures of the fuel plate are sensitive to the variations of the heat transfer coefficients whose values are lower, but their effects are weakened and slight when the heat transfer coefficients increase and reach a certain extent. Increasing the heat generation rate leads to elevating the internal temperatures. The temperatures and the maximum temperature differences within the plate increase along with the particle volume fractions. The surface thermal flux goes up along with particle volume fractions and heat generation rates, but the effects of surface heat transfer coefficients are not evident.

  10. Artificial intelligence and nuclear power. Report by the Technology Transfer Artificial Intelligence Task Team

    SciTech Connect

    Not Available

    1985-06-01

    The Artificial Intelligence Task Team was organized to review the status of Artificial Intelligence (AI) technology, identify guidelines for AI work, and to identify work required to allow the nuclear industry to realize maximum benefit from this technology. The state of the nuclear industry was analyzed to determine where the application of AI technology could be of greatest benefit. Guidelines and criteria were established to focus on those particular problem areas where AI could provide the highest possible payoff to the industry. Information was collected from government, academic, and private organizations. Very little AI work is now being done to specifically support the nuclear industry. The AI Task Team determined that the establishment of a Strategic Automation Initiative (SAI) and the expansion of the DOE Technology Transfer program would ensure that AI technology could be used to develop software for the nuclear industry that would have substantial financial payoff to the industry. The SAI includes both long and short term phases. The short-term phase includes projects which would demonstrate that AI can be applied to the nuclear industry safely, and with substantial financial benefit. The long term phase includes projects which would develop AI technologies with specific applicability to the nuclear industry that would not be developed by people working in any other industry.

  11. Interspecies interactions are an integral determinant of microbial community dynamics.

    PubMed

    Aziz, Fatma A A; Suzuki, Kenshi; Ohtaki, Akihiro; Sagegami, Keita; Hirai, Hidetaka; Seno, Jun; Mizuno, Naoko; Inuzuka, Yuma; Saito, Yasuhisa; Tashiro, Yosuke; Hiraishi, Akira; Futamata, Hiroyuki

    2015-01-01

    This study investigated the factors that determine the dynamics of bacterial communities in a complex system using multidisciplinary methods. Since natural and engineered microbial ecosystems are too complex to study, six types of synthetic microbial ecosystems (SMEs) were constructed under chemostat conditions with phenol as the sole carbon and energy source. Two to four phenol-degrading, phylogenetically and physiologically different bacterial strains were used in each SME. Phylogeny was based on the nucleotide sequence of 16S rRNA genes, while physiologic traits were based on kinetic and growth parameters on phenol. Two indices, J parameter and "interspecies interaction," were compared to predict which strain would become dominant in an SME. The J parameter was calculated from kinetic and growth parameters. On the other hand, "interspecies interaction," a new index proposed in this study, was evaluated by measuring the specific growth activity, which was determined on the basis of relative growth of a strain with or without the supernatant prepared from other bacterial cultures. Population densities of strains used in SMEs were enumerated by real-time quantitative PCR (qPCR) targeting the gene encoding the large subunit of phenol hydroxylase and were compared to predictions made from J parameter and interspecies interaction calculations. In 4 of 6 SEMs tested the final dominant strain shown by real-time qPCR analyses coincided with the strain predicted by both the J parameter and the interspecies interaction. However, in SMEII-2 and SMEII-3 the final dominant Variovorax strains coincided with prediction of the interspecies interaction but not the J parameter. These results demonstrate that the effects of interspecies interactions within microbial communities contribute to determining the dynamics of the microbial ecosystem.

  12. Interspecies interactions are an integral determinant of microbial community dynamics.

    PubMed

    Aziz, Fatma A A; Suzuki, Kenshi; Ohtaki, Akihiro; Sagegami, Keita; Hirai, Hidetaka; Seno, Jun; Mizuno, Naoko; Inuzuka, Yuma; Saito, Yasuhisa; Tashiro, Yosuke; Hiraishi, Akira; Futamata, Hiroyuki

    2015-01-01

    This study investigated the factors that determine the dynamics of bacterial communities in a complex system using multidisciplinary methods. Since natural and engineered microbial ecosystems are too complex to study, six types of synthetic microbial ecosystems (SMEs) were constructed under chemostat conditions with phenol as the sole carbon and energy source. Two to four phenol-degrading, phylogenetically and physiologically different bacterial strains were used in each SME. Phylogeny was based on the nucleotide sequence of 16S rRNA genes, while physiologic traits were based on kinetic and growth parameters on phenol. Two indices, J parameter and "interspecies interaction," were compared to predict which strain would become dominant in an SME. The J parameter was calculated from kinetic and growth parameters. On the other hand, "interspecies interaction," a new index proposed in this study, was evaluated by measuring the specific growth activity, which was determined on the basis of relative growth of a strain with or without the supernatant prepared from other bacterial cultures. Population densities of strains used in SMEs were enumerated by real-time quantitative PCR (qPCR) targeting the gene encoding the large subunit of phenol hydroxylase and were compared to predictions made from J parameter and interspecies interaction calculations. In 4 of 6 SEMs tested the final dominant strain shown by real-time qPCR analyses coincided with the strain predicted by both the J parameter and the interspecies interaction. However, in SMEII-2 and SMEII-3 the final dominant Variovorax strains coincided with prediction of the interspecies interaction but not the J parameter. These results demonstrate that the effects of interspecies interactions within microbial communities contribute to determining the dynamics of the microbial ecosystem. PMID:26539177

  13. Interspecies interactions are an integral determinant of microbial community dynamics

    PubMed Central

    Aziz, Fatma A. A.; Suzuki, Kenshi; Ohtaki, Akihiro; Sagegami, Keita; Hirai, Hidetaka; Seno, Jun; Mizuno, Naoko; Inuzuka, Yuma; Saito, Yasuhisa; Tashiro, Yosuke; Hiraishi, Akira; Futamata, Hiroyuki

    2015-01-01

    This study investigated the factors that determine the dynamics of bacterial communities in a complex system using multidisciplinary methods. Since natural and engineered microbial ecosystems are too complex to study, six types of synthetic microbial ecosystems (SMEs) were constructed under chemostat conditions with phenol as the sole carbon and energy source. Two to four phenol-degrading, phylogenetically and physiologically different bacterial strains were used in each SME. Phylogeny was based on the nucleotide sequence of 16S rRNA genes, while physiologic traits were based on kinetic and growth parameters on phenol. Two indices, J parameter and “interspecies interaction,” were compared to predict which strain would become dominant in an SME. The J parameter was calculated from kinetic and growth parameters. On the other hand, “interspecies interaction,” a new index proposed in this study, was evaluated by measuring the specific growth activity, which was determined on the basis of relative growth of a strain with or without the supernatant prepared from other bacterial cultures. Population densities of strains used in SMEs were enumerated by real-time quantitative PCR (qPCR) targeting the gene encoding the large subunit of phenol hydroxylase and were compared to predictions made from J parameter and interspecies interaction calculations. In 4 of 6 SEMs tested the final dominant strain shown by real-time qPCR analyses coincided with the strain predicted by both the J parameter and the interspecies interaction. However, in SMEII-2 and SMEII-3 the final dominant Variovorax strains coincided with prediction of the interspecies interaction but not the J parameter. These results demonstrate that the effects of interspecies interactions within microbial communities contribute to determining the dynamics of the microbial ecosystem. PMID:26539177

  14. Reprogramming within hours following nuclear transfer into mouse but not human zygotes.

    PubMed

    Egli, Dieter; Chen, Alice E; Saphier, Genevieve; Ichida, Justin; Fitzgerald, Claire; Go, Kathryn J; Acevedo, Nicole; Patel, Jay; Baetscher, Manfred; Kearns, William G; Goland, Robin; Leibel, Rudolph L; Melton, Douglas A; Eggan, Kevin

    2011-01-01

    Fertilized mouse zygotes can reprogram somatic cells to a pluripotent state. Human zygotes might therefore be useful for producing patient-derived pluripotent stem cells. However, logistical, legal and social considerations have limited the availability of human eggs for research. Here we show that a significant number of normal fertilized eggs (zygotes) can be obtained for reprogramming studies. Using these zygotes, we found that when the zygotic genome was replaced with that of a somatic cell, development progressed normally throughout the cleavage stages, but then arrested before the morula stage. This arrest was associated with a failure to activate transcription in the transferred somatic genome. In contrast to human zygotes, mouse zygotes reprogrammed the somatic cell genome to a pluripotent state within hours after transfer. Our results suggest that there may be a previously unappreciated barrier to successful human nuclear transfer, and that future studies could focus on the requirements for genome activation. PMID:21971503

  15. PRESTO polarization transfer to quadrupolar nuclei: Implications for dynamic nuclear polarization

    DOE PAGESBeta

    Perras, Frederic A.; Kobayashi, Takeshi; Pruski, Marek

    2015-08-04

    In this study, we show both experimentally and numerically on a series of model systems that in experiments involving transfer of magnetization from 1H to the quadrupolar nuclei under magic-angle-spinning (MAS), the PRESTO technique consistently outperforms traditionally used cross polarization (CP), affording more quantitative intensities, improved lineshapes, better overall sensitivity, and straightforward optimization. This advantage derives from the fact that PRESTO circumvents the convoluted and uncooperative spin dynamics during the CP transfer under MAS, by replacing the spin-locking of quadrupolar nuclei with a single central transition selective 90° pulse and using a symmetry-based recoupling sequence in the 1H channel. Thismore » is important in the context of dynamic nuclear polarization (DNP) NMR of quadrupolar nuclei, where the efficient transfer of enhanced 1H polarization is desired to obtain the highest sensitivity.« less

  16. PRESTO polarization transfer to quadrupolar nuclei: Implications for dynamic nuclear polarization

    SciTech Connect

    Perras, Frederic A.; Kobayashi, Takeshi; Pruski, Marek

    2015-08-04

    In this study, we show both experimentally and numerically on a series of model systems that in experiments involving transfer of magnetization from 1H to the quadrupolar nuclei under magic-angle-spinning (MAS), the PRESTO technique consistently outperforms traditionally used cross polarization (CP), affording more quantitative intensities, improved lineshapes, better overall sensitivity, and straightforward optimization. This advantage derives from the fact that PRESTO circumvents the convoluted and uncooperative spin dynamics during the CP transfer under MAS, by replacing the spin-locking of quadrupolar nuclei with a single central transition selective 90° pulse and using a symmetry-based recoupling sequence in the 1H channel. This is important in the context of dynamic nuclear polarization (DNP) NMR of quadrupolar nuclei, where the efficient transfer of enhanced 1H polarization is desired to obtain the highest sensitivity.

  17. Epigenetic reprogramming in embryonic and foetal development upon somatic cell nuclear transfer cloning.

    PubMed

    Niemann, Heiner; Tian, X Cindy; King, W Allan; Lee, Rita S F

    2008-02-01

    The birth of 'Dolly', the first mammal cloned from an adult donor cell, has sparked a flurry of research activities to improve cloning technology and to understand the underlying mechanism of epigenetic reprogramming of the transferred somatic cell nucleus. Especially in ruminants, somatic cell nuclear transfer (SCNT) is frequently associated with pathological changes in the foetal and placental phenotype and has significant consequences for development both before and after birth. The most critical factor is epigenetic reprogramming of the transferred somatic cell nucleus from its differentiated status into the totipotent state of the early embryo. This involves an erasure of the gene expression program of the respective donor cell and the establishment of the well-orchestrated sequence of expression of an estimated number of 10 000-12 000 genes regulating embryonic and foetal development. The following article reviews the present knowledge on the epigenetic reprogramming of the transferred somatic cell nucleus, with emphasis on DNA methylation, imprinting, X-chromosome inactivation and telomere length restoration in bovine development. Additionally, we briefly discuss other approaches towards epigenetic nuclear reprogramming, including the fusion of somatic and embryonic stem cells and the overexpression of genes crucial in the formation and maintenance of the pluripotent status. Improvements in our understanding of this dramatic epigenetic reprogramming event will be instrumental in realising the great potential of SCNT for basic biological research and for various agricultural and biomedical applications.

  18. Heat transfer analysis of fuel assemblies in a heterogeneous gas core nuclear rocket

    NASA Technical Reports Server (NTRS)

    Watanabe, Yoichi; Appelbaum, Jacob; Diaz, Nils; Maya, Isaac

    1991-01-01

    Heat transfer problems of a heterogeneous gaseous core nuclear rocket were studied. The reactor core consists of 1.5-m long hexagonal fuel assemblies filled with pressurized uranium tetrafluoride (UF4) gas. The fuel gas temperature ranges from 3500 to 7000 K at a nominal operating condition of 40 atm. Each fuel assembly has seven coolant tubes, through which hydrogen propellant flows. The propellant temperature is not constrained by the fuel temperature but by the maximum temperature of the graphite coolant tube. For a core achieving a fission power density of 1000 MW/cu m, the propellant core exit temperature can be as high as 3200 K. The physical size of a 1250 MW gaseous core nuclear rocket is comparable with that of a NERVA-type solid core nuclear rocket. The engine can deliver a specific impulse of 1020 seconds and a thrust of 330 kN.

  19. Rabbit embryonic stem cell lines derived from fertilized, parthenogenetic or somatic cell nuclear transfer embryos

    SciTech Connect

    Fang, Zhen F.; Gai, Hui; Huang, You Z.; Li, Shan G.; Chen, Xue J.; Shi, Jian J.; Wu, Li; Liu, Ailian; Xu, Ping; Sheng, Hui Z. . E-mail: hzsheng2003@yahoo.com

    2006-11-01

    Embryonic stem cells were isolated from rabbit blastocysts derived from fertilization (conventional rbES cells), parthenogenesis (pES cells) and nuclear transfer (ntES cells), and propagated in a serum-free culture system. Rabbit ES (rbES) cells proliferated for a prolonged time in an undifferentiated state and maintained a normal karyotype. These cells grew in a monolayer with a high nuclear/cytoplasm ratio and contained a high level of alkaline phosphate activity. In addition, rbES cells expressed the pluripotent marker Oct-4, as well as EBAF2, FGF4, TDGF1, but not antigens recognized by antibodies against SSEA-1, SSEA-3, SSEA-4, TRA-1-10 and TRA-1-81. All 3 types of ES cells formed embryoid bodies and generated teratoma that contained tissue types of all three germ layers. rbES cells exhibited a high cloning efficiency, were genetically modified readily and were used as nuclear donors to generate a viable rabbit through somatic cell nuclear transfer. In combination with genetic engineering, the ES cell technology should facilitate the creation of new rabbit lines.

  20. Interspecies stress in momentum equations for dense binary particulate systems.

    PubMed

    Zhang, D Z; Ma, X; Rauenzahn, R M

    2006-07-28

    For two-species particulate systems, ensemble averaged continuity and momentum equations for each species are derived based on the Liouville equation of the system. The ensemble average used is species specific. It is found that the interaction between species results in not only the interspecies force but also a stress in the momentum equations. In the limit that particles of one of the species can be considered as a continuum, the existence of the interspecies stress enables us to reduce the derived equations to the familiar form for dispersed two-phase flows.

  1. Production of the first cloned camel by somatic cell nuclear transfer.

    PubMed

    Wani, Nisar A; Wernery, U; Hassan, F A H; Wernery, R; Skidmore, J A

    2010-02-01

    In this study, we demonstrate the use of somatic cell nuclear transfer to produce the first cloned camelid, a dromedary camel (Camelus dromedarius) belonging to the family Camelidae. Donor karyoplasts were obtained from adult skin fibroblasts, cumulus cells, or fetal fibroblasts, and in vivo-matured oocytes, obtained from preovulatory follicles of superstimulated female camels by transvaginal ultrasound guided ovum pick-up, were used as cytoplasts. Reconstructed embryos were cultured in vitro for 7 days up to the hatching/hatched blastocyst stage before they were transferred to synchronized recipients on Day 6 after ovulation. Pregnancies were achieved from the embryos reconstructed from all cell types, and a healthy calf, named Injaz, was born from the pregnancy by an embryo reconstructed with cumulus cells. Genotype analyses, using 25 dromedary camel microsatellite markers, confirmed that the cloned calf was derived from the donor cell line and the ovarian tissue. In conclusion, the present study reports, for the first time, establishment of pregnancies and birth of the first cloned camelid, a dromedary camel (C. dromedarius), by use of somatic cell nuclear transfer. This has opened doors for the amelioration and preservation of genetically valuable animals like high milk producers, racing champions, and males of high genetic merit in camelids. We also demonstrated, for the first time, that adult and fetal fibroblasts can be cultured, expanded, and frozen without losing their ability to support the development of nuclear transfer embryos, a technology that may potentially be used to modify fibroblast genome by homologous recombination so as to generate genetically altered cloned animals.

  2. Generation of cloned mice from adult neurons by direct nuclear transfer.

    PubMed

    Mizutani, Eiji; Oikawa, Mami; Kassai, Hidetoshi; Inoue, Kimiko; Shiura, Hirosuke; Hirasawa, Ryutaro; Kamimura, Satoshi; Matoba, Shogo; Ogonuki, Narumi; Nagatomo, Hiroaki; Abe, Kuniya; Wakayama, Teruhiko; Aiba, Atsu; Ogura, Atsuo

    2015-03-01

    Whereas cloning mammals by direct somatic cell nuclear transfer has been successful using a wide range of donor cell types, neurons from adult brain remain "unclonable" for unknown reasons. Here, using a combination of two epigenetic approaches, we examined whether neurons from adult mice could be cloned. First, we used a specific antibody to discover cell types with reduced amounts of a repressive histone mark-dimethylated histone H3 lysine 9 (H3K9me2)-and identified CA1 pyramidal cells in the hippocampus and Purkinje cells in the cerebellum as candidates. Second, reconstructed embryos were treated with trichostatin A (TSA), a potent histone deacetylase inhibitor. Using CA1 cells, cloned offspring were obtained at high rates, reaching 10.2% and 4.6% (of embryos transferred) for male and female donors, respectively. Cerebellar Purkinje cell nuclei were too large to maintain their genetic integrity during nuclear transfer, leading to developmental arrest of embryos. However, gene expression analysis using cloned blastocysts corroborated a high rate of genomic reprogrammability of CA1 pyramidal and Purkinje cells. Neurons from the hippocampal dentate gyrus and cerebral cortex, which had higher amounts of H3K9me2, could also be used for producing cloned offspring, but the efficiencies were low. A more thorough analysis revealed that TSA treatment was essential for cloning adult neuronal cells. This study demonstrates, to our knowledge for the first time, that adult neurons can be cloned by nuclear transfer. Furthermore, our data imply that reduced amounts of H3K9me2 and increased histone acetylation appear to act synergistically to improve the development of cloned embryos.

  3. Coherent transfer of nuclear spin polarization in field-cycling NMR experiments

    SciTech Connect

    Pravdivtsev, Andrey N.; Yurkovskaya, Alexandra V.; Ivanov, Konstantin L.; Vieth, Hans-Martin

    2013-12-28

    Coherent polarization transfer effects in a coupled spin network have been studied over a wide field range. The transfer mechanism is based on exciting zero-quantum coherences between the nuclear spin states by means of non-adiabatic field jump from high to low magnetic field. Subsequent evolution of these coherences enables conversion of spin order in the system, which is monitored after field jump back to high field. Such processes are most efficient when the spin system passes through an avoided level crossing during the field variation. The polarization transfer effects have been demonstrated for N-acetyl histidine, which has five scalar coupled protons; the initial spin order has been prepared by applying RF-pulses at high magnetic field. The observed oscillatory transfer kinetics is taken as a clear indication of a coherent mechanism; level crossing effects have also been demonstrated. The experimental data are in very good agreement with the theoretical model of coherent polarization transfer. The method suggested is also valid for other types of initial polarization in the spin system, most notably, for spin hyperpolarization.

  4. Coherent transfer of nuclear spin polarization in field-cycling NMR experiments.

    PubMed

    Pravdivtsev, Andrey N; Yurkovskaya, Alexandra V; Vieth, Hans-Martin; Ivanov, Konstantin L

    2013-12-28

    Coherent polarization transfer effects in a coupled spin network have been studied over a wide field range. The transfer mechanism is based on exciting zero-quantum coherences between the nuclear spin states by means of non-adiabatic field jump from high to low magnetic field. Subsequent evolution of these coherences enables conversion of spin order in the system, which is monitored after field jump back to high field. Such processes are most efficient when the spin system passes through an avoided level crossing during the field variation. The polarization transfer effects have been demonstrated for N-acetyl histidine, which has five scalar coupled protons; the initial spin order has been prepared by applying RF-pulses at high magnetic field. The observed oscillatory transfer kinetics is taken as a clear indication of a coherent mechanism; level crossing effects have also been demonstrated. The experimental data are in very good agreement with the theoretical model of coherent polarization transfer. The method suggested is also valid for other types of initial polarization in the spin system, most notably, for spin hyperpolarization. PMID:24387362

  5. Development of Interspecies Correlation Models for Petroleum Hydrocarbons

    EPA Science Inventory

    Estimating the consequences of petroleum products to water column organisms has commonly been hampered by limited acute toxicity data, which exists only for a relatively small number of test species. In this study, we developed petroleum-specific Interspecies Correlation Estimati...

  6. Effect of the Compaction and the Size of DNA on the Nuclear Transfer Efficiency after Microinjection in Synchronized Cells

    PubMed Central

    Akita, Hidetaka; Kurihara, Dai; Schmeer, Marco; Schleef, Martin; Harashima, Hideyoshi

    2015-01-01

    The nuclear transfer process is one of the critical rate-limiting processes in transgene expression. In the present study, we report on the effect of compaction and the size of the DNA molecule on nuclear transfer efficiency by microinjection. A DNA/protamine complex- or variously-sized naked DNA molecules were injected into the cytoplasm or nucleus of synchronized HeLa cells. To evaluate the nuclear transfer process, a nuclear transfer score (NT score), calculated based on transgene expression after cytoplasmic microinjection divided by that after nuclear microinjection, was employed. The compaction of DNA with protamine decreased the NT score in comparison with the injection of naked DNA when the N/P ratio was increased to >2.0. Moreover, when naked DNA was microinjected, gene expression increased in parallel with the size of the DNA in the following order: minicircle DNA (MC07.CMV-EGFP; 2257 bp) > middle-sized plasmid DNA (pBS-EGFP; 3992 bp) > conventional plasmid DNA (pcDNA3.1-EGFP; 6172 bp), while the level of gene expression was quite comparable among them when the DNAs were injected into the nucleus. The above findings suggest that the intrinsic size of the DNA molecule is a major determinant for nuclear entry and that minicircle DNA has a great advantage in nuclear transfer. PMID:26066769

  7. Production of Cloned Korean Native Pig by Somatic Cell Nuclear Transfer

    PubMed Central

    Hwang, In-Sul; Kwon, Dae-Jin; Oh, Keun Bong; Ock, Sun-A; Chung, Hak-Jae; Cho, In-Cheol; Lee, Jeong-Woong; Im, Gi-Sun; Hwang, Seongsoo

    2015-01-01

    The Korean native pig (KNP) have been considered as animal models for animal biotechnology research because of their relatively small body size and their presumably highly inbred status due to the closed breeding program. However, little is reported about the use of KNP for animal biotechnology researches. This study was performed to establish the somatic cell nuclear transfer (SCNT) protocol for the production of swine leukocyte antigens (SLA) homotype-defined SCNT KNP. The ear fibroblast cells originated from KNP were cultured and used as donor cell. After thawing, the donor cells were cultured for 1 hour with 15 μM roscovitine prior to the nuclear transfer. The numbers of reconstructed and parthenogenetic embryos transferred were 98 ± 35.2 and 145 ± 11.2, respectively. The pregnancy and delivery rate were 3/5 (60%) and 2/5 (40%). One healthy SLA homotype-defined SCNT KNP was successfully generated. The recipient-based individual cloning efficiency ranged from 0.65 to 1.08%. Taken together, it can be postulated that the methodological establishment of the production of SLA homotype-defined cloned KNP can be applied to the generation of transgenic cloned KNP as model animals for human disease and xenotransplantation researches. PMID:27004264

  8. Production of Cloned Korean Native Pig by Somatic Cell Nuclear Transfer.

    PubMed

    Hwang, In-Sul; Kwon, Dae-Jin; Oh, Keun Bong; Ock, Sun-A; Chung, Hak-Jae; Cho, In-Cheol; Lee, Jeong-Woong; Im, Gi-Sun; Hwang, Seongsoo

    2015-06-01

    The Korean native pig (KNP) have been considered as animal models for animal biotechnology research because of their relatively small body size and their presumably highly inbred status due to the closed breeding program. However, little is reported about the use of KNP for animal biotechnology researches. This study was performed to establish the somatic cell nuclear transfer (SCNT) protocol for the production of swine leukocyte antigens (SLA) homotype-defined SCNT KNP. The ear fibroblast cells originated from KNP were cultured and used as donor cell. After thawing, the donor cells were cultured for 1 hour with 15 μM roscovitine prior to the nuclear transfer. The numbers of reconstructed and parthenogenetic embryos transferred were 98 ± 35.2 and 145 ± 11.2, respectively. The pregnancy and delivery rate were 3/5 (60%) and 2/5 (40%). One healthy SLA homotype-defined SCNT KNP was successfully generated. The recipient-based individual cloning efficiency ranged from 0.65 to 1.08%. Taken together, it can be postulated that the methodological establishment of the production of SLA homotype-defined cloned KNP can be applied to the generation of transgenic cloned KNP as model animals for human disease and xenotransplantation researches.

  9. Identification of nuclear effects in neutrino-carbon interactions at low three-momentum transfer

    DOE PAGESBeta

    Rodrigues, P. A.

    2016-02-17

    Two different nuclear-medium effects are isolated using a low three-momentum transfer subsample of neutrino-carbon scattering data from the MINERvA neutrino experiment. The observed hadronic energy in charged-current νμ interactions is combined with muon kinematics to permit separation of the quasielastic and Δ(1232) resonance processes. First, we observe a small cross section at very low energy transfer that matches the expected screening effect of long-range nucleon correlations. Second, additions to the event rate in the kinematic region between the quasielastic and Δ resonance processes are needed to describe the data. The data in this kinematic region also have an enhanced populationmore » of multiproton final states. Contributions predicted for scattering from a nucleon pair have both properties; the model tested in this analysis is a significant improvement but does not fully describe the data. We present the results as a double-differential cross section to enable further investigation of nuclear models. Furthermore, improved description of the effects of the nuclear environment are required by current and future neutrino oscillation experiments.« less

  10. Identification of Nuclear Effects in Neutrino-Carbon Interactions at Low Three-Momentum Transfer.

    PubMed

    Rodrigues, P A; Demgen, J; Miltenberger, E; Aliaga, L; Altinok, O; Bellantoni, L; Bercellie, A; Betancourt, M; Bodek, A; Bravar, A; Budd, H; Cai, T; Carneiro, M F; Chvojka, J; Devan, J; Dytman, S A; Díaz, G A; Eberly, B; Elkins, M; Felix, J; Fields, L; Fine, R; Gago, A M; Galindo, R; Gallagher, H; Ghosh, A; Golan, T; Gran, R; Harris, D A; Higuera, A; Hurtado, K; Kiveni, M; Kleykamp, J; Kordosky, M; Le, T; Leistico, J R; Lovlein, A; Maher, E; Manly, S; Mann, W A; Marshall, C M; Martinez Caicedo, D A; McFarland, K S; McGivern, C L; McGowan, A M; Messerly, B; Miller, J; Mislivec, A; Morfín, J G; Mousseau, J; Muhlbeier, T; Naples, D; Nelson, J K; Norrick, A; Nuruzzaman; Osta, J; Paolone, V; Patrick, C E; Perdue, G N; Ramirez, M A; Ransome, R D; Ray, H; Ren, L; Rimal, D; Ruterbories, D; Schellman, H; Schmitz, D W; Solano Salinas, C J; Tagg, N; Tice, B G; Valencia, E; Walton, T; Wolcott, J; Wospakrik, M; Zavala, G; Zhang, D

    2016-02-19

    Two different nuclear-medium effects are isolated using a low three-momentum transfer subsample of neutrino-carbon scattering data from the MINERvA neutrino experiment. The observed hadronic energy in charged-current ν_{μ} interactions is combined with muon kinematics to permit separation of the quasielastic and Δ(1232) resonance processes. First, we observe a small cross section at very low energy transfer that matches the expected screening effect of long-range nucleon correlations. Second, additions to the event rate in the kinematic region between the quasielastic and Δ resonance processes are needed to describe the data. The data in this kinematic region also have an enhanced population of multiproton final states. Contributions predicted for scattering from a nucleon pair have both properties; the model tested in this analysis is a significant improvement but does not fully describe the data. We present the results as a double-differential cross section to enable further investigation of nuclear models. Improved description of the effects of the nuclear environment are required by current and future neutrino oscillation experiments.

  11. Identification of Nuclear Effects in Neutrino-Carbon Interactions at Low Three-Momentum Transfer

    NASA Astrophysics Data System (ADS)

    Rodrigues, P. A.; Demgen, J.; Miltenberger, E.; Aliaga, L.; Altinok, O.; Bellantoni, L.; Bercellie, A.; Betancourt, M.; Bodek, A.; Bravar, A.; Budd, H.; Cai, T.; Carneiro, M. F.; Chvojka, J.; Devan, J.; Dytman, S. A.; Díaz, G. A.; Eberly, B.; Elkins, M.; Felix, J.; Fields, L.; Fine, R.; Gago, A. M.; Galindo, R.; Gallagher, H.; Ghosh, A.; Golan, T.; Gran, R.; Harris, D. A.; Higuera, A.; Hurtado, K.; Kiveni, M.; Kleykamp, J.; Kordosky, M.; Le, T.; Leistico, J. R.; Lovlein, A.; Maher, E.; Manly, S.; Mann, W. A.; Marshall, C. M.; Martinez Caicedo, D. A.; McFarland, K. S.; McGivern, C. L.; McGowan, A. M.; Messerly, B.; Miller, J.; Mislivec, A.; Morfín, J. G.; Mousseau, J.; Muhlbeier, T.; Naples, D.; Nelson, J. K.; Norrick, A.; Nuruzzaman; Osta, J.; Paolone, V.; Patrick, C. E.; Perdue, G. N.; Ramirez, M. A.; Ransome, R. D.; Ray, H.; Ren, L.; Rimal, D.; Ruterbories, D.; Schellman, H.; Schmitz, D. W.; Solano Salinas, C. J.; Tagg, N.; Tice, B. G.; Valencia, E.; Walton, T.; Wolcott, J.; Wospakrik, M.; Zavala, G.; Zhang, D.; Minerva Collaboration

    2016-02-01

    Two different nuclear-medium effects are isolated using a low three-momentum transfer subsample of neutrino-carbon scattering data from the MINERvA neutrino experiment. The observed hadronic energy in charged-current νμ interactions is combined with muon kinematics to permit separation of the quasielastic and Δ (1232 ) resonance processes. First, we observe a small cross section at very low energy transfer that matches the expected screening effect of long-range nucleon correlations. Second, additions to the event rate in the kinematic region between the quasielastic and Δ resonance processes are needed to describe the data. The data in this kinematic region also have an enhanced population of multiproton final states. Contributions predicted for scattering from a nucleon pair have both properties; the model tested in this analysis is a significant improvement but does not fully describe the data. We present the results as a double-differential cross section to enable further investigation of nuclear models. Improved description of the effects of the nuclear environment are required by current and future neutrino oscillation experiments.

  12. Genetic Drift Can Compromise Mitochondrial Replacement by Nuclear Transfer in Human Oocytes.

    PubMed

    Yamada, Mitsutoshi; Emmanuele, Valentina; Sanchez-Quintero, Maria J; Sun, Bruce; Lallos, Gregory; Paull, Daniel; Zimmer, Matthew; Pagett, Shardonay; Prosser, Robert W; Sauer, Mark V; Hirano, Michio; Egli, Dieter

    2016-06-01

    Replacement of mitochondria through nuclear transfer between oocytes of two different women has emerged recently as a strategy for preventing inheritance of mtDNA diseases. Although experiments in human oocytes have shown effective replacement, the consequences of small amounts of mtDNA carryover have not been studied sufficiently. Using human mitochondrial replacement stem cell lines, we show that, even though the low levels of heteroplasmy introduced into human oocytes by mitochondrial carryover during nuclear transfer often vanish, they can sometimes instead result in mtDNA genotypic drift and reversion to the original genotype. Comparison of cells with identical oocyte-derived nuclear DNA but different mtDNA shows that either mtDNA genotype is compatible with the nucleus and that drift is independent of mitochondrial function. Thus, although functional replacement of the mitochondrial genome is possible, even low levels of heteroplasmy can affect the stability of the mtDNA genotype and compromise the efficacy of mitochondrial replacement.

  13. Identification of Nuclear Effects in Neutrino-Carbon Interactions at Low Three-Momentum Transfer.

    PubMed

    Rodrigues, P A; Demgen, J; Miltenberger, E; Aliaga, L; Altinok, O; Bellantoni, L; Bercellie, A; Betancourt, M; Bodek, A; Bravar, A; Budd, H; Cai, T; Carneiro, M F; Chvojka, J; Devan, J; Dytman, S A; Díaz, G A; Eberly, B; Elkins, M; Felix, J; Fields, L; Fine, R; Gago, A M; Galindo, R; Gallagher, H; Ghosh, A; Golan, T; Gran, R; Harris, D A; Higuera, A; Hurtado, K; Kiveni, M; Kleykamp, J; Kordosky, M; Le, T; Leistico, J R; Lovlein, A; Maher, E; Manly, S; Mann, W A; Marshall, C M; Martinez Caicedo, D A; McFarland, K S; McGivern, C L; McGowan, A M; Messerly, B; Miller, J; Mislivec, A; Morfín, J G; Mousseau, J; Muhlbeier, T; Naples, D; Nelson, J K; Norrick, A; Nuruzzaman; Osta, J; Paolone, V; Patrick, C E; Perdue, G N; Ramirez, M A; Ransome, R D; Ray, H; Ren, L; Rimal, D; Ruterbories, D; Schellman, H; Schmitz, D W; Solano Salinas, C J; Tagg, N; Tice, B G; Valencia, E; Walton, T; Wolcott, J; Wospakrik, M; Zavala, G; Zhang, D

    2016-02-19

    Two different nuclear-medium effects are isolated using a low three-momentum transfer subsample of neutrino-carbon scattering data from the MINERvA neutrino experiment. The observed hadronic energy in charged-current ν_{μ} interactions is combined with muon kinematics to permit separation of the quasielastic and Δ(1232) resonance processes. First, we observe a small cross section at very low energy transfer that matches the expected screening effect of long-range nucleon correlations. Second, additions to the event rate in the kinematic region between the quasielastic and Δ resonance processes are needed to describe the data. The data in this kinematic region also have an enhanced population of multiproton final states. Contributions predicted for scattering from a nucleon pair have both properties; the model tested in this analysis is a significant improvement but does not fully describe the data. We present the results as a double-differential cross section to enable further investigation of nuclear models. Improved description of the effects of the nuclear environment are required by current and future neutrino oscillation experiments. PMID:26943528

  14. Coupled electron-nuclear dynamics: Charge migration and charge transfer initiated near a conical intersection

    SciTech Connect

    Mendive-Tapia, David; Vacher, Morgane; Bearpark, Michael J.; Robb, Michael A.

    2013-07-28

    Coupled electron-nuclear dynamics, implemented using the Ehrenfest method, has been used to study charge migration with fixed nuclei, together with charge transfer when nuclei are allowed to move. Simulations were initiated at reference geometries of neutral benzene and 2-phenylethylamine (PEA), and at geometries close to potential energy surface crossings in the cations. Cationic eigenstates, and the so-called sudden approximation, involving removal of an electron from a correlated ground-state wavefunction for the neutral species, were used as initial conditions. Charge migration without coupled nuclear motion could be observed if the Ehrenfest simulation, using the sudden approximation, was started near a conical intersection where the states were both strongly coupled and quasi-degenerate. Further, the main features associated with charge migration were still recognizable when the nuclear motion was allowed to couple. In the benzene radical cation, starting from the reference neutral geometry with the sudden approximation, one could observe sub-femtosecond charge migration with a small amplitude, which results from weak interaction with higher electronic states. However, we were able to engineer large amplitude charge migration, with a period between 10 and 100 fs, corresponding to oscillation of the electronic structure between the quinoid and anti-quinoid cationic electronic configurations, by distorting the geometry along the derivative coupling vector from the D{sub 6h} Jahn-Teller crossing to lower symmetry where the states are not degenerate. When the nuclear motion becomes coupled, the period changes only slightly. In PEA, in an Ehrenfest trajectory starting from the D{sub 2} eigenstate and reference geometry, a partial charge transfer occurs after about 12 fs near the first crossing between D{sub 1}, D{sub 2} (N{sup +}-Phenyl, N-Phenyl{sup +}). If the Ehrenfest propagation is started near this point, using the sudden approximation without coupled

  15. Coupled electron-nuclear dynamics: Charge migration and charge transfer initiated near a conical intersection

    NASA Astrophysics Data System (ADS)

    Mendive-Tapia, David; Vacher, Morgane; Bearpark, Michael J.; Robb, Michael A.

    2013-07-01

    Coupled electron-nuclear dynamics, implemented using the Ehrenfest method, has been used to study charge migration with fixed nuclei, together with charge transfer when nuclei are allowed to move. Simulations were initiated at reference geometries of neutral benzene and 2-phenylethylamine (PEA), and at geometries close to potential energy surface crossings in the cations. Cationic eigenstates, and the so-called sudden approximation, involving removal of an electron from a correlated ground-state wavefunction for the neutral species, were used as initial conditions. Charge migration without coupled nuclear motion could be observed if the Ehrenfest simulation, using the sudden approximation, was started near a conical intersection where the states were both strongly coupled and quasi-degenerate. Further, the main features associated with charge migration were still recognizable when the nuclear motion was allowed to couple. In the benzene radical cation, starting from the reference neutral geometry with the sudden approximation, one could observe sub-femtosecond charge migration with a small amplitude, which results from weak interaction with higher electronic states. However, we were able to engineer large amplitude charge migration, with a period between 10 and 100 fs, corresponding to oscillation of the electronic structure between the quinoid and anti-quinoid cationic electronic configurations, by distorting the geometry along the derivative coupling vector from the D6h Jahn-Teller crossing to lower symmetry where the states are not degenerate. When the nuclear motion becomes coupled, the period changes only slightly. In PEA, in an Ehrenfest trajectory starting from the D2 eigenstate and reference geometry, a partial charge transfer occurs after about 12 fs near the first crossing between D1, D2 (N+-Phenyl, N-Phenyl+). If the Ehrenfest propagation is started near this point, using the sudden approximation without coupled nuclear motion, one observes an

  16. Coupled electron-nuclear dynamics: charge migration and charge transfer initiated near a conical intersection.

    PubMed

    Mendive-Tapia, David; Vacher, Morgane; Bearpark, Michael J; Robb, Michael A

    2013-07-28

    Coupled electron-nuclear dynamics, implemented using the Ehrenfest method, has been used to study charge migration with fixed nuclei, together with charge transfer when nuclei are allowed to move. Simulations were initiated at reference geometries of neutral benzene and 2-phenylethylamine (PEA), and at geometries close to potential energy surface crossings in the cations. Cationic eigenstates, and the so-called sudden approximation, involving removal of an electron from a correlated ground-state wavefunction for the neutral species, were used as initial conditions. Charge migration without coupled nuclear motion could be observed if the Ehrenfest simulation, using the sudden approximation, was started near a conical intersection where the states were both strongly coupled and quasi-degenerate. Further, the main features associated with charge migration were still recognizable when the nuclear motion was allowed to couple. In the benzene radical cation, starting from the reference neutral geometry with the sudden approximation, one could observe sub-femtosecond charge migration with a small amplitude, which results from weak interaction with higher electronic states. However, we were able to engineer large amplitude charge migration, with a period between 10 and 100 fs, corresponding to oscillation of the electronic structure between the quinoid and anti-quinoid cationic electronic configurations, by distorting the geometry along the derivative coupling vector from the D6h Jahn-Teller crossing to lower symmetry where the states are not degenerate. When the nuclear motion becomes coupled, the period changes only slightly. In PEA, in an Ehrenfest trajectory starting from the D2 eigenstate and reference geometry, a partial charge transfer occurs after about 12 fs near the first crossing between D1, D2 (N(+)-Phenyl, N-Phenyl(+)). If the Ehrenfest propagation is started near this point, using the sudden approximation without coupled nuclear motion, one observes an

  17. Coupled electron-nuclear dynamics: charge migration and charge transfer initiated near a conical intersection.

    PubMed

    Mendive-Tapia, David; Vacher, Morgane; Bearpark, Michael J; Robb, Michael A

    2013-07-28

    Coupled electron-nuclear dynamics, implemented using the Ehrenfest method, has been used to study charge migration with fixed nuclei, together with charge transfer when nuclei are allowed to move. Simulations were initiated at reference geometries of neutral benzene and 2-phenylethylamine (PEA), and at geometries close to potential energy surface crossings in the cations. Cationic eigenstates, and the so-called sudden approximation, involving removal of an electron from a correlated ground-state wavefunction for the neutral species, were used as initial conditions. Charge migration without coupled nuclear motion could be observed if the Ehrenfest simulation, using the sudden approximation, was started near a conical intersection where the states were both strongly coupled and quasi-degenerate. Further, the main features associated with charge migration were still recognizable when the nuclear motion was allowed to couple. In the benzene radical cation, starting from the reference neutral geometry with the sudden approximation, one could observe sub-femtosecond charge migration with a small amplitude, which results from weak interaction with higher electronic states. However, we were able to engineer large amplitude charge migration, with a period between 10 and 100 fs, corresponding to oscillation of the electronic structure between the quinoid and anti-quinoid cationic electronic configurations, by distorting the geometry along the derivative coupling vector from the D6h Jahn-Teller crossing to lower symmetry where the states are not degenerate. When the nuclear motion becomes coupled, the period changes only slightly. In PEA, in an Ehrenfest trajectory starting from the D2 eigenstate and reference geometry, a partial charge transfer occurs after about 12 fs near the first crossing between D1, D2 (N(+)-Phenyl, N-Phenyl(+)). If the Ehrenfest propagation is started near this point, using the sudden approximation without coupled nuclear motion, one observes an

  18. Full-term development of nuclear transfer calves produced from open-pulled straw (OPS) vitrified cytoplasts: work in progress.

    PubMed

    Booth, P J; Vajta, G; Høj, A; Holm, P; Jacobsen, H; Greve, T; Callesen, H

    1999-04-01

    Cryopreservation of cytoplasts would help to resolve the logistics of matching the availability of oocytes with embryo donors in nuclear transfer. Therefore, the developmental potential of nuclear transfer bovine embryos reconstructed using vitrified cytoplasts was investigated. In vitro matured oocytes were denuded, enucleated, activated with calcium ionophore (10 microM, 5 min) and cycloheximide (10 microg/mL, 6 h) and then vitrified by the open pulled straw (OPS) method. After immediate warming, the nuclear transfer embryos were reconstructed using blastomeres from nonvitrified,in vitro-produced embryo donors. Compared with control nuclear transfer embryos that were reconstructed using nonvitrified cytoplasts, fusion rates (% +/- SEM) were not affected (83.7+/-9.2 vs. 79.8+/-4.6; P>0.05), but cleavage (55.7+/-2.9 vs. 92.8+/-3.9; P = 0.0002) and blastocyst rates (7.2+/-5.0 vs. 32.6+/-7.8; P = 0.0025, vitrified vs. nonvitrified cytoplasts, respectively) per successful fusion were reduced. One nuclear transfer blastocyst reconstructed from a vitrified cytoplast was transferred to a synchronized recipient. After a normal length gestation (265 d), twin calves (21 and 26 kg) were delivered. Microsatellite analysis confirmed that the calves were homozygotic (the embryo split in utero), and were derived from the in vitro-produced embryo donor. The twins were dead at birth, but post-mortem analysis of the calves indicated no abnormalities or infections, suggesting that their death was related to the twin pregnancy and the known fragility of nuclear transfer calves. These data demonstrate that open pulled straw-vitrified cytoplasts are capable of supporting full-term development of nuclear transfer embryos. PMID:10729021

  19. Conservation of the Sapsaree (Canis familiaris), a Korean Natural Monument, using somatic cell nuclear transfer.

    PubMed

    Jang, Goo; Hong, SoGun; Kang, JungTaek; Park, JungEun; Oh, HyunJu; Park, ChanKyu; Ha, JiHong; Kim, DaeYong; Kim, MinKyu; Lee, ByeongChun

    2009-09-01

    A recent emerging technology, somatic cell nuclear transfer (SCNT), has been considered for conserving threatened or endangered species. Sapsaree is a native breed in Korea and has been designated as a Natural Monument. The aim of this study was to produce a Sapsaree by SCNT for breed conservation. Donor fibroblasts from a 9-year-old male Sapsaree were placed into the perivitelline spaces of enucleated in vivo matured oocytes and fused electrically. A total of 309 cloned embryos were transferred into the oviducts of 15 naturally synchronized recipients. Two recipients were diagnosed as pregnant, and each delivered one cloned puppy, both of which weighed 530 g. Overall, this study demonstrated that an endangered canine breed can be conserved by SCNT.

  20. Animal cloning by nuclear transfer: state-of-the-art and future perspectives.

    PubMed

    Dinnyes, Andras; Szmolenszky, Agnes

    2005-01-01

    Model organisms are essential to study the genetic basis of human diseases. Transgenic mammalian models, especially genetic knock-out mice have catalysed the progress in this area. To continue the advancement, further sophisticated and refined models are crucially needed to study the genetic basis and manifestations of numerous human diseases. Coinciding with the start of the new era of post-genomic research, new tools for establishment of transgenesis, such as nuclear transfer and gene targeting in somatic cells, have become available, offering a unique opportunity for the generation of transgenic animal models. The new technology provides important tools for comparative functional genomics to promote the interpretation and increase the practical value of the data generated in numerous mouse models. This paper discusses the state-of-the-art of the nuclear replacement technology and presents future perspectives.

  1. Widespread horizontal gene transfer from double-stranded RNA viruses to eukaryotic nuclear genomes.

    PubMed

    Liu, Huiquan; Fu, Yanping; Jiang, Daohong; Li, Guoqing; Xie, Jiatao; Cheng, Jiasen; Peng, Youliang; Ghabrial, Said A; Yi, Xianhong

    2010-11-01

    Horizontal gene transfer commonly occurs from cells to viruses but rarely occurs from viruses to their host cells, with the exception of retroviruses and some DNA viruses. However, extensive sequence similarity searches in public genome databases for various organisms showed that the capsid protein and RNA-dependent RNA polymerase genes from totiviruses and partitiviruses have widespread homologs in the nuclear genomes of eukaryotic organisms, including plants, arthropods, fungi, nematodes, and protozoa. PCR amplification and sequencing as well as comparative evidence of junction coverage between virus and host sequences support the conclusion that these viral homologs are real and occur in eukaryotic genomes. Sequence comparison and phylogenetic analysis suggest that these genes were likely transferred horizontally from viruses to eukaryotic genomes. Furthermore, we present evidence showing that some of the transferred genes are conserved and expressed in eukaryotic organisms and suggesting that these viral genes are also functional in the recipient genomes. Our findings imply that horizontal transfer of double-stranded RNA viral genes is widespread among eukaryotes and may give rise to functionally important new genes, thus entailing that RNA viruses may play significant roles in the evolution of eukaryotes.

  2. Somatic cell nuclear transfer-derived embryonic stem cell lines in humans: pros and cons.

    PubMed

    Langerova, Alena; Fulka, Helena; Fulka, Josef

    2013-12-01

    The recent paper, published by Mitalipov's group in Cell (Tachibana et al., 2013 ), reporting the production of human somatic cell nuclear transfer (SCNT) embryonic stem cells (ESCs), opens again the debate if, in the era of induced pluripotent stem cells (iPSCs), the production of these cells is indeed necessary and, if so, whether they are different from ESCs produced from spare embryos and iPSCs. It is our opinion that these questions are very difficult to answer because it is still unclear whether and how normal ESCs differ from iPSCs. PMID:24180743

  3. Optimization of procedures for cloning by somatic cell nuclear transfer in mice.

    PubMed

    Chung, Young Gie; Gao, Shaorong; Latham, Keith E

    2006-01-01

    Cloning by somatic cell nuclear transfer is a complex procedure that is dependent on correct interactions between oocyte and donor cell genome. These interactions require minimal insult to either the oocyte or the transplanted nucleus. Available data also indicate that reprogramming the donor cell genome may be slow, so that the cloned embryo expresses genes typical of the donor cell, and thus has different characteristics from normal embryos. Procedures that minimize damage to the donor genome and that address the unique characteristics of the cloned construct should enhance the efficacy of the method.

  4. Somatic cell nuclear transfer-derived embryonic stem cell lines in humans: pros and cons.

    PubMed

    Langerova, Alena; Fulka, Helena; Fulka, Josef

    2013-12-01

    The recent paper, published by Mitalipov's group in Cell (Tachibana et al., 2013 ), reporting the production of human somatic cell nuclear transfer (SCNT) embryonic stem cells (ESCs), opens again the debate if, in the era of induced pluripotent stem cells (iPSCs), the production of these cells is indeed necessary and, if so, whether they are different from ESCs produced from spare embryos and iPSCs. It is our opinion that these questions are very difficult to answer because it is still unclear whether and how normal ESCs differ from iPSCs.

  5. Development of porcine tetraploid somatic cell nuclear transfer embryos is influenced by oocyte nuclei.

    PubMed

    Fu, Bo; Liu, Di; Ma, Hong; Guo, Zhen-Hua; Wang, Liang; Li, Zhong-Qiu; Peng, Fu-Gang; Bai, Jing

    2016-02-01

    Cloning efficiency in mammalian systems remains low because reprogramming of donor cells is frequently incomplete. Nuclear factors in the oocyte are removed by enucleation, and this removal may adversely affect reprogramming efficiency. Here, we investigated the role of porcine oocyte nuclear factors during reprogramming. We introduced somatic cell nuclei into intact MII oocytes to establish tetraploid somatic cell nuclear transfer (SCNT) embryos containing both somatic nuclei and oocyte nuclei. We then examined the influence of the oocyte nucleus on tetraploid SCNT embryo development by assessing characteristics including pronucleus formation, cleavage rate, and blastocyst formation. Overall, tetraploid SCNT embryos have a higher developmental competence than do standard diploid SCNT embryos. Therefore, we have established an embryonic model in which a fetal fibroblast nucleus and an oocyte metaphase II plate coexist. Tetraploid SCNT represents a new research platform that is potentially useful for examining interactions between donor nuclei and oocyte nuclei. This platform should facilitate further understanding of the roles played by nuclear factors during reprogramming.

  6. Nuclear event time histories and computed site transfer functions for locations in the Los Angeles region

    USGS Publications Warehouse

    Rogers, A.M.; Covington, P.A.; Park, R.B.; Borcherdt, R.D.; Perkins, D.M.

    1980-01-01

    This report presents a collection of Nevada Test Site (NTS) nuclear explosion recordings obtained at sites in the greater Los Angeles, Calif., region. The report includes ground velocity time histories, as well as, derived site transfer functions. These data have been collected as part of a study to evaluate the validity of using low-level ground motions to predict the frequency-dependent response of a site during an earthquake. For this study 19 nuclear events were recorded at 98 separate locations. Some of these sites have recorded more than one of the nuclear explosions, and, consequently, there are a total of 159, three-component station records. The location of all the recording sites are shown in figures 1–5, the station coordinates and abbreviations are given in table 1. The station addresses are listed in table 2, and the nuclear explosions that were recorded are listed in table 3. The recording sites were chosen on the basis of three criteria: (1) that the underlying geological conditions were representative of conditions over significant areas of the region, (2) that the site was the location of a strong-motion recording of the 1971 San Fernando earthquake, or (3) that more complete geographical coverage was required in that location.

  7. Human-mouse interspecies collagen I heterotrimer is functional during embryonic development of Mov13 mutant mouse embryos.

    PubMed

    Wu, H; Bateman, J F; Schnieke, A; Sharpe, A; Barker, D; Mascara, T; Eyre, D; Bruns, R; Krimpenfort, P; Berns, A

    1990-04-01

    To investigate whether the human pro alpha 1(I) collagen chain could form an in vivo functional interspecies heterotrimer with the mouse pro alpha 2(I) collagen chain, we introduced the human COL1A1 gene into Mov13 mice which have a functional deletion of the endogenous COL1A1 gene. Transgenic mouse strains (HucI and HucII) carrying the human COL1A1 gene were first generated by microinjecting the COL1A1 gene into wild-type mouse embryos. Genetic evidence indicated that the transgene in the HucI strain was closely linked to the endogenous mouse COL1A1 gene and was X linked in the HucII transgenic strain. Northern (RNA) blot and S1 protection analyses showed that the transgene was expressed in the appropriate tissue-specific manner and as efficiently as the endogenous COL1A1 gene. HucII mice were crossed with Mov13 mice to transfer the human transgene into the mutant strain. Whereas homozygous Mov13 embryos die between days 13 and 14 of gestation, the presence of the transgene permitted apparently normal development of the mutant embryos to birth. This indicated that the mouse-human interspecies collagen I heterotrimer was functional in the animal. The rescue was, however, only partial, as all homozygotes died within 36 h after delivery, with signs of internal bleeding. This could have been due to a functional defect in the interspecies hybrid collagen. Extensive analysis failed to reveal any biochemical or morphological abnormalities of the collagen I molecules in Mov13-HucII embryos. This may indicate that there was a subtle functional defect of the interspecies hybrid protein which was not revealed by our analysis or that another gene has been mutated by the retroviral insertion in the Mov13 mutant strain.

  8. Histone deacetylase inhibitor significantly improved the cloning efficiency of porcine somatic cell nuclear transfer embryos.

    PubMed

    Huang, Yongye; Tang, Xiaochun; Xie, Wanhua; Zhou, Yan; Li, Dong; Yao, Chaogang; Zhou, Yang; Zhu, Jianguo; Lai, Liangxue; Ouyang, Hongsheng; Pang, Daxin

    2011-12-01

    Valproic acid (VPA), a histone deacetylase inbibitor, has been shown to generate inducible pluripotent stem (iPS) cells from mouse and human fibroblasts with a significant higher efficiency. Because successful cloning by somatic cell nuclear transfer (SCNT) undergoes a full reprogramming process in which the epigenetic state of a differentiated donor nuclear is converted into an embryonic totipotent state, we speculated that VPA would be useful in promoting cloning efficiency. Therefore, in the present study, we examined whether VPA can promote the developmental competence of SCNT embryos by improving the reprogramming state of donor nucleus. Here we report that 1 mM VPA for 14 to 16 h following activation significantly increased the rate of blastocyst formation of porcine SCNT embryos constructed from Landrace fetal fibroblast cells compared to the control (31.8 vs. 11.4%). However, we found that the acetylation level of Histone H3 lysine 14 and Histone H4 lysine 5 and expression level of Oct4, Sox2, and Klf4 was not significantly changed between VPA-treated and -untreated groups at the blastocyst stage. The SCNT embryos were transferred to 38 surrogates, and the cloning efficiency in the treated group was significantly improved compared with the control group. Taken together, we have demonstrated that VPA can improve both in vitro and in vivo development competence of porcine SCNT embryos.

  9. Embryo production and possible species preservation by nuclear transfer of somatic cells isolated from bovine semen.

    PubMed

    Liu, Jie; Westhusin, Mark; Long, Charles; Johnson, Gregory; Burghardt, Robert; Kraemer, Duane

    2010-12-01

    Somatic cells in semen are a potential source of nuclei for nuclear transfer to produce genetically identical animals; this is especially important when an animal has died and the only viable genetic material available is frozen semen. Usefulness of somatic cells obtained from fresh (cultured) and frozen (isolated, not cultured) bovine semen for nuclear transfer was evaluated. Twelve ejaculates were collected from nine bulls representing three breeds: Charolais, Brahman, and crossbred Rodeo bull. All samples were processed immediately and cell growth was obtained from seven of the twelve ejaculates (58.3%). Cells from three bulls (with the best growth rates) were evaluated by optical microscopy and used in cloning experiments. In culture, these cells exhibited classic epithelial morphology and expressed cytokeratin and vimentin, indicating they were of epithelial origin. When cells from the three bulls were used as donor cells, 15.9% (18/113), 34.5% (29/84), and 14.4% (13/90) of the fused embryos developed into blastocysts, respectively. Of the blastocyst stage embryos, 38.9% (7/18), 72.4% (21/29), and 61.5% (8/13) hatched, respectively. Somatic cells isolated (not cultured) from frozen bovine semen were also used in the cloning experiments. Although cleavage occurred, no compact morulae or blastocysts were obtained. In conclusion, epithelial cell growth was obtained from fresh bovine ejaculates with relatively high efficiency. Somatic cells from semen can be used as nucleus donors to produce cloned blastocyst-stage embryos.

  10. Nuclear transfer with apoptotic bovine fibroblasts: can programmed cell death be reprogrammed?

    PubMed

    Miranda, Moyses dos Santos; Bressan, Fabiana Fernandes; De Bem, Tiago Henrique Camara; Merighe, Giovana Krempel Fonseca; Ohashi, Otávio Mitio; King, William Alan; Meirelles, Flavio Viera

    2012-06-01

    Cell death by apoptosis is considered to be irreversible. However, reports have indicated that its reversibility is possible if the cells have not yet reached the "point of no return." In order to add new information about this topic, we used cells at different moments of apoptotic process as nuclear donors in somatic cell nuclear transfer (SCNT) in order to test if programmed cell death can be reversed. Adult bovine fibroblasts were treated with 10 μM of staurosporine (STP) for 3 h and analyzed for phosphatidylserine externalization (Annexin assay) and presence of active caspase-9. Annexin-positive (Anx+) and Caspase-9-positive (Casp-9+) cells were isolated by FACS and immediately transferred into enucleated in vitro matured bovine oocytes. After STP treatment, 89.9% of cells were Anx+ (4.6% in control cells; p<0.01) and 24.9% were Casp-9+ (2.4% in control cells; p<0.01). Fusion and cleavage were not affected by the use apoptotic cells (p>0.05). Also, the use of Anx+ cells did not affect blastocyst production compared to control (26.4% vs. 22.9%, respectively; p>0.05). However, blastocyst formation was affected by the use of Casp-9+ cells (12.3%; p<0.05). These findings contribute to the idea of that apoptosis is reversible only at early stages. Additionally, we hypothesize that the "point of no return" for apoptosis may be located around activation of Caspase-9.

  11. Targeted disruption of Ataxia-telangiectasia mutated gene in miniature pigs by somatic cell nuclear transfer

    SciTech Connect

    Kim, Young June; Ahn, Kwang Sung; Kim, Minjeong; Kim, Min Ju; Park, Sang-Min; Ryu, Junghyun; Ahn, Jin Seop; Heo, Soon Young; Kang, Jee Hyun; Choi, You Jung; Choi, Seong-Jun; Shim, Hosup

    2014-10-03

    Highlights: • ATM gene-targeted pigs were produced by somatic cell nuclear transfer. • A novel large animal model for ataxia telangiectasia was developed. • The new model may provide an alternative to the mouse model. - Abstract: Ataxia telangiectasia (A-T) is a recessive autosomal disorder associated with pleiotropic phenotypes, including progressive cerebellar degeneration, gonad atrophy, and growth retardation. Even though A-T is known to be caused by the mutations in the Ataxia telangiectasia mutated (ATM) gene, the correlation between abnormal cellular physiology caused by ATM mutations and the multiple symptoms of A-T disease has not been clearly determined. None of the existing ATM mouse models properly reflects the extent to which neurological degeneration occurs in human. In an attempt to provide a large animal model for A-T, we produced gene-targeted pigs with mutations in the ATM gene by somatic cell nuclear transfer. The disrupted allele in the ATM gene of cloned piglets was confirmed via PCR and Southern blot analysis. The ATM gene-targeted pigs generated in the present study may provide an alternative to the current mouse model for the study of mechanisms underlying A-T disorder and for the development of new therapies.

  12. Somatic cell nuclear transfer: Infinite reproduction of a unique diploid genome

    SciTech Connect

    Kishigami, Satoshi Wakayama, Sayaka; Hosoi, Yoshihiko; Iritani, Akira; Wakayama, Teruhiko

    2008-06-10

    In mammals, a diploid genome of an individual following fertilization of an egg and a spermatozoon is unique and irreproducible. This implies that the generated unique diploid genome is doomed with the individual ending. Even as cultured cells from the individual, they cannot normally proliferate in perpetuity because of the 'Hayflick limit'. However, Dolly, the sheep cloned from an adult mammary gland cell, changes this scenario. Somatic cell nuclear transfer (SCNT) enables us to produce offspring without germ cells, that is, to 'passage' a unique diploid genome. Animal cloning has also proven to be a powerful research tool for reprogramming in many mammals, notably mouse and cow. The mechanism underlying reprogramming, however, remains largely unknown and, animal cloning has been inefficient as a result. More momentously, in addition to abortion and fetal mortality, some cloned animals display possible premature aging phenotypes including early death and short telomere lengths. Under these inauspicious conditions, is it really possible for SCNT to preserve a diploid genome? Delightfully, in mouse and recently in primate, using SCNT we can produce nuclear transfer ES cells (ntES) more efficiently, which can preserve the eternal lifespan for the 'passage' of a unique diploid genome. Further, new somatic cloning technique using histone-deacetylase inhibitors has been developed which can significantly increase the previous cloning rates two to six times. Here, we introduce SCNT and its value as a preservation tool for a diploid genome while reviewing aging of cloned animals on cellular and individual levels.

  13. Aberrant expression of Xist in aborted porcine fetuses derived from somatic cell nuclear transfer embryos.

    PubMed

    Yuan, Lin; Wang, Anfeng; Yao, Chaogang; Huang, Yongye; Duan, Feifei; Lv, Qinyan; Wang, Dongxu; Ouyang, Hongsheng; Li, Zhanjun; Lai, Liangxue

    2014-01-01

    Cloned pigs generated by somatic cell nuclear transfer (SCNT) show a greater ratio of early abortion during mid-gestation than normal controls. X-linked genes have been demonstrated to be important for the development of cloned embryos. To determine the relationship between the expression of X-linked genes and abortion of cloned porcine fetuses, the expression of X-linked genes were investigated by quantitative real-time polymerase chain reaction (q-PCR) and the methylation status of Xist DMR was performed by bisulfate-specific PCR (BSP). q-PCR analysis indicated that there was aberrant expression of X-linked genes, especially the upregulated expression of Xist in both female and male aborted fetuses compared to control fetuses. Results of BSP suggested that hypomethylation of Xist occurred in aborted fetuses, whether male or female. These results suggest that the abnormal expression of Xist may be associated with the abortion of fetuses derived from somatic cell nuclear transfer embryos. PMID:25429426

  14. Somatic cell nuclear transfer: infinite reproduction of a unique diploid genome.

    PubMed

    Kishigami, Satoshi; Wakayama, Sayaka; Hosoi, Yoshihiko; Iritani, Akira; Wakayama, Teruhiko

    2008-06-10

    In mammals, a diploid genome of an individual following fertilization of an egg and a spermatozoon is unique and irreproducible. This implies that the generated unique diploid genome is doomed with the individual ending. Even as cultured cells from the individual, they cannot normally proliferate in perpetuity because of the "Hayflick limit". However, Dolly, the sheep cloned from an adult mammary gland cell, changes this scenario. Somatic cell nuclear transfer (SCNT) enables us to produce offspring without germ cells, that is, to "passage" a unique diploid genome. Animal cloning has also proven to be a powerful research tool for reprogramming in many mammals, notably mouse and cow. The mechanism underlying reprogramming, however, remains largely unknown and, animal cloning has been inefficient as a result. More momentously, in addition to abortion and fetal mortality, some cloned animals display possible premature aging phenotypes including early death and short telomere lengths. Under these inauspicious conditions, is it really possible for SCNT to preserve a diploid genome? Delightfully, in mouse and recently in primate, using SCNT we can produce nuclear transfer ES cells (ntES) more efficiently, which can preserve the eternal lifespan for the "passage" of a unique diploid genome. Further, new somatic cloning technique using histone-deacetylase inhibitors has been developed which can significantly increase the previous cloning rates two to six times. Here, we introduce SCNT and its value as a preservation tool for a diploid genome while reviewing aging of cloned animals on cellular and individual levels. PMID:18346729

  15. Mass transfer and transport of radionuclides through backfill in a geologic nuclear waster repository

    SciTech Connect

    Kang, Chulhyung.

    1989-01-01

    Analytical models for the mass transfer and transport of radionuclides through waste package backfill material are developed and their numerical illustrations are demonstrated. These models can aid in the performance assessment of the waste package in nuclear waste repositories. The following analyses are considered to study the performance of the backfill barrier in a geologic nuclear waste repository. (1) The analysis for time and space dependent mass transfer and mass transport of a radioactive decay chain for a congruent release with the matrix species is studied. Non-recursive analytical solutions are derived for a semi-infinite medium and a finite medium. The theory is illustrated with the important decay chain {sup 234}U {yields} {sup 230}Th {yields} {sup 226}Ra. (2) The time and space dependent mass transport for high solubility nuclides, such as iodine or cesium, is analyzed. The approximate solution that is valid for all times is developed, and validated by comparison with an asymptotic solution and the solution obtained by the numerical inversion of a Laplace transform covering the entire time span. From this model one can determine the thickness of the backfill to satisfy a prescribed release rate limit for high soluble radionuclides. (3) The analysis of the transient radionuclide migration through the backfill into a fracture intersecting the waste form is developed. The nuclides are advected by water flowing in the fracture. The numerical illustrations show that the species transport for the short half-life nuclide can be stopped effectively within the backfill, but the backfill can no longer retard the migration for stable nuclides or long half-life nuclides after some initial time period. (4) The steady state mass transfer from a cylindrical waste form with flow through the backfill layer is examined.

  16. Transfer of 137Cs from Chernobyl debris and nuclear weapons fallout to different Swedish population groups.

    PubMed

    Rääf, C L; Hubbard, L; Falk, R; Agren, G; Vesanen, R

    2006-08-15

    Data from measurements on the body burden of (134)Cs, (137)Cs and (40)K in various Swedish populations between 1959 and 2001 has been compiled into a national database. The compilation is a co-operation between the Departments of Radiation Physics in Malmö and Göteborg, the National Radiation Protection Authority (SSI) and the Swedish Defense Research Agency (FOI). In a previous study the effective ecological half time and the associated effective dose to various Swedish populations due to internal contamination of (134)Cs and (137)Cs have been assessed using the database. In this study values of human body burden have been combined with data on the local and regional ground deposition of fallout from nuclear weapons tests (only (137)Cs) and Chernobyl debris (both (134)Cs and (137)Cs), which have enabled estimates of the radioecological transfer in the studied populations. The assessment of the database shows that the transfer of radiocesium from Chernobyl fallout to humans varies considerably between various populations in Sweden. In terms of committed effective dose over a 70 y period from internal contamination per unit activity deposition, the general (predominantly urban) Swedish population obtains 20-30 microSv/kBq m(-2). Four categories of populations exhibit higher radioecological transfer than the general population; i.) reindeer herders ( approximately 700 microSv/kBq m(-2)), ii.) hunters in the counties dominated by forest vegetation ( approximately 100 microSv/kBq m(-2)), iii.) rural non-farming populations living in sub-arctic areas (40-150 microSv/kBq m(-2)), and iv.) farmers ( approximately 50 microSv/kBq m(-2)). Two important factors determine the aggregate transfer from ground deposition to man; i.) dietary habits (intakes of foodstuff originating from natural and semi-natural ecosystems), and ii.) inclination to follow the recommended food restriction by the authorities. The transfer to the general population is considerably lower

  17. Application of Direct Assessment Approaches and Methodologies to Cathodically Protected Nuclear Waste Transfer Lines

    SciTech Connect

    Dahl, Megan M.; Pikas, Joseph; Edgemon, Glenn L.; Philo, Sarah

    2013-01-22

    The U.S. Department of Energy's (DOE) Hanford Site is responsible for the safe storage, retrieval, treatment, and disposal of approximately 54 million gallons (204 million liters) of radioactive waste generated since the site's inception in 1943. Today, the major structures involved in waste management at Hanford include 149 carbon steel single-shell tanks, 28 carbon-steel double-shell tanks, plus a network of buried metallic transfer lines and ancillary systems (pits, vaults, catch tanks, etc.) required to store, retrieve, and transfer waste within the tank farm system. Many of the waste management systems at Hanford are still in use today. In response to uncertainties regarding the structural integrity of these systems,' an independent, comprehensive integrity assessment of the Hanford Site piping system was performed. It was found that regulators do not require the cathodically protected pipelines located within the Hanford Site to be assessed by External Corrosion Direct Assessment (ECDA) or any other method used to ensure integrity. However, a case study is presented discussing the application of the direct assessment process on pipelines in such a nuclear environment. Assessment methodology and assessment results are contained herein. An approach is described for the monitoring, integration of outside data, and analysis of this information in order to identify whether coating deterioration accompanied by external corrosion is a threat for these waste transfer lines.

  18. Piglets born from vitrified cloned blastocysts produced with a simplified method of delipation and nuclear transfer.

    PubMed

    Du, Yutao; Li, Juan; Kragh, Peter M; Zhang, Yunhai; Schmidt, Mette; Bøgh, Ingrid B; Zhang, Xiuqing; Purup, Stig; Kuwayama, M; Jørgensen, Arne L; Pedersen, Anette M; Villemoes, Klaus; Yang, Huanming; Bolund, Lars; Vajta, Gábor

    2007-01-01

    Successful cryopreservation of porcine embryos offers a promising perspective in the fields of agriculture, animal science, and human medical research. The objective of the present work was to establish a system facilitating the cryopreservation of porcine embryos produced by somatic cell nuclear transfer (SCNT). Several key techniques including micromanipulator-based enucleation, noninvasive delipation, zona-free fusion, and activation were combined with high efficiency. After a partial zona digestion and high-speed centrifugation, 89.8+/-2.1% (mean+/-SEM) of enucleated oocytes were successfully delipated. Delipated cytoplasts were incubated for an additional 0.5 or 2 h before fusion with somatic cells. After activation and 6 days of in vitro culture, no significant difference in the rate of blastocysts per reconstructed embryo was observed between the two groups (33.1+/-1.8% and 26.0+/-4.3% for 0.5 and 2 h recovery time, respectively). Cryopreservation of the blastocysts was performed with a Cryotop device and factory-prepared vitrification and warming solutions. One hundred fifty-five vitrified SCNT embryos were transferred surgically into two recipient sows to test their developmental capacity in vivo. One recipient became pregnant and delivered six piglets. In conclusion, our simplified delipation and SCNT procedure resulted in viable piglets after vitrification and embryo transfer at the blastocyst stage. PMID:18154508

  19. Unusual horizontal transfer of a long interspersed nuclear element between distant vertebrate classes.

    PubMed

    Kordis, D; Gubensek, F

    1998-09-01

    We have shown previously by Southern blot analysis that Bov-B long interspersed nuclear elements (LINEs) are present in different Viperidae snake species. To address the question as to whether Bov-B LINEs really have been transmitted horizontally between vertebrate classes, the analysis has been extended to a larger number of vertebrate, invertebrate, and plant species. In this paper, the evolutionary origin of Bov-B LINEs is shown unequivocally to be in Squamata. The previously proposed horizontal transfer of Bov-B LINEs in vertebrates has been confirmed by their discontinuous phylogenetic distribution in Squamata (Serpentes and two lizard infra-orders) as well as in Ruminantia, by the high level of nucleotide identity, and by their phylogenetic relationships. The horizontal transfer of Bov-B LINEs from Squamata to the ancestor of Ruminantia is evident from the genetic distances and discontinuous phylogenetic distribution. The ancestor of Colubroidea snakes is a possible donor of Bov-B LINEs to Ruminantia. The timing of horizontal transfer has been estimated from the distribution of Bov-B LINEs in Ruminantia and the fossil data of Ruminantia to be 40-50 My ago. The phylogenetic relationships of Bov-B LINEs from the various Squamata species agrees with that of the species phylogeny, suggesting that Bov-B LINEs have been maintained stably by vertical transmission since the origin of Squamata in the Mesozoic era.

  20. International remote monitoring project Argentina Nuclear Power Station Spent Fuel Transfer Remote Monitoring System

    SciTech Connect

    Schneider, S.; Lucero, R.; Glidewell, D.

    1997-08-01

    The Autoridad Regulataria Nuclear (ARN) and the United States Department of Energy (DOE) are cooperating on the development of a Remote Monitoring System for nuclear nonproliferation efforts. A Remote Monitoring System for spent fuel transfer will be installed at the Argentina Nuclear Power Station in Embalse, Argentina. The system has been designed by Sandia National Laboratories (SNL), with Los Alamos National Laboratory (LANL) and Oak Ridge National Laboratory (ORNL) providing gamma and neutron sensors. This project will test and evaluate the fundamental design and implementation of the Remote Monitoring System in its application to regional and international safeguards efficiency. This paper provides a description of the monitoring system and its functions. The Remote Monitoring System consists of gamma and neutron radiation sensors, RF systems, and video systems integrated into a coherent functioning whole. All sensor data communicate over an Echelon LonWorks Network to a single data logger. The Neumann DCM 14 video module is integrated into the Remote Monitoring System. All sensor and image data are stored on a Data Acquisition System (DAS) and archived and reviewed on a Data and Image Review Station (DIRS). Conventional phone lines are used as the telecommunications link to transmit on-site collected data and images to remote locations. The data and images are authenticated before transmission. Data review stations will be installed at ARN in Buenos Aires, Argentina, ABACC in Rio De Janeiro, IAEA Headquarters in Vienna, and Sandia National Laboratories in Albuquerque, New Mexico. 2 refs., 2 figs.

  1. Explaining dehumanization among children: the interspecies model of prejudice.

    PubMed

    Costello, Kimberly; Hodson, Gordon

    2014-03-01

    Although many theoretical approaches have emerged to explain prejudices expressed by children, none incorporate outgroup dehumanization, a key predictor of prejudice among adults. According to the Interspecies Model of Prejudice, beliefs in the human-animal divide facilitate outgroup prejudice through fostering animalistic dehumanization (Costello & Hodson, 2010). In the present investigation, White children attributed Black children fewer 'uniquely human' characteristics, representing the first systematic evidence of racial dehumanization among children (Studies 1 and 2). In Study 2, path analyses supported the Interspecies Model of Prejudice: children's human-animal divide beliefs predicted greater racial prejudice, an effect explained by heightened racial dehumanization. Similar patterns emerged among parents. Furthermore, parent Social Dominance Orientation predicted child prejudice indirectly through children's endorsement of a hierarchical human-animal divide and subsequent dehumanizing tendencies. Encouragingly, children's human-animal divide perceptions were malleable to an experimental prime highlighting animal-human similarity. Implications for prejudice interventions are considered.

  2. Control of interspecies electron flow during anaerobic digestion: role of floc formation in syntrophic methanogenesis

    SciTech Connect

    Thiele, J.H.; Chartrain, M.; Zeikus, J.G.

    1988-01-01

    The flora of an anaerobic whey-processing chemostat was separated by anaerobic sedimentation techniques into a free-living bacterial fraction and a bacterial floc fraction. The floc fraction constituted a major part (i.e., 57% total protein) of the total microbial population in the digestor, and it accounted for 87% of the total CO/sub 2/-dependent methanogenic activity and 76% of the total ethanol-consuming acetogenic activity. Lactose was degraded by both cellular fractions, but in the free flora fraction it was associated with higher intermediary levels of H/sub 2/, ethanol, butyrate, and propionate production. Electron microscopic analysis of flocs showed bacterial diversity and juxtapositioning of tentative Desulfovibrio and Methanobacterium species without significant microcolony formation. Ethanol, an intermediary product of lactose-hydrolyzing bacteria, was converted to acetate and methane within the flocs by interspecies electron transfer. Ethanol-dependent methane formation was compartmentalized and closely coupled kinetically within the flocs but without significant formation of H/sub 2/ gas. Physical disruption of flocs into fragments of 10- to 20-..mu..m diameter initially increased the H/sub 2/ partial pressure but did not change the carbon transformation kinetic patterns of ethanol metabolism or demonstrate a significant role for H/sub 2/ in CO/sub 2/ reduction to methane. The data demonstrate that floc formation in a whey-processing anaerobic digestor functions in juxtapositioning cells for interspecies electron transfer during syntrophic ethanol conversion into acetate and methane but by a mechanism which was independent of the available dissolved H/sub 2/ gas pool in the ecosystem.

  3. Mechanisms Underlying Mammalian Hybrid Sterility in Two Feline Interspecies Models

    PubMed Central

    Davis, Brian W.; Seabury, Christopher M.; Brashear, Wesley A.; Li, Gang; Roelke-Parker, Melody; Murphy, William J.

    2015-01-01

    The phenomenon of male sterility in interspecies hybrids has been observed for over a century, however, few genes influencing this recurrent phenotype have been identified. Genetic investigations have been primarily limited to a small number of model organisms, thus limiting our understanding of the underlying molecular basis of this well-documented “rule of speciation.” We utilized two interspecies hybrid cat breeds in a genome-wide association study employing the Illumina 63 K single-nucleotide polymorphism array. Collectively, we identified eight autosomal genes/gene regions underlying associations with hybrid male sterility (HMS) involved in the function of the blood-testis barrier, gamete structural development, and transcriptional regulation. We also identified several candidate hybrid sterility regions on the X chromosome, with most residing in close proximity to complex duplicated regions. Differential gene expression analyses revealed significant chromosome-wide upregulation of X chromosome transcripts in testes of sterile hybrids, which were enriched for genes involved in chromatin regulation of gene expression. Our expression results parallel those reported in Mus hybrids, supporting the “Large X-Effect” in mammalian HMS and the potential epigenetic basis for this phenomenon. These results support the value of the interspecies feline model as a powerful tool for comparison to rodent models of HMS, demonstrating unique aspects and potential commonalities that underpin mammalian reproductive isolation. PMID:26006188

  4. Mechanisms Underlying Mammalian Hybrid Sterility in Two Feline Interspecies Models.

    PubMed

    Davis, Brian W; Seabury, Christopher M; Brashear, Wesley A; Li, Gang; Roelke-Parker, Melody; Murphy, William J

    2015-10-01

    The phenomenon of male sterility in interspecies hybrids has been observed for over a century, however, few genes influencing this recurrent phenotype have been identified. Genetic investigations have been primarily limited to a small number of model organisms, thus limiting our understanding of the underlying molecular basis of this well-documented "rule of speciation." We utilized two interspecies hybrid cat breeds in a genome-wide association study employing the Illumina 63 K single-nucleotide polymorphism array. Collectively, we identified eight autosomal genes/gene regions underlying associations with hybrid male sterility (HMS) involved in the function of the blood-testis barrier, gamete structural development, and transcriptional regulation. We also identified several candidate hybrid sterility regions on the X chromosome, with most residing in close proximity to complex duplicated regions. Differential gene expression analyses revealed significant chromosome-wide upregulation of X chromosome transcripts in testes of sterile hybrids, which were enriched for genes involved in chromatin regulation of gene expression. Our expression results parallel those reported in Mus hybrids, supporting the "Large X-Effect" in mammalian HMS and the potential epigenetic basis for this phenomenon. These results support the value of the interspecies feline model as a powerful tool for comparison to rodent models of HMS, demonstrating unique aspects and potential commonalities that underpin mammalian reproductive isolation. PMID:26006188

  5. Mechanisms Underlying Mammalian Hybrid Sterility in Two Feline Interspecies Models.

    PubMed

    Davis, Brian W; Seabury, Christopher M; Brashear, Wesley A; Li, Gang; Roelke-Parker, Melody; Murphy, William J

    2015-10-01

    The phenomenon of male sterility in interspecies hybrids has been observed for over a century, however, few genes influencing this recurrent phenotype have been identified. Genetic investigations have been primarily limited to a small number of model organisms, thus limiting our understanding of the underlying molecular basis of this well-documented "rule of speciation." We utilized two interspecies hybrid cat breeds in a genome-wide association study employing the Illumina 63 K single-nucleotide polymorphism array. Collectively, we identified eight autosomal genes/gene regions underlying associations with hybrid male sterility (HMS) involved in the function of the blood-testis barrier, gamete structural development, and transcriptional regulation. We also identified several candidate hybrid sterility regions on the X chromosome, with most residing in close proximity to complex duplicated regions. Differential gene expression analyses revealed significant chromosome-wide upregulation of X chromosome transcripts in testes of sterile hybrids, which were enriched for genes involved in chromatin regulation of gene expression. Our expression results parallel those reported in Mus hybrids, supporting the "Large X-Effect" in mammalian HMS and the potential epigenetic basis for this phenomenon. These results support the value of the interspecies feline model as a powerful tool for comparison to rodent models of HMS, demonstrating unique aspects and potential commonalities that underpin mammalian reproductive isolation.

  6. Natural interspecies recombinant bovine/porcine enterovirus in sheep.

    PubMed

    Boros, Akos; Pankovics, Péter; Knowles, Nick J; Reuter, Gábor

    2012-09-01

    Members of the genus Enterovirus (family Picornaviridae) are believed to be common and widespread among humans and different animal species, although only a few enteroviruses have been identified from animal sources. Intraspecies recombination among human enteroviruses is a well-known phenomenon, but only a few interspecies examples have been reported and, to our current knowledge, none of these have involved non-primate enteroviruses. In this study, we report the detection and complete genome characterization (using RT-PCR and long-range PCR) of a natural interspecies recombinant bovine/porcine enterovirus (ovine enterovirus type 1; OEV-1) in seven (44 %) of 16 faecal samples from 3-week-old domestic sheep (Ovis aries) collected in two consecutive years. Phylogenetic analysis of the complete coding region revealed that OEV-1 (ovine/TB4-OEV/2009/HUN; GenBank accession no. JQ277724) was a novel member of the species Porcine enterovirus B (PEV-B), implying the endemic presence of PEV-B viruses among sheep. However, the 5' UTR of OEV-1 showed a high degree of sequence and structural identity to bovine enteroviruses. The presumed recombination breakpoint was mapped to the end of the 5' UTR at nucleotide position 814 using sequence and SimPlot analyses. The interspecies-recombinant nature of OEV-1 suggests a closer relationship among bovine and porcine enteroviruses, enabling the exchange of at least some modular genetic elements that may evolve independently.

  7. Nuclear transfer of adult and genetically modified fetal cells of the rat.

    PubMed

    Hayes, E; Galea, S; Verkuylen, A; Pera, M; Morrison, J; Lacham-Kaplan, O; Trounson, A

    2001-04-27

    The present study examines the handling, activation, and micromanipulation of rat eggs in an attempt to produce live young using nuclear transfer (NT) of adult and genetically modified rat fetal cells. Mature rat eggs cultured in calcium-free medium showed reduced rates (24%) of chromosomal dispersion ("spontaneous activation" characteristic of this species) compared with eggs cultured in calcium-containing medium (47%), but failed to survive micromanipulation procedures. High rates of parthenogenetic cleavage were obtained with chemical activation using ethanol/cycloheximide (65%) compared with other standard chemical activation methods (4-28%). This type of activation was also effective in reestablishing cleavage capability (19-71%), in a time-dependent manner, of spontaneously activated eggs arrested at a second prophase-like state. At most, two of four tested micromanipulation procedures were effective in producing NT embryos capable of morula or blastocyst development (14-16%) in vivo following transfer to mouse oviducts. NT blastocysts produced from cumulus cells and transfected rat fetal fibroblasts appeared morphologically and karyotypically normal (2n = 42). Nocodazole-assisted metaphase enucleation and piezoelectric-assisted donor cell injection produced significant and equivocal effects on survival and cleavage rates of reconstructed embryos but failed to significantly improve in vivo morula/blastocyst development rates (16-28%) compared with unassisted micromanipulation (16%). Live births have not yet been obtained from early cleavage stage embryos (n = 269) transferred to pseudopregnant recipient rat oviducts. Improvements in reconstituted NT embryo culture and transfer are required for these methods to be an effective means of transgenic rat production.

  8. Heat Transfer Enhancement By Three-Dimensional Surface Roughness Technique In Nuclear Fuel Rod Bundles

    NASA Astrophysics Data System (ADS)

    Najeeb, Umair

    This thesis experimentally investigates the enhancement of single-phase heat transfer, frictional loss and pressure drop characteristics in a Single Heater Element Loop Tester (SHELT). The heater element simulates a single fuel rod for Pressurized Nuclear reactor. In this experimental investigation, the effect of the outer surface roughness of a simulated nuclear rod bundle was studied. The outer surface of a simulated fuel rod was created with a three-dimensional (Diamond-shaped blocks) surface roughness. The angle of corrugation for each diamond was 45 degrees. The length of each side of a diamond block is 1 mm. The depth of each diamond block was 0.3 mm. The pitch of the pattern was 1.614 mm. The simulated fuel rod had an outside diameter of 9.5 mm and wall thickness of 1.5 mm and was placed in a test-section made of 38.1 mm inner diameter, wall thickness 6.35 mm aluminum pipe. The Simulated fuel rod was made of Nickel 200 and Inconel 625 materials. The fuel rod was connected to 10 KW DC power supply. The Inconel 625 material of the rod with an electrical resistance of 32.3 kO was used to generate heat inside the test-section. The heat energy dissipated from the Inconel tube due to the flow of electrical current flows into the working fluid across the rod at constant heat flux conditions. The DI water was employed as working fluid for this experimental investigation. The temperature and pressure readings for both smooth and rough regions of the fuel rod were recorded and compared later to find enhancement in heat transfer coefficient and increment in the pressure drops. Tests were conducted for Reynold's Numbers ranging from 10e4 to 10e5. Enhancement in heat transfer coefficient at all Re was recorded. The maximum heat transfer co-efficient enhancement recorded was 86% at Re = 4.18e5. It was also observed that the pressure drop and friction factor increased by 14.7% due to the increased surface roughness.

  9. Probing and Exploiting the Interplay between Nuclear and Electronic Motion in Charge Transfer Processes.

    PubMed

    Delor, Milan; Sazanovich, Igor V; Towrie, Michael; Weinstein, Julia A

    2015-04-21

    The Born-Oppenheimer approximation refers to the assumption that the nuclear and electronic wave functions describing a molecular system evolve and can be determined independently. It is now well-known that this approximation often breaks down and that nuclear-electronic (vibronic) coupling contributes greatly to the ultrafast photophysics and photochemistry observed in many systems ranging from simple molecules to biological organisms. In order to probe vibronic coupling in a time-dependent manner, one must use spectroscopic tools capable of correlating the motions of electrons and nuclei on an ultrafast time scale. Recent developments in nonlinear multidimensional electronic and vibrational spectroscopies allow monitoring both electronic and structural factors with unprecedented time and spatial resolution. In this Account, we present recent studies from our group that make use of different variants of frequency-domain transient two-dimensional infrared (T-2DIR) spectroscopy, a pulse sequence combining electronic and vibrational excitations in the form of a UV-visible pump, a narrowband (12 cm(-1)) IR pump, and a broadband (400 cm(-1)) IR probe. In the first example, T-2DIR is used to directly compare vibrational dynamics in the ground and relaxed electronic excited states of Re(Cl)(CO)3(4,4'-diethylester-2,2'-bipyridine) and Ru(4,4'-diethylester-2,2'-bipyridine)2(NCS)2, prototypical charge transfer complexes used in photocatalytic CO2 reduction and electron injection in dye-sensitized solar cells. The experiments show that intramolecular vibrational redistribution (IVR) and vibrational energy transfer (VET) are up to an order of magnitude faster in the triplet charge transfer excited state than in the ground state. These results show the influence of electronic arrangement on vibrational coupling patterns, with direct implications for vibronic coupling mechanisms in charge transfer excited states. In the second example, we show unambiguously that electronic and

  10. Large scale in vivo risk assessment of bovine viral diarrhea virus (BVDV) transmission through transfer of bovine embryos produced via somatic cell nuclear transfer (SCNT).

    PubMed

    Gregg, K; Gosch, G; Guerra, T; Chen, S H; Xiang, T; Broek, D; Bruner, B; Polejaeva, I

    2010-10-15

    The objective was to use the bovine viral diarrhea virus (BVDV) as a model to assess the risk of infectious disease transmission in the system of in vitro embryo production and transfer via somatic cell nuclear transfer (SCNT) technology. The risks of BVDV transmission in the SCNT embryo production were previously evaluated. In that in vitro study, following standard operating procedures (SOP), including pre-nuclear transfer donor cell testing, oocyte decontamination and virus-free cell and embryo culture conditions, SCNT embryos produced were free of detectable viral RNA. The current study focused on the evaluation of the potential risk of disease transmission from SCNT embryos to recipients, and the risk of producing persistently infected animals via SCNT embryo transfer. Blood samples were collected from 553 recipients of SCNT embryos and 438 cloned calves and tested for the presence of BVDV viral RNA via a sensitive real time PCR method. All samples tested were negative. These results, in conjunction with the previous in vitro study, confirmed that the established SCNT embryo production and transfer system is safe and presents no detectable risk of disease transmission.

  11. Genomic Stability of Lyophilized Sheep Somatic Cells before and after Nuclear Transfer

    PubMed Central

    Iuso, Domenico; Czernik, Marta; Di Egidio, Fiorella; Sampino, Silvestre; Zacchini, Federica; Bochenek, Michal; Smorag, Zdzislaw; Modlinski, Jacek A.; Ptak, Grazyna; Loi, Pasqualino

    2013-01-01

    The unprecedented decline of biodiversity worldwide is urging scientists to collect and store biological material from seriously threatened animals, including large mammals. Lyophilization is being explored as a low-cost system for storage in bio-banks of cells that might be used to expand or restore endangered or extinct species through the procedure of Somatic Cell Nuclear Transfer (SCNT). Here we report that the genome is intact in about 60% of lyophylized sheep lymphocytes, whereas DNA damage occurs randomly in the remaining 40%. Remarkably, lyophilized nuclei injected into enucleated oocytes are repaired by a robust DNA repairing activity of the oocytes, and show normal developmental competence. Cloned embryos derived from lyophylized cells exhibited chromosome and cellular composition comparable to those of embryos derived from fresh donor cells. These findings support the feasibility of lyophylization as a storage procedure of mammalian cells to be used for SCNT. PMID:23308098

  12. Alternate nuclear transfer is no alternative for embryonic stem cell research.

    PubMed

    Fennel, John A

    2008-02-01

    Recent developments allow for the creation of human stem cells without the creation of human embryos, a process called alternate nuclear transfer ('ANT'). Pursuing this method of stem cell research makes sense for pro-lifers if arguments for the sanctity of the human embryo do not apply to ANT. However, the technology that makes ANT possible undermines the erstwhile technical barrier between human embryos and somatic cell DNA. These advances bring home the force of hypothetical arguments about the potential of the DNA in somatic cells, showing that there is not a morally relevant difference between the potential of an embryo and the potential of the DNA in a somatic cell. Therefore, the supposed distinction between entities that are potential human life and entities that are human life does not give any support to arguments for the sanctity of the human embryo because those arguments extend value to too many entities. PMID:18251768

  13. Femtosecond laser based enucleation of porcine oocytes for somatic cell nuclear transfer

    NASA Astrophysics Data System (ADS)

    Kütemeyer, K.; Lucas-Hahn, A.; Petersen, B.; Hassel, P.; Lemme, E.; Niemann, H.; Heisterkamp, A.

    2009-07-01

    Cloning of several mammalian species has been achieved by somatic cell nuclear transfer (SCNT) in recent years. However, this method still results in very low efficiencies around 1% which originate from suboptimal culture conditions and highly invasive techniques for oocyte enucleation and injection of the donor cell using micromanipulators. In this paper, we present a new minimal invasive method for oocyte imaging and enucleation based on the application of femtosecond (fs) laser pulses. After imaging of the oocyte with multiphoton microscopy, ultrashort pulses are focused onto the metaphase plate of MII-oocytes in order to ablate the DNA molecules. We show that fs laser based enucleation of porcine oocytes completely inhibits the first mitotic cleavage after parthenogenetic activation while maintaining intact oocyte morphology in most cases. In contrast, control groups without previous irradiation of the metaphase plate are able to develop to the blastocyst stage. Further experiments have to clarify the suitability of fs laser based enucleated oocytes for SCNT.

  14. Differentiated cells are more efficient than adult stem cells for cloning by somatic cell nuclear transfer.

    PubMed

    Sung, Li-Ying; Gao, Shaorong; Shen, Hongmei; Yu, Hui; Song, Yifang; Smith, Sadie L; Chang, Ching-Chien; Inoue, Kimiko; Kuo, Lynn; Lian, Jin; Li, Ao; Tian, X Cindy; Tuck, David P; Weissman, Sherman M; Yang, Xiangzhong; Cheng, Tao

    2006-11-01

    Since the creation of Dolly via somatic cell nuclear transfer (SCNT), more than a dozen species of mammals have been cloned using this technology. One hypothesis for the limited success of cloning via SCNT (1%-5%) is that the clones are likely to be derived from adult stem cells. Support for this hypothesis comes from the findings that the reproductive cloning efficiency for embryonic stem cells is five to ten times higher than that for somatic cells as donors and that cloned pups cannot be produced directly from cloned embryos derived from differentiated B and T cells or neuronal cells. The question remains as to whether SCNT-derived animal clones can be derived from truly differentiated somatic cells. We tested this hypothesis with mouse hematopoietic cells at different differentiation stages: hematopoietic stem cells, progenitor cells and granulocytes. We found that cloning efficiency increases over the differentiation hierarchy, and terminally differentiated postmitotic granulocytes yield cloned pups with the greatest cloning efficiency.

  15. Somatic cell nuclear transfer and transgenesis in large animals: current and future insights.

    PubMed

    Galli, C; Lagutina, I; Perota, A; Colleoni, S; Duchi, R; Lucchini, F; Lazzari, G

    2012-06-01

    Somatic cell nuclear transfer (SCNT) was first developed in livestock for the purpose of accelerating the widespread use of superior genotypes. Although many problems still exist now after fifteen years of research owing to the limited understanding of genome reprogramming, SCNT has provided a powerful tool to make copies of selected individuals in different species, to study genome pluripotency and differentiation, opening new avenues of research in regenerative medicine and representing the main route for making transgenic livestock. Besides well-established methods to deliver transgenes, recent development in enzymatic engineering to edit the genome provides more precise and reproducible tools to target-specific genomic loci especially for producing knockout animals. The interest in generating transgenic livestock lies in the agricultural and biomedical areas and it is, in most cases, at the stage of research and development, with few exceptions that are making the way into practical applications.

  16. In vitro development of canine somatic cell nuclear transfer embryos in different culture media.

    PubMed

    Kim, Dong-Hoon; No, Jin-Gu; Choi, Mi-Kyung; Yeom, Dong-Hyeon; Kim, Dong-Kyo; Yang, Byoung-Chul; Yoo, Jae Gyu; Kim, Min Kyu; Kim, Hong-Tea

    2015-01-01

    The objective of the present study was to investigate the effects of three different culture media on the development of canine somatic cell nuclear transfer (SCNT) embryos. Canine cloned embryos were cultured in modified synthetic oviductal fluid (mSOF), porcine zygote medium-3 (PZM-3), or G1/G2 sequential media. Our results showed that the G1/G2 media yielded significantly higher morula and blastocyst development in canine SCNT embryos (26.1% and 7.8%, respectively) compared to PZM-3 (8.5% and 0%or mSOF (2.3% and 0%) media. In conclusion, this study suggests that blastocysts can be produced more efficiently using G1/G2 media to culture canine SCNT embryos.

  17. Modeling Electronic-Nuclear Interactions for Excitation Energy Transfer Processes in Light-Harvesting Complexes.

    PubMed

    Lee, Mi Kyung; Coker, David F

    2016-08-18

    An accurate approach for computing intermolecular and intrachromophore contributions to spectral densities to describe the electronic-nuclear interactions relevant for modeling excitation energy transfer processes in light harvesting systems is presented. The approach is based on molecular dynamics (MD) calculations of classical correlation functions of long-range contributions to excitation energy fluctuations and a separate harmonic analysis and single-point gradient quantum calculations for electron-intrachromophore vibrational couplings. A simple model is also presented that enables detailed analysis of the shortcomings of standard MD-based excitation energy fluctuation correlation function approaches. The method introduced here avoids these problems, and its reliability is demonstrated in accurate predictions for bacteriochlorophyll molecules in the Fenna-Matthews-Olson pigment-protein complex, where excellent agreement with experimental spectral densities is found. This efficient approach can provide instantaneous spectral densities for treating the influence of fluctuations in environmental dissipation on fast electronic relaxation.

  18. Development of buffalo (Bubalus bubalis) embryonic stem cell lines from somatic cell nuclear transferred blastocysts.

    PubMed

    Shah, Syed Mohmad; Saini, Neha; Ashraf, Syma; Singh, Manoj K; Manik, Radheysham; Singla, Suresh K; Palta, Prabhat; Chauhan, Manmohan Singh

    2015-11-01

    We developed buffalo embryonic stem cell lines from somatic cell nuclear transfer derived blastocysts, produced by hand-guided cloning technique. The inner cell mass of the blastocyst was cut mechanically using a Microblade and cultured onto feeder cells in buffalo embryonic stem (ES) cell culture medium at 38 °C in a 5% CO2 incubator. The stem cell colonies were characterized for alkaline phosphatase activity, karyotype, pluripotency and self-renewal markers like OCT4, NANOG, SOX2, c-Myc, FOXD3, SSEA-1, SSEA-4, TRA-1-60, TRA-1-81 and CD90. The cell lines also possessed the capability to differentiate across all the three germ layers under spontaneous differentiation conditions. PMID:26987926

  19. Murine somatic cell nuclear transfer using reprogrammed donor cells expressing male germ cell-specific genes.

    PubMed

    Kang, Hoin; Park, Jong Im; Roh, Sangho

    2016-01-01

    In vivo-matured mouse oocytes were enucleated, and a single murine embryonic fibroblast (control or reprogrammed by introducing extracts from murine testis tissue, which showed expression of male germ cell-specific genes) was injected into the cytoplasm of the oocytes. The rate of blastocyst development and expression levels of Oct-4, Eomes and Cdx-2 were not significantly different in both experimental groups. However, the expression levels of Nanog, Sox9 and Glut-1 were significantly increased when reprogrammed cells were used as donor nuclei. Increased expression of Nanog can be supportive of complete reprogramming of somatic cell nuclear transfer murine embryos. The present study suggested that donor cells expressing male germ cell-specific genes can be reconstructed and can develop into embryos with normal high expression of developmentally essential genes. PMID:26369430

  20. Study of Two-Phase Heat Transfer in Nano-fluids for Nuclear Applications

    SciTech Connect

    Kim, S.J.; Truong, B.; Buongiorno, J.; Hu, L.W.; Bang, I.C.

    2006-07-01

    Nano-fluids are engineered colloidal suspensions of nano-particles in a base fluid. We are investigating the two-phase heat transfer behavior of water-based nano-fluids, to evaluate their potential use in nuclear applications, including the PWR primary coolant and PWR and BWR safety systems. A simple pool boiling wire experiment shows that a significant increase in Critical Heat Flux (CHF) can be achieved at modest nano-particle concentrations. For example, the CHF increases by 50% in nano-fluids with alumina nano-particles at 0.001%v concentration. The CHF enhancement appears to correlate with the presence of a layer of nano-particles that builds up on the heated surface during nucleate boiling. A review of the prevalent Departure from Nucleate Boiling (DNB) theories suggests that an alteration of the nucleation site density (brought about by the nano-particle layer) could plausibly explain the CHF enhancement. (authors)

  1. Modeling Electronic-Nuclear Interactions for Excitation Energy Transfer Processes in Light-Harvesting Complexes.

    PubMed

    Lee, Mi Kyung; Coker, David F

    2016-08-18

    An accurate approach for computing intermolecular and intrachromophore contributions to spectral densities to describe the electronic-nuclear interactions relevant for modeling excitation energy transfer processes in light harvesting systems is presented. The approach is based on molecular dynamics (MD) calculations of classical correlation functions of long-range contributions to excitation energy fluctuations and a separate harmonic analysis and single-point gradient quantum calculations for electron-intrachromophore vibrational couplings. A simple model is also presented that enables detailed analysis of the shortcomings of standard MD-based excitation energy fluctuation correlation function approaches. The method introduced here avoids these problems, and its reliability is demonstrated in accurate predictions for bacteriochlorophyll molecules in the Fenna-Matthews-Olson pigment-protein complex, where excellent agreement with experimental spectral densities is found. This efficient approach can provide instantaneous spectral densities for treating the influence of fluctuations in environmental dissipation on fast electronic relaxation. PMID:27472379

  2. Bioactive conformation of stromelysin inhibitors determined by transferred nuclear Overhauser effects.

    PubMed Central

    Gonnella, N C; Bohacek, R; Zhang, X; Kolossváry, I; Paris, C G; Melton, R; Winter, C; Hu, S I; Ganu, V

    1995-01-01

    The transferred nuclear Overhauser effect has been used to determine the biologically active conformations of two stromelysin inhibitors. Both inhibitors used in this study were hydroxamic acids generated via chemical synthesis. These structures, representing the conformation of each inhibitor bound to stromelysin, superimposed with excellent agreement. The study also provided information on the shape and orientation of the S2' and S1' pockets of the enzyme relative to thermolysin. Comparisons were made between stromelysin and thermolysin inhibitors to critically examine thermolysin as a template for stromelysin-inhibitor design. The enzyme-bound conformations of these stromelysin inhibitors were determined for use as a template in conformationally restricted drug design. Images Fig. 4 Fig. 5 Fig. 6 Fig. 7 PMID:7831311

  3. Live embryo imaging to follow cell cycle and chromosomes stability after nuclear transfer.

    PubMed

    Balbach, Sebastian T; Boiani, Michele

    2015-01-01

    Nuclear transfer (NT) into mouse oocytes yields a transcriptionally and functionally heterogeneous population of cloned embryos. Most studies of NT embryos consider only embryos at predefined key stages (e.g., morula or blastocyst), that is, after the bulk of reprogramming has taken place. These retrospective approaches are of limited use to elucidate mechanisms of reprogramming and to predict developmental success. Observing cloned embryo development using live embryo cinematography has the potential to reveal otherwise undetectable embryo features. However, light exposure necessary for live cell cinematography is highly toxic to cloned embryos. Here we describe a protocol for combined bright-field and fluorescence live-cell imaging of histone H2b-GFP expressing mouse embryos, to record cell divisions up to the blastocyst stage. This protocol, which can be adapted to observe other reporters such as Oct4-GFP or Nanog-GFP, allowed us to quantitatively analyze cleavage kinetics of cloned embryos. PMID:25287344

  4. Potential of primary kidney cells for somatic cell nuclear transfer mediated transgenesis in pig

    PubMed Central

    2012-01-01

    Background Somatic cell nuclear transfer (SCNT) is currently the most efficient and precise method to generate genetically tailored pig models for biomedical research. However, the efficiency of this approach is crucially dependent on the source of nuclear donor cells. In this study, we evaluate the potential of primary porcine kidney cells (PKCs) as cell source for SCNT, including their proliferation capacity, transfection efficiency, and capacity to support full term development of SCNT embryos after additive gene transfer or homologous recombination. Results PKCs could be maintained in culture with stable karyotype for up to 71 passages, whereas porcine fetal fibroblasts (PFFs) and porcine ear fibroblasts (PEFs) could be hardly passaged more than 20 times. Compared with PFFs and PEFs, PKCs exhibited a higher proliferation rate and resulted in a 2-fold higher blastocyst rate after SCNT and in vitro cultivation. Among the four transfection methods tested with a GFP expression plasmid, best results were obtained with the NucleofectorTM technology, resulting in transfection efficiencies of 70% to 89% with high fluorescence intensity, low cytotoxicity, good cell proliferation, and almost no morphological signs of cell stress. Usage of genetically modified PKCs in SCNT resulted in approximately 150 piglets carrying at least one of 18 different transgenes. Several of those pigs originated from PKCs that underwent homologous recombination and antibiotic selection before SCNT. Conclusion The high proliferation capacity of PKCs facilitates the introduction of precise and complex genetic modifications in vitro. PKCs are thus a valuable cell source for the generation of porcine biomedical models by SCNT. PMID:23140586

  5. Spent Nuclear Fuel Dry Transfer System Cold Demonstration Project Final Report

    SciTech Connect

    Christensen, Max R; McKinnon, M. A.

    1999-12-01

    The spent nuclear fuel dry transfer system (DTS) provides an interface between large and small casks and between storage-only and transportation casks. It permits decommissioning of reactor pools after shutdown and allows the use of large storage-only casks for temporary onsite storage of spent nuclear fuel irrespective of reactor or fuel handling limitations at a reactor site. A cold demonstration of the DTS prototype was initiated in August 1996 at the Idaho National Engineering and Environmental Laboratory (INEEL). The major components demonstrated included the fuel assembly handling subsystem, the shield plug/lid handling subsystem, the cask interface subsystem, the demonstration control subsystem, a support frame, and a closed circuit television and lighting system. The demonstration included a complete series of DTS operations from source cask receipt and opening through fuel transfer and closure of the receiving cask. The demonstration included both normal operations and recovery from off-normal events. It was designed to challenge the system to determine whether there were any activities that could be made to jeopardize the activities of another function or its safety. All known interlocks were challenged. The equipment ran smoothly and functioned as designed. A few "bugs" were corrected. Prior to completion of the demonstration testing, a number of DTS prototype systems were modified to apply lessons learned to date. Additional testing was performed to validate the modifications. In general, all the equipment worked exceptionally well. The demonstration also helped confirm cost estimates that had been made at several points in the development of the system.

  6. Improved efficiency of bovine cloning by autologous somatic cell nuclear transfer.

    PubMed

    Yang, Xiao-Yu; Li, Hua; Ma, Qing-Wen; Yan, Jing-Bin; Zhao, Jiang-Guo; Li, Hua-Wei; Shen, Hai-Qing; Liu, Hai-Feng; Huang, Ying; Huang, Shu-Zhen; Zeng, Yi-Tao; Zeng, Fanyi

    2006-11-01

    Somatic cell nuclear transfer (SCNT) has been used for the cloning of various mammals. However, the rates of successful, healthy birth are generally poor. To improve cloning efficiency, we report the utilization of an 'autologous SCNT' cloning technique in which the somatic nucleus of a female bovine donor is transferred to its own enucleated oocyte recovered by ovum pick up, in contrast to the routine 'allogeneic SCNT' procedure using oocytes from unrelated females. Our results showed that embryos derived from autologous SCNThave significantly higher developmental competence than those derived from allogeneic SCNT, especiallyat the eight-cell (60 vs 44%), morula (45 vs 36%), and blastocyst (38 vs 23%) stages. The pregnancy and birth rates were also higher for the autologous (39 and 23%), compared to the allogeneic (22 and 6%) SCNT groups. Genome-wide histone3-lysine9 methylation profiles reveal that autologous SCNTembryos have less epigenetic defects than the allogeneic SCNTembryos. This study indicates that autologous SCNT can improve the efficiency of bovine cloning with less reprogramming deficiency.

  7. Rhesus monkey embryos produced by nuclear transfer from embryonic blastomeres or somatic cells.

    PubMed

    Mitalipov, Shoukhrat M; Yeoman, Richard R; Nusser, Kevin D; Wolf, Don P

    2002-05-01

    Production of genetically identical nonhuman primates would reduce the number of animals required for biomedical research and dramatically impact studies pertaining to immune system function, such as development of the human-immunodeficiency-virus vaccine. Our long-term goal is to develop robust somatic cell cloning and/or twinning protocols in the rhesus macaque. The objective of this study was to determine the developmental competence of nuclear transfer (NT) embryos derived from embryonic blastomeres (embryonic cell NT) or fetal fibroblasts (somatic cell NT) as a first step in the production of rhesus monkeys by somatic cell cloning. Development of cleaved embryos up to the 8-cell stage was similar among embryonic and somatic cell NT embryos and comparable to controls created by intracytoplasmic sperm injection (ICSI; mean +/- SEM, 81 +/- 5%, 88 +/- 7%, and 87 +/- 4%, respectively). However, significantly lower rates of development to the blastocyst stage were observed with somatic cell NT embryos (1%) in contrast to embryonic cell NT (34 +/- 15%) or ICSI control embryos (46 +/- 6%). Development of somatic cell NT embryos was not markedly affected by donor cell treatment, timing of activation, or chemical activation protocol. Transfer of embryonic, but not of somatic cell NT embryos, into recipients resulted in term pregnancy. Future efforts will focus on optimizing the production of somatic cell NT embryos that develop in high efficiency to the blastocyst stage in vitro.

  8. Phenotypes of Aging Postovulatory Oocytes After Somatic Cell Nuclear Transfer in Mice.

    PubMed

    Lee, Ah Reum; Shimoike, Takashi; Wakayama, Teruhiko; Kishigami, Satoshi

    2016-06-01

    Oocytes rapidly lose their developmental potential after ovulation, termed postovulatory oocyte aging, and often exhibit characteristic phenotypes, such as cytofragmentation, abnormal spindle shapes, and chromosome misalignments. Here, we reconstructed mouse oocytes using somatic cell nuclear transfer (SCNT) to reveal the effect of somatic cell-derived nuclei on oocyte physiology during aging. Normal oocytes started undergoing cytofragmentation 24 hours after oocyte collection; however, this occurred earlier in SCNT oocytes and was more severe at 48 hours, suggesting that the transferred somatic cell nuclei affected oocyte physiology. We found no difference in the status of acetylated α-tubulin (Ac-Tub) and α-tubulin (Tub) between normal and SCNT aging oocytes, but unlike normal oocytes, aging SCNT oocytes did not have astral microtubules. Interestingly, aging SCNT oocytes displayed more severely scattered chromosomes or irregularly shaped spindles. Observations of the microfilaments showed that, in normal oocytes, there was a clear actin ring beneath the plasma membrane and condensed microfilaments around the spindle (the actin cap) at 0 hours, and the actin filaments started degenerating at 1 hour, becoming completely disrupted and distributed to the cytoplasm at 24 hours. By contrast, in SCNT oocytes, an actin cap formed around the transplanted nuclei within 1 hour of SCNT, which was still present at 24 hours. Thus, SCNT oocytes age in a similar but distinct way, suggesting that they not only contain nuclei with abnormal epigenetics but are also physiologically different. PMID:27253626

  9. Transfer-induced fission in inverse kinematics: Impact on experimental and evaluated nuclear data bases

    NASA Astrophysics Data System (ADS)

    Farget, F.; Caamaño, M.; Ramos, D.; Rodrıguez-Tajes, C.; Schmidt, K.-H.; Audouin, L.; Benlliure, J.; Casarejos, E.; Clément, E.; Cortina, D.; Delaune, O.; Derkx, X.; Dijon, A.; Doré, D.; Fernández-Domınguez, B.; Gaudefroy, L.; Golabek, C.; Heinz, A.; Jurado, B.; Lemasson, A.; Paradela, C.; Roger, T.; Salsac, M. D.; Schmitt, C.

    2015-12-01

    Inverse kinematics is a new tool to study nuclear fission. Its main advantage is the possibility to measure with an unmatched resolution the atomic number of fission fragments, leading to new observables in the properties of fission-fragment distributions. In addition to the resolution improvement, the study of fission based on nuclear collisions in inverse kinematics beneficiates from a larger view with respect to the neutron-induced fission, as in a single experiment the number of fissioning systems and the excitation energy range are widden. With the use of spectrometers, mass and kinetic-energy distributions may now be investigated as a function of the proton and neutron number sharing. The production of fissioning nuclei in transfer reactions allows studying the isotopic yields of fission fragments as a function of the excitation energy. The higher excitation energy resulting in the fusion reaction leading to the compound nucleus 250Cf at an excitation energy of 45MeV is also presented. With the use of inverse kinematics, the charge polarisation of fragments at scission is now revealed with high precision, and it is shown that it cannot be neglected, even at higher excitation energies. In addition, the kinematical properties of the fragments inform on the deformation configuration at scission.

  10. Multi-Scale-Structured Composite Coatings by Plasma-Transferred Arc for Nuclear Applications

    NASA Astrophysics Data System (ADS)

    Werry, A.; Chazelas, C.; Denoirjean, A.; Valette, S.; Vardelle, A.; Meillot, E.

    2016-01-01

    In nuclear plants, the replacement of hardfacing Stellite, a cobalt-based alloy, on parts of the piping system in connection with the reactor has been investigated since the late 60's. Various Fe-based or Ni-based alloys, Co-free or with a low content of Co, have been developed but with mechanical properties generally lower than that of Stellites. The 4th generation nuclear plants impose additional or more stringent requirements for hardfacing materials. Plasma-transferred arc (PTA) coatings of cobalt-free nickel-based alloys with the addition of sub-micrometric or micrometric alumina particles are thought to be a potential solution for tribological applications in the primary system of sodium-cooled fast reactors. In this study, PTA coatings of nickel-based alloys reinforced with alumina particles were deposited on 316L stainless steel substrates. Under the conditions of this study, the addition of alumina particles resulted in a refinement of coating microstructure and the improvement of their resistance to abrasive wear. However, it does not bring about any change in coating micro-hardness.

  11. Production efficiency and telomere length of the cloned pigs following serial somatic cell nuclear transfer.

    PubMed

    Kurome, Mayuko; Hisatomi, Hisashi; Matsumoto, Shirou; Tomii, Ryo; Ueno, Satoshi; Hiruma, Katsumi; Saito, Hitoshi; Nakamura, Kimitoshi; Okumura, Kenji; Matsumoto, Mitsuhito; Kaji, Yuji; Endo, Fumio; Nagashima, Hiroshi

    2008-08-01

    The aim of the present study was to examine the production efficiency of cloned pigs by serial somatic cell nuclear transfer (SCNT) and to ascertain any changes in the telomere lengths of multiple generations of pigs. Using fetal fibroblasts as the starting nuclear donor cells, porcine salivary gland progenitor cells were collected from the resultant first-generation cloned pigs to successively produce second- and third-generation clones, with no significant differences in production efficiency, which ranged from 1.4% (2/140) to 3.3% (13/391) among the 3 generations. The average telomere lengths (terminal restriction fragment values) for the first, second and third generation clones were 16.3, 18.1 and 20.5 kb, respectively, and were comparable to those in age-matched controls. These findings suggest that third-generation cloned pigs can be produced by serial somatic cell cloning without compromising production efficiency and that the telomere lengths of cloned pigs from the first to third generations are normal. PMID:18490858

  12. Inheritance of mitochondrial DNA in serially recloned pigs by somatic cell nuclear transfer (SCNT)

    SciTech Connect

    Do, Minhwa; Jang, Won-Gu; Hwang, Jeong Hee; Jang, Hoon; Kim, Eun-Jung; Jeong, Eun-Jeong; Shim, Hosup; Hwang, Sung Soo; Oh, Keon Bong; Byun, Sung June; Kim, Jin-Hoi; Lee, Jeong Woong

    2012-08-10

    Highlights: Black-Right-Pointing-Pointer We success serial SCNT through the third generation using pig fibroblasts. Black-Right-Pointing-Pointer Donor-specific mtDNA in the recloned pigs was detected. Black-Right-Pointing-Pointer SCNT affect mtDNA mounts. -- Abstract: Somatic cell nuclear transfer (SCNT) has been established for the transmission of specific nuclear DNA. However, the fate of donor mitochondrial DNA (mtDNA) remains unclear. Here, we examined the fate of donor mtDNA in recloned pigs through third generations. Fibroblasts of recloned pigs were obtained from offspring of each generation produced by fusion of cultured fibroblasts from a Minnesota miniature pig (MMP) into enucleated oocytes of a Landrace pig. The D-loop regions from the mtDNA of donor and recipient differ at nucleotide sequence positions 16050 (A{yields}T), 16062 (T{yields}C), and 16135 (G{yields}A). In order to determine the fate of donor mtDNA in recloned pigs, we analyzed the D-loop region of the donor's mtDNA by allele-specific PCR (AS-PCR) and real-time PCR. Donor mtDNA was successfully detected in all recloned offspring (F1, F2, and F3). These results indicate that heteroplasmy that originate from donor and recipient mtDNA is maintained in recloned pigs, resulting from SCNT, unlike natural reproduction.

  13. Longitudinal study of reproductive performance of female cattle produced by somatic cell nuclear transfer.

    PubMed

    Polejaeva, Irina A; Broek, Diane M; Walker, Shawn C; Zhou, Wenli; Walton, Mark; Benninghoff, Abby D; Faber, David C

    2013-01-01

    The objective of this study was to determine whether or not reproductive performance in cattle produced by somatic cell nuclear transfer (SCNT) is significantly different from that of their genetic donors. To address this question, we directed two longitudinal studies using different embryo production procedures: (1) superovulation followed by artificial insemination (AI) and embryo collection and (2) ultrasound-guided ovum pick-up followed by in vitro fertilization (OPU-IVF). Collectively, these two studies represent the largest data set available for any species on the reproductive performance of female clones and their genetic donors as measured by their embryo production outcomes in commercial embryo production program. The large-scale study described herein was conducted over a six-year period of time and provides a unique comparison of 96 clones to the 40 corresponding genetic donors. To our knowledge, this is the first longitudinal study on the reproductive performance of cattle clones using OPU-IVF. With nearly 2,000 reproductive procedures performed and more than 9,200 transferable embryos produced, our observations show that the reproductive performance of cattle produced by SCNT is not different compared to their genetic donors for the production of transferable embryos after either AI followed by embryo collection (P = 0.77) or OPU-IVF (P = 0.97). These data are in agreement with previous reports showing that the reproductive capabilities of cloned cattle are equal to that of conventionally produced cattle. In conclusion, results of this longitudinal study once again demonstrate that cloning technology, in combination with superovulation, AI and embryo collection or OPU-IVF, provides a valuable tool for faster dissemination of superior maternal genetics.

  14. Longitudinal Study of Reproductive Performance of Female Cattle Produced by Somatic Cell Nuclear Transfer

    PubMed Central

    Polejaeva, Irina A.; Broek, Diane M.; Walker, Shawn C.; Zhou, Wenli; Walton, Mark; Benninghoff, Abby D.; Faber, David C.

    2013-01-01

    The objective of this study was to determine whether or not reproductive performance in cattle produced by somatic cell nuclear transfer (SCNT) is significantly different from that of their genetic donors. To address this question, we directed two longitudinal studies using different embryo production procedures: (1) superovulation followed by artificial insemination (AI) and embryo collection and (2) ultrasound-guided ovum pick-up followed by in vitro fertilization (OPU-IVF). Collectively, these two studies represent the largest data set available for any species on the reproductive performance of female clones and their genetic donors as measured by their embryo production outcomes in commercial embryo production program. The large-scale study described herein was conducted over a six-year period of time and provides a unique comparison of 96 clones to the 40 corresponding genetic donors. To our knowledge, this is the first longitudinal study on the reproductive performance of cattle clones using OPU-IVF. With nearly 2,000 reproductive procedures performed and more than 9,200 transferable embryos produced, our observations show that the reproductive performance of cattle produced by SCNT is not different compared to their genetic donors for the production of transferable embryos after either AI followed by embryo collection (P = 0.77) or OPU-IVF (P = 0.97). These data are in agreement with previous reports showing that the reproductive capabilities of cloned cattle are equal to that of conventionally produced cattle. In conclusion, results of this longitudinal study once again demonstrate that cloning technology, in combination with superovulation, AI and embryo collection or OPU-IVF, provides a valuable tool for faster dissemination of superior maternal genetics. PMID:24391930

  15. Finite-thrust optimization of interplanetary transfers of space vehicle with bimodal nuclear thermal propulsion

    NASA Astrophysics Data System (ADS)

    Kharytonov, Oleksii M.; Kiforenko, Boris M.

    2011-08-01

    The nuclear thermal rocket (NTR) propulsion is one of the leading promising technologies for primary space propulsion for manned exploration of the solar system due to its high specific impulse capability and sufficiently high thrust-to-weight ratio. Another benefit of NTR is its possible bimodal design, when nuclear reactor is used for generation of a jet thrust in a high-thrust mode and (with an appropriate power conversion system) as a source of electric power to supply the payload and the electric engines in a low-thrust mode. The model of the NTR thrust control was developed considering high-thrust NTR as a propulsion system of limited power and exhaust velocity. For the proposed model the control of the thrust value is accomplished by the regulation of reactor thermal power and propellant mass flow rate. The problem of joint optimization of the combination of high- and low-thrust arcs and the parameters of bimodal NTR (BNTR) propulsion system is considered for the interplanetary transfers. The interplanetary trajectory of the space vehicle is formed by the high-thrust NTR burns, which define planet-centric maneuvers and by the low-thrust heliocentric arcs where the nuclear electric propulsion (NEP) is used. The high-thrust arcs are analyzed using finite-thrust approach. The motion of the corresponding dynamical system is realized in three phase spaces concerning the departure planet-centric maneuver by means of high-thrust NTR propulsion, the low-thrust NEP heliocentric maneuver and the approach high-thrust NTR planet-centric maneuver. The phase coordinates are related at the time instants of the change of the phase spaces due to the relations between the space vehicle masses. The optimal control analysis is performed using Pontryagin's maximum principle. The numerical results are analyzed for Earth-Mars "sprint" transfer. The optimal values of the parameters that define the masses of NTR and NEP subsystems have been evaluated. It is shown that the low

  16. Double Species Bose-Einstein Condensate with Tunable Interspecies Interactions

    SciTech Connect

    Thalhammer, G.; Barontini, G.; De Sarlo, L.; Catani, J.; Minardi, F.; Inguscio, M.

    2008-05-30

    We produce Bose-Einstein condensates of two different species, {sup 87}Rb and {sup 41}K, in an optical dipole trap in proximity of interspecies Feshbach resonances. We discover and characterize two Feshbach resonances, located around 35 and 79 G, by observing the three-body losses and the elastic cross section. The narrower resonance is exploited to create a double species condensate with tunable interactions. Our system opens the way to the exploration of double species Mott insulators and, more in general, of the quantum phase diagram of the two-species Bose-Hubbard model.

  17. Coupled quantum-classical method for long range charge transfer: relevance of the nuclear motion to the quantum electron dynamics

    NASA Astrophysics Data System (ADS)

    da Silva, Robson; Hoff, Diego A.; Rego, Luis G. C.

    2015-04-01

    Charge and excitonic-energy transfer phenomena are fundamental for energy conversion in solar cells as well as artificial photosynthesis. Currently, much interest is being paid to light-harvesting and energy transduction processes in supramolecular structures, where nuclear dynamics has a major influence on electronic quantum dynamics. For this reason, the simulation of long range electron transfer in supramolecular structures, under environmental conditions described within an atomistic framework, has been a difficult problem to study. This work describes a coupled quantum mechanics/molecular mechanics method that aims at describing long range charge transfer processes in supramolecular systems, taking into account the atomistic details of large molecular structures, the underlying nuclear motion, and environmental effects. The method is applied to investigate the relevance of electron-nuclei interaction on the mechanisms for photo-induced electron-hole pair separation in dye-sensitized interfaces as well as electronic dynamics in molecular structures.

  18. Assessment of nuclear transfer techniques to prevent the transmission of heritable mitochondrial disorders without compromising embryonic development competence in mice.

    PubMed

    Neupane, Jitesh; Vandewoestyne, Mado; Ghimire, Sabitri; Lu, Yuechao; Qian, Chen; Van Coster, Rudy; Gerris, Jan; Deroo, Tom; Deforce, Dieter; De Sutter, Petra; Heindryckx, Björn

    2014-09-01

    To evaluate and compare mitochondrial DNA (mtDNA) carry-over and embryonic development potential between different nuclear transfer techniques we performed germinal vesicle nuclear transfer (GV NT), metaphase-II spindle-chromosome-complex (MII-SCC) transfer and pronuclear transfer (PNT) in mice. No detectable mtDNA carry-over was seen in most of the reconstructed oocytes and embryos. No significant differences were seen in mtDNA carry-over rate between GV NT (n=20), MII-SCC transfer (0.29 ± 0.63; n=21) and PNT (0.29 ± 0.75; n=25). Blastocyst formation was not compromised after either PNT (88%; n=18) or MII-SCC transfer (86%; n=27). Further analysis of blastomeres from cleaving embryos (n=8) demonstrated undetectable mtDNA carry-over in all but one blastomere. We show that NT in the germ line is potent to prevent transmission of heritable mtDNA disorders with the applicability for patients attempting reproduction.

  19. Uncoupled Embryonic and Extra-Embryonic Tissues Compromise Blastocyst Development after Somatic Cell Nuclear Transfer

    PubMed Central

    Degrelle, Séverine A.; Jaffrezic, Florence; Campion, Evelyne; Lê Cao, Kim-Anh; Le Bourhis, Daniel; Richard, Christophe; Rodde, Nathalie; Fleurot, Renaud; Everts, Robin E.; Lecardonnel, Jérôme; Heyman, Yvan; Vignon, Xavier; Tian, Xiuchun C.; Lewin, Harris A.; Renard, Jean-Paul; Hue, Isabelle

    2012-01-01

    Somatic cell nuclear transfer (SCNT) is the most efficient cell reprogramming technique available, especially when working with bovine species. Although SCNT blastocysts performed equally well or better than controls in the weeks following embryo transfer at Day 7, elongation and gastrulation defects were observed prior to implantation. To understand the developmental implications of embryonic/extra-embryonic interactions, the morphological and molecular features of elongating and gastrulating tissues were analysed. At Day 18, 30 SCNT conceptuses were compared to 20 controls (AI and IVP: 10 conceptuses each); one-half of the SCNT conceptuses appeared normal while the other half showed signs of atypical elongation and gastrulation. SCNT was also associated with a high incidence of discordance in embryonic and extra-embryonic patterns, as evidenced by morphological and molecular “uncoupling”. Elongation appeared to be secondarily affected; only 3 of 30 conceptuses had abnormally elongated shapes and there were very few differences in gene expression when they were compared to the controls. However, some of these differences could be linked to defects in microvilli formation or extracellular matrix composition and could thus impact extra-embryonic functions. In contrast to elongation, gastrulation stages included embryonic defects that likely affected the hypoblast, the epiblast, or the early stages of their differentiation. When taking into account SCNT conceptus somatic origin, i.e. the reprogramming efficiency of each bovine ear fibroblast (Low: 0029, Med: 7711, High: 5538), we found that embryonic abnormalities or severe embryonic/extra-embryonic uncoupling were more tightly correlated to embryo loss at implantation than were elongation defects. Alternatively, extra-embryonic differences between SCNT and control conceptuses at Day 18 were related to molecular plasticity (high efficiency/high plasticity) and subsequent pregnancy loss. Finally, because it alters

  20. Coupled thermochemical, isotopic evolution and heat transfer simulations in highly irradiated UO2 nuclear fuel

    NASA Astrophysics Data System (ADS)

    Piro, M. H. A.; Banfield, J.; Clarno, K. T.; Simunovic, S.; Besmann, T. M.; Lewis, B. J.; Thompson, W. T.

    2013-10-01

    Predictive capabilities for simulating irradiated nuclear fuel behavior are enhanced in the current work by coupling thermochemistry, isotopic evolution and heat transfer. Thermodynamic models that are incorporated into this framework not only predict the departure from stoichiometry of UO2, but also consider dissolved fission and activation products in the fluorite oxide phase, noble metal inclusions, secondary oxides including uranates, zirconates, molybdates and the gas phase. Thermochemical computations utilize the spatial and temporal evolution of the fission and activation product inventory in the pellet, which is typically neglected in nuclear fuel performance simulations. Isotopic computations encompass the depletion, decay and transmutation of more than 2000 isotopes that are calculated at every point in space and time. These computations take into consideration neutron flux depression and the increased production of fissile plutonium near the fuel pellet periphery (i.e., the so-called “rim effect”). Thermochemical and isotopic predictions are in very good agreement with reported experimental measurements of highly irradiated UO2 fuel with an average burnup of 102 GW d t(U)-1. Simulation results demonstrate that predictions are considerably enhanced when coupling thermochemical and isotopic computations in comparison to empirical correlations. Notice: This manuscript has been authored by UT-Battelle, LLC, under Contract No. DE-AC05-00OR22725 with the U.S. Department of Energy. The United States Government retains and the publisher, by accepting the article for publication, acknowledges that the United States Government retains a non-exclusive, paid-up, irrevocable, world-wide license to publish or reproduce the published form of this manuscript, or allow others to do so, for United States Government purposes.

  1. Evolutionary Role of Interspecies Hybridization and Genetic Exchanges in Yeasts

    PubMed Central

    Dujon, Bernard

    2012-01-01

    Summary: Forced interspecific hybridization has been used in yeasts for many years to study speciation or to construct artificial strains with novel fermentative and metabolic properties. Recent genome analyses indicate that natural hybrids are also generated spontaneously between yeasts belonging to distinct species, creating lineages with novel phenotypes, varied genetic stability, or altered virulence in the case of pathogens. Large segmental introgressions from evolutionarily distant species are also visible in some yeast genomes, suggesting that interspecific genetic exchanges occur during evolution. The origin of this phenomenon remains unclear, but it is likely based on weak prezygotic barriers, limited Dobzhansky-Muller (DM) incompatibilities, and rapid clonal expansions. Newly formed interspecies hybrids suffer rapid changes in the genetic contribution of each parent, including chromosome loss or aneuploidy, translocations, and loss of heterozygosity, that, except in a few recently studied cases, remain to be characterized more precisely at the genomic level by use of modern technologies. We review here known cases of natural or artificially formed interspecies hybrids between yeasts and discuss their potential importance in terms of genome evolution. Problems of meiotic fertility, ploidy constraint, gene and gene product compatibility, and nucleomitochondrial interactions are discussed and placed in the context of other known mechanisms of yeast genome evolution as a model for eukaryotes. PMID:23204364

  2. Human-mouse interspecies collagen I heterotrimer is functional during embryonic development of Mov13 mutant mouse embryos.

    PubMed Central

    Wu, H; Bateman, J F; Schnieke, A; Sharpe, A; Barker, D; Mascara, T; Eyre, D; Bruns, R; Krimpenfort, P; Berns, A

    1990-01-01

    To investigate whether the human pro alpha 1(I) collagen chain could form an in vivo functional interspecies heterotrimer with the mouse pro alpha 2(I) collagen chain, we introduced the human COL1A1 gene into Mov13 mice which have a functional deletion of the endogenous COL1A1 gene. Transgenic mouse strains (HucI and HucII) carrying the human COL1A1 gene were first generated by microinjecting the COL1A1 gene into wild-type mouse embryos. Genetic evidence indicated that the transgene in the HucI strain was closely linked to the endogenous mouse COL1A1 gene and was X linked in the HucII transgenic strain. Northern (RNA) blot and S1 protection analyses showed that the transgene was expressed in the appropriate tissue-specific manner and as efficiently as the endogenous COL1A1 gene. HucII mice were crossed with Mov13 mice to transfer the human transgene into the mutant strain. Whereas homozygous Mov13 embryos die between days 13 and 14 of gestation, the presence of the transgene permitted apparently normal development of the mutant embryos to birth. This indicated that the mouse-human interspecies collagen I heterotrimer was functional in the animal. The rescue was, however, only partial, as all homozygotes died within 36 h after delivery, with signs of internal bleeding. This could have been due to a functional defect in the interspecies hybrid collagen. Extensive analysis failed to reveal any biochemical or morphological abnormalities of the collagen I molecules in Mov13-HucII embryos.(ABSTRACT TRUNCATED AT 250 WORDS) Images PMID:1690840

  3. TRANSFER OF EXCESS NUCLEAR MATERIAL FROM LOS ALAMOS TO SAVANNAH RIVER SITE FOR LONG-TERM DISPOSITION

    SciTech Connect

    C. W. HOTH; L. A. FOSTER; T. F YARBRO

    2001-06-01

    Los Alamos National Laboratory is preparing excess nuclear material for shipment to Savannah River Site (SRS) for final disposition. Prior to shipment the nuclear material will be stabilized and packaged to meet strict criteria. The criterion that must be met include: (1) the DOE stabilization, packaging and storage requirements for plutonium bearing materials, DOE-STD-3013, (2) shipping container packaging requirements, (3) SRS packaging and storage criteria, and (4) DOE Material Disposition criteria for either immobilization or MOX reactor fuel. Another issue in preparing for this transfer is the DOE certification of shipping containers and the availability of shipping containers. This transfer of the nuclear material is fully supported by the EM, DP and NN Sections of the DOE, as well as, by LANL and SRS, yet a strong collaboration is needed to meet all established requirements relating to stabilization, packaging, shipment, storage and final disposition. This paper will present the overall objectives, the issues and the planned strategy to accomplish this nuclear material transfer.

  4. Transcriptomic Features of Bovine Blastocysts Derived by Somatic Cell Nuclear Transfer.

    PubMed

    Min, Byungkuk; Cho, Sunwha; Park, Jung Sun; Lee, Yun-Gyeong; Kim, Namshin; Kang, Yong-Kook

    2015-09-03

    Reprogramming incompletely occurs in most somatic cell nuclear transfer (SCNT) embryos, which results in misregulation of developmentally important genes and subsequent embryonic malfunction and lethality. Here we examined transcriptome profiles in single bovine blastocysts derived by in vitro fertilization (IVF) and SCNT. Different types of donor cells, cumulus cell and ear-skin fibroblast, were used to derive cSCNT and fSCNT blastocysts, respectively. SCNT blastocysts expressed 13,606 genes on average, similar to IVF (13,542). Correlation analysis found that both cSCNT and fSCNT blastocyst groups had transcriptomic features distinctive from the IVF group, with the cSCNT transcriptomes closer to the IVF ones than the fSCNT. Gene expression analysis identified 56 underrepresented and 78 overrepresented differentially expressed genes in both SCNT groups. A 400-kb locus harboring zinc-finger protein family genes in chromosome 18 were found coordinately down-regulated in fSCNT blastocysts, showing a feature of reprogramming-resistant regions. Probing into different categories of genes important for blastocyst development revealed that genes involved in trophectoderm development frequently were underrepresented, and those encoding epigenetic modifiers tended to be overrepresented in SCNT blastocysts. Our effort to identify reprogramming-resistant, differentially expressed genes can help map reprogramming error-prone loci onto the genome and elucidate how to handle the stochastic events of reprogramming to improve cloning efficiency.

  5. In vitro and in vivo genotoxic effects of somatic cell nuclear transfer cloned cattle meat.

    PubMed

    Lee, Nam-Jin; Yang, Byoung-Chul; Jung, Yu-Ri; Lee, Jung-Won; Im, Gi-Sun; Seong, Hwan-Hoo; Park, Jin-Ki; Kang, Jong-Koo; Hwang, Seongsoo

    2011-09-01

    Although the nutritional composition and health status after consumption of the meat and milk derived from both conventionally bred (normal) and somatic cell nuclear transferred (cloned) animals and their progeny are not different, little is known about their food safeties like genetic toxicity. This study is performed to examine both in vitro (bacterial mutation and chromosome aberration) and in vivo (micronucleus) genotoxicity studies of cloned cattle meat. The concentrations of both normal and cloned cattle meat extracts (0-10×) were tested to five strains of bacteria (Salmonella typhimurium: TA98, TA100, TA1535, and TA1537; Escherichia coli: WP2uvrA) for bacterial mutation and to Chinese hamster lung (CHL/IU) cells for chromosome aberration, respectively. For micronucleus test, ICR mice were divided into five dietary groups: commercial pellets (control), pellets containing 5% (N-5) and 10% (N-10) normal cattle meat, and pellets containing 5% (C-5) and 10% (C-10) cloned cattle meat. No test substance-related genotoxicity was noted in the five bacterial strains, CHL/IU cells, or mouse bone marrow cells, suggesting that the cloned cattle meat potentially may be safe in terms of mutagenic hazards. Thus, it can be postulated that the cloned cattle meat do not induce any harmful genotoxic effects in vitro and in vivo.

  6. Generation of biallelic knock-out sheep via gene-editing and somatic cell nuclear transfer

    PubMed Central

    Li, Honghui; Wang, Gui; Hao, Zhiqiang; Zhang, Guozhong; Qing, Yubo; Liu, Shuanghui; Qing, Lili; Pan, Weirong; Chen, Lei; Liu, Guichun; Zhao, Ruoping; Jia, Baoyu; Zeng, Luyao; Guo, Jianxiong; Zhao, Lixiao; Zhao, Heng; Lv, Chaoxiang; Xu, Kaixiang; Cheng, Wenmin; Li, Hushan; Zhao, Hong-Ye; Wang, Wen; Wei, Hong-Jiang

    2016-01-01

    Transgenic sheep can be used to achieve genetic improvements in breeds and as an important large-animal model for biomedical research. In this study, we generated a TALEN plasmid specific for ovine MSTN and transfected it into fetal fibroblast cells of STH sheep. MSTN biallelic-KO somatic cells were selected as nuclear donor cells for SCNT. In total, cloned embryos were transferred into 37 recipient gilts, 28 (75.7%) becoming pregnant and 15 delivering, resulting in 23 lambs, 12 of which were alive. Mutations in the lambs were verified via sequencing and T7EI assay, and the gene mutation site was consistent with that in the donor cells. Off-target analysis was performed, and no off-target mutations were detected. MSTN KO affected the mRNA expression of MSTN relative genes. The growth curve for the resulting sheep suggested that MSTN KO caused a remarkable increase in body weight compared with those of wild-type sheep. Histological analyses revealed that MSTN KO resulted in muscle fiber hypertrophy. These findings demonstrate the successful generation of MSTN biallelic-KO STH sheep via gene editing in somatic cells using TALEN technology and SCNT. These MSTN mutant sheep developed and grew normally, and exhibited increased body weight and muscle growth. PMID:27654750

  7. Transcriptomic Features of Bovine Blastocysts Derived by Somatic Cell Nuclear Transfer

    PubMed Central

    Min, Byungkuk; Cho, Sunwha; Park, Jung Sun; Lee, Yun-Gyeong; Kim, Namshin; Kang, Yong-Kook

    2015-01-01

    Reprogramming incompletely occurs in most somatic cell nuclear transfer (SCNT) embryos, which results in misregulation of developmentally important genes and subsequent embryonic malfunction and lethality. Here we examined transcriptome profiles in single bovine blastocysts derived by in vitro fertilization (IVF) and SCNT. Different types of donor cells, cumulus cell and ear-skin fibroblast, were used to derive cSCNT and fSCNT blastocysts, respectively. SCNT blastocysts expressed 13,606 genes on average, similar to IVF (13,542). Correlation analysis found that both cSCNT and fSCNT blastocyst groups had transcriptomic features distinctive from the IVF group, with the cSCNT transcriptomes closer to the IVF ones than the fSCNT. Gene expression analysis identified 56 underrepresented and 78 overrepresented differentially expressed genes in both SCNT groups. A 400-kb locus harboring zinc-finger protein family genes in chromosome 18 were found coordinately down-regulated in fSCNT blastocysts, showing a feature of reprogramming-resistant regions. Probing into different categories of genes important for blastocyst development revealed that genes involved in trophectoderm development frequently were underrepresented, and those encoding epigenetic modifiers tended to be overrepresented in SCNT blastocysts. Our effort to identify reprogramming-resistant, differentially expressed genes can help map reprogramming error-prone loci onto the genome and elucidate how to handle the stochastic events of reprogramming to improve cloning efficiency. PMID:26342001

  8. Vitamin C enhances in vitro and in vivo development of porcine somatic cell nuclear transfer embryos.

    PubMed

    Huang, Yongye; Tang, Xiaochun; Xie, Wanhua; Zhou, Yan; Li, Dong; Zhou, Yang; Zhu, Jianguo; Yuan, Ting; Lai, Liangxue; Pang, Daxin; Ouyang, Hongsheng

    2011-07-29

    The reprogramming of differentiated cells into a totipotent embryonic state through somatic cell nuclear transfer (SCNT) is still an inefficient process. Previous studies revealed that the generation of induced pluripotent stem (iPS) cells from mouse and human fibroblasts could be significantly enhanced with vitamin C treatment. Here, we investigated the effects of vitamin C, to our knowledge for the first time, on the in vitro and in vivo development of porcine SCNT embryos. The rate of blastocyst development in SCNT embryos treated with 50μg/mL vitamin C 15h after activation (36.0%) was significantly higher than that of untreated SCNT embryos (11.5%). The enhanced in vitro development rate of vitamin C-treated embryos was associated with an increased acetylation level of histone H4 lysine 5 and higher Oct4, Sox2 and Klf4 expression levels in blastocysts, as determined by real-time PCR. In addition, treatment with vitamin C resulted in an increased pregnancy rate in pigs. These findings suggest that treatment with vitamin C is beneficial for enhancement of the in vitro and in vivo development of porcine SCNT embryos. PMID:21749856

  9. Lymphoid lineage differentiation potential of mouse nuclear transfer embryonic stem cells.

    PubMed

    Eslami-Arshaghi, Tarlan; Salehi, Mohammad; Soleimani, Masoud; Gholipourmalekabadi, Mazaher; Mossahebi-Mohammadi, Majid; Ardeshirylajimi, Abdolreza; Rajabi, Hoda

    2015-09-01

    Stem cells therapy is considered as an efficient strategy for the treatment of some diseases. Nevertheless, some obstacles such as probability of rejection by the immune system limit applications of this strategy. Therefore, several efforts have been made to overcome this among which using the induced pluripotent stem cells (iPSCs) and nuclear transfer embryonic stem cell (nt-ESCs) are the most efficient strategies. The objective of this study was to evaluate the differentiation potential of the nt-ESCs to lymphoid lineage in the presence of IL-7, IL-3, FLT3-ligand and TPO growth factors in vitro. To this end, the nt-ESCs cells were prepared and treated with aforementioned growth factors for 7 and 14 days. Then, the cells were examined for expression of lymphoid markers (CD3, CD25, CD127 and CD19) by quantitative PCR (q-PCR) and flow cytometry. An increased expression of CD19 and CD25 markers was observed in the treated cells compared with the negative control samples by day 7. After 14 days, the expression level of all the tested CD markers significantly increased in the treated groups in comparison with the control. The current study reveals the potential of the nt-ESCs in differentiation to lymphoid lineage in the presence of defined growth factors.

  10. Nuclear retinoid receptors and pregnancy: placental transfer, functions, and pharmacological aspects.

    PubMed

    Comptour, Aurélie; Rouzaire, Marion; Belville, Corinne; Bouvier, Damien; Gallot, Denis; Blanchon, Loïc; Sapin, Vincent

    2016-10-01

    Animal models of vitamin A (retinol) deficiency have highlighted its crucial role in reproduction and placentation, whereas an excess of retinoids (structurally or functionally related entities) can cause toxic and teratogenic effects in the embryo and foetus, especially in the first trimester of human pregnancy. Knock-out experimental strategies-targeting retinoid nuclear receptors RARs and RXRs have confirmed that the effects of vitamin A are mediated by retinoic acid (especially all-trans retinoic acid) and that this vitamin is essential for the developmental process. All these data show that the vitamin A pathway and metabolism are as important for the well-being of the foetus, as they are for that of the adult. Accordingly, during this last decade, extensive research on retinoid metabolism has yielded detailed knowledge on all the actors in this pathway, spurring the development of antagonists and agonists for therapeutic and research applications. Natural and synthetic retinoids are currently used in clinical practice, most often on the skin for the treatment of acne, and as anti-oncogenic agents in acute promyelocytic leukaemia. However, because of the toxicity and teratogenicity of retinoids during pregnancy, their pharmacological use needs a sound knowledge of their metabolism, molecular aspects, placental transfer, and action. PMID:27502420

  11. Nuclear retinoid receptors and pregnancy: placental transfer, functions, and pharmacological aspects.

    PubMed

    Comptour, Aurélie; Rouzaire, Marion; Belville, Corinne; Bouvier, Damien; Gallot, Denis; Blanchon, Loïc; Sapin, Vincent

    2016-10-01

    Animal models of vitamin A (retinol) deficiency have highlighted its crucial role in reproduction and placentation, whereas an excess of retinoids (structurally or functionally related entities) can cause toxic and teratogenic effects in the embryo and foetus, especially in the first trimester of human pregnancy. Knock-out experimental strategies-targeting retinoid nuclear receptors RARs and RXRs have confirmed that the effects of vitamin A are mediated by retinoic acid (especially all-trans retinoic acid) and that this vitamin is essential for the developmental process. All these data show that the vitamin A pathway and metabolism are as important for the well-being of the foetus, as they are for that of the adult. Accordingly, during this last decade, extensive research on retinoid metabolism has yielded detailed knowledge on all the actors in this pathway, spurring the development of antagonists and agonists for therapeutic and research applications. Natural and synthetic retinoids are currently used in clinical practice, most often on the skin for the treatment of acne, and as anti-oncogenic agents in acute promyelocytic leukaemia. However, because of the toxicity and teratogenicity of retinoids during pregnancy, their pharmacological use needs a sound knowledge of their metabolism, molecular aspects, placental transfer, and action.

  12. The cell agglutination agent, phytohemagglutinin-L, improves the efficiency of somatic nuclear transfer cloning in cattle (Bos taurus).

    PubMed

    Du, Fuliang; Shen, Perng-Chih; Xu, Jie; Sung, Li-Ying; Jeong, B-Seon; Lucky Nedambale, Tshimangadzo; Riesen, John; Cindy Tian, X; Cheng, Winston T K; Lee, Shan-Nan; Yang, Xiangzhong

    2006-02-01

    One of the several factors that contribute to the low efficiency of mammalian somatic cloning is poor fusion between the small somatic donor cell and the large recipient oocyte. This study was designed to test phytohemagglutinin (PHA) agglutination activity on fusion rate, and subsequent developmental potential of cloned bovine embryos. The toxicity of PHA was established by examining its effects on the development of parthenogenetic bovine oocytes treated with different doses (Experiment 1), and for different durations (Experiment 2). The effective dose and duration of PHA treatment (150 microg/mL, 20 min incubation) was selected and used to compare membrane fusion efficiency and embryo development following somatic cell nuclear transfer (Experiment 3). Cloning with somatic donor fibroblasts versus cumulus cells was also compared, both with and without PHA treatment (150 microg/mL, 20 min). Fusion rate of nuclear donor fibroblasts, after phytohemagglutinin treatment, was increased from 33 to 61% (P < 0.05), and from 59 to 88% (P < 0.05) with cumulus cell nuclear donors. The nuclear transfer (NT) efficiency per oocyte used was improved following PHA treatment, for both fibroblast (13% versus 22%) as well as cumulus cells (17% versus 34%; P < 0.05). The cloned embryos, both with and without PHA treatment, were subjected to vitrification and embryo transfer testing, and resulted in similar survival (approximately 90% hatching) and pregnancy rates (17-25%). Three calves were born following vitrification and embryo transfer of these embryos; two from the PHA-treated group, and one from non-PHA control group. We concluded that PHA treatment significantly improved the fusion efficiency of somatic NT in cattle, and therefore, increased the development of cloned blastocysts. Furthermore, within a determined range of dose and duration, PHA had no detrimental effect on embryo survival post-vitrification, nor on pregnancy or calving rates following embryo transfer.

  13. Serial cloning of pigs by somatic cell nuclear transfer: restoration of phenotypic normality during serial cloning.

    PubMed

    Cho, Seong-Keun; Kim, Jae-Hwan; Park, Jong-Yi; Choi, Yun-Jung; Bang, Jae-Il; Hwang, Kyu-Chan; Cho, Eun-Jeong; Sohn, Sea-Hwan; Uhm, Sang Jun; Koo, Deog-Bon; Lee, Kyung-Kwang; Kim, Teoan; Kim, Jin-Hoi

    2007-12-01

    Somatic cell nuclear transfer (scNT) is a useful way to create cloned animals. However, scNT clones exhibit high levels of phenotypic instability. This instability may be due to epigenetic reprogramming and/or genomic damage in the donor cells. To test this, we produced transgenic pig fibroblasts harboring the truncated human thrombopoietin (hTPO) gene and used them as donor cells in scNT to produce first-generation (G1) cloned piglets. In this study, 2,818 scNT embryos were transferred to 11 recipients and five G1 piglets were obtained. Among them, a clone had a dimorphic facial appearance with severe hypertelorism and a broad prominent nasal bridge. The other clones looked normal. Second-generation (G2) scNT piglets were then produced using ear cells from a G1 piglet that had an abnormal nose phenotype. We reasoned that, if the phenotypic abnormality of the G1 clone was not present in the G2 and third-generation (G3) clones, or was absent in the G2 clones but reappeared in the G3 clones, the phenotypic instability of the G1 clone could be attributed to faulty epigenetic reprogramming rather than to inherent/accidental genomic damage to the donor cells. Blastocyst rates, cell numbers in blastocyst, pregnancy rates, term placenta weight and ponderal index, and birth weight between G1 and G2 clones did not differ, but were significantly (P < 0.05) lower than control age- and sex-matched piglets. Next, we analyzed global methylation changes during development of the preimplantation embryos reconstructed by donor cells used for the production of G1 and G2 clones and could not find any significant differences in the methylation patterns between G1 and G2 clones. Indeed, we failed to detect the phenotypic abnormality in the G2 and G3 clones. Thus, the phenotypic abnormality of the G1 clone is likely to be due to epigenetic dysregulation. Additional observations then suggested that expression of the hTPO gene in the transgenic clones did not appear to be the cause of the

  14. Distribution and interspecies contact of feral swine and cattle on rangeland in south Texas: implications for disease transmission.

    PubMed

    Cooper, Susan M; Scott, H Morgan; de la Garza, Guadalupe R; Deck, Aubrey L; Cathey, James C

    2010-01-01

    The last outbreak of foot-and-mouth disease (FMD) in the United States occurred in 1929. Since that time, numbers and distribution of feral swine (Sus scrofa) have increased greatly, especially in the southern states. This creates a potential risk to livestock production because swine are susceptible to, and can be carriers of, several economically harmful diseases of livestock. Most importantly, swine are potent amplifiers of FMD virus. In this study, global positioning system (GPS) collars were placed on rangeland cattle (Bos indicus x taurus) and feral swine to determine shared habitat use by these species on a large ranch in south Texas from 2004 to 2006. The aim was to identify locations and rates of interspecies contact that may result in effective transfer of FMD virus, should an outbreak occur. In shrubland and riparian areas, animals were dispersed, so contacts within and between species were relatively infrequent. Indirect contacts, whereby cattle and feral swine used the same location (within 20 m) within a 360-min period, occurred primarily at water sources, and seasonally in irrigated forage fields and along ranch roads. Direct contacts between species (animals <20 m apart and within 15 min) were rare and occurred primarily at water sources. Changes in ranch management practices are suggested to reduce interspecies contact should an FMD disease outbreak occur. This information can also be used to improve current epidemiologic models to better fit free-ranging animal populations.

  15. Nuclear transfer of synchronized African wild cat somatic cells into enucleated domestic cat oocytes

    USGS Publications Warehouse

    Gomez, M.C.; Jenkins, J.A.; Giraldo, A.; Harris, R.F.; King, A.; Dresser, B.L.; Pope, C.E.

    2003-01-01

    The African wild cat is one of the smallest wild cats and its future is threatened by hybridization with domestic cats. Nuclear transfer, a valuable tool for retaining genetic variability, offers the possibility of species continuation rather than extinction. The aim of this study was to investigate the ability of somatic cell nuclei of the African wild cat (AWC) to dedifferentiate within domestic cat (DSH) cytoplasts and to support early development after nuclear transplantation. In experiment 1, distributions of AWC and DSH fibroblasts in each cell-cycle phase were assessed by flow cytometry using cells cultured to confluency and disaggregated with pronase, trypsin, or mechanical separation. Trypsin (89.0%) and pronase (93.0%) yielded higher proportions of AWC nuclei in the G0/G1 phase than mechanical separation (82.0%). In contrast, mechanical separation yielded higher percentages of DSH nuclei in the G0/G1 phase (86.6%) than pronase (79.7%) or trypsin (74.2%) treatments. In both species, pronase induced less DNA damage than trypsin. In experiment 2, the effects of serum starvation, culture to confluency, and exposure to roscovitine on the distribution of AWC and DSH fibroblasts in various phases of the cell cycle were determined. Flow cytometry analyses revealed that the dynamics of the cell cycle varied as culture conditions were modified. Specifically, a higher percentage of AWC and DSH nuclei were in the G0/G1 phase after cells were serum starved (83% vs. 96%) than were present in cycling cells (50% vs. 64%), after contact inhibition (61% vs. 88%), or after roscovitine (56% vs. 84%) treatment, respectively. In experiment 3, we evaluated the effects of cell synchronization and oocyte maturation (in vivo vs. in vitro) on the reconstruction and development of AWC-DSH- and DSH-DSH-cloned embryos. The method of cell synchronization did not affect the fusion and cleavage rate because only a slightly higher percentage of fused couplets cleaved when donor nuclei

  16. Frequent somatic transfer of mitochondrial DNA into the nuclear genome of human cancer cells.

    PubMed

    Ju, Young Seok; Tubio, Jose M C; Mifsud, William; Fu, Beiyuan; Davies, Helen R; Ramakrishna, Manasa; Li, Yilong; Yates, Lucy; Gundem, Gunes; Tarpey, Patrick S; Behjati, Sam; Papaemmanuil, Elli; Martin, Sancha; Fullam, Anthony; Gerstung, Moritz; Nangalia, Jyoti; Green, Anthony R; Caldas, Carlos; Borg, Åke; Tutt, Andrew; Lee, Ming Ta Michael; van't Veer, Laura J; Tan, Benita K T; Aparicio, Samuel; Span, Paul N; Martens, John W M; Knappskog, Stian; Vincent-Salomon, Anne; Børresen-Dale, Anne-Lise; Eyfjörd, Jórunn Erla; Flanagan, Adrienne M; Foster, Christopher; Neal, David E; Cooper, Colin; Eeles, Rosalind; Lakhani, Sunil R; Desmedt, Christine; Thomas, Gilles; Richardson, Andrea L; Purdie, Colin A; Thompson, Alastair M; McDermott, Ultan; Yang, Fengtang; Nik-Zainal, Serena; Campbell, Peter J; Stratton, Michael R

    2015-06-01

    Mitochondrial genomes are separated from the nuclear genome for most of the cell cycle by the nuclear double membrane, intervening cytoplasm, and the mitochondrial double membrane. Despite these physical barriers, we show that somatically acquired mitochondrial-nuclear genome fusion sequences are present in cancer cells. Most occur in conjunction with intranuclear genomic rearrangements, and the features of the fusion fragments indicate that nonhomologous end joining and/or replication-dependent DNA double-strand break repair are the dominant mechanisms involved. Remarkably, mitochondrial-nuclear genome fusions occur at a similar rate per base pair of DNA as interchromosomal nuclear rearrangements, indicating the presence of a high frequency of contact between mitochondrial and nuclear DNA in some somatic cells. Transmission of mitochondrial DNA to the nuclear genome occurs in neoplastically transformed cells, but we do not exclude the possibility that some mitochondrial-nuclear DNA fusions observed in cancer occurred years earlier in normal somatic cells. PMID:25963125

  17. Frequent somatic transfer of mitochondrial DNA into the nuclear genome of human cancer cells

    PubMed Central

    Ju, Young Seok; Tubio, Jose M.C.; Mifsud, William; Fu, Beiyuan; Davies, Helen R.; Ramakrishna, Manasa; Li, Yilong; Yates, Lucy; Gundem, Gunes; Tarpey, Patrick S.; Behjati, Sam; Papaemmanuil, Elli; Martin, Sancha; Fullam, Anthony; Gerstung, Moritz; Nangalia, Jyoti; Green, Anthony R.; Caldas, Carlos; Borg, Åke; Tutt, Andrew; Lee, Ming Ta Michael; van't Veer, Laura J.; Tan, Benita K.T.; Aparicio, Samuel; Span, Paul N.; Martens, John W.M.; Knappskog, Stian; Vincent-Salomon, Anne; Børresen-Dale, Anne-Lise; Eyfjörd, Jórunn Erla; Flanagan, Adrienne M.; Foster, Christopher; Neal, David E.; Cooper, Colin; Eeles, Rosalind; Lakhani, Sunil R.; Desmedt, Christine; Thomas, Gilles; Richardson, Andrea L.; Purdie, Colin A.; Thompson, Alastair M.; McDermott, Ultan; Yang, Fengtang; Nik-Zainal, Serena; Campbell, Peter J.; Stratton, Michael R.

    2015-01-01

    Mitochondrial genomes are separated from the nuclear genome for most of the cell cycle by the nuclear double membrane, intervening cytoplasm, and the mitochondrial double membrane. Despite these physical barriers, we show that somatically acquired mitochondrial-nuclear genome fusion sequences are present in cancer cells. Most occur in conjunction with intranuclear genomic rearrangements, and the features of the fusion fragments indicate that nonhomologous end joining and/or replication-dependent DNA double-strand break repair are the dominant mechanisms involved. Remarkably, mitochondrial-nuclear genome fusions occur at a similar rate per base pair of DNA as interchromosomal nuclear rearrangements, indicating the presence of a high frequency of contact between mitochondrial and nuclear DNA in some somatic cells. Transmission of mitochondrial DNA to the nuclear genome occurs in neoplastically transformed cells, but we do not exclude the possibility that some mitochondrial-nuclear DNA fusions observed in cancer occurred years earlier in normal somatic cells. PMID:25963125

  18. Frequent somatic transfer of mitochondrial DNA into the nuclear genome of human cancer cells.

    PubMed

    Ju, Young Seok; Tubio, Jose M C; Mifsud, William; Fu, Beiyuan; Davies, Helen R; Ramakrishna, Manasa; Li, Yilong; Yates, Lucy; Gundem, Gunes; Tarpey, Patrick S; Behjati, Sam; Papaemmanuil, Elli; Martin, Sancha; Fullam, Anthony; Gerstung, Moritz; Nangalia, Jyoti; Green, Anthony R; Caldas, Carlos; Borg, Åke; Tutt, Andrew; Lee, Ming Ta Michael; van't Veer, Laura J; Tan, Benita K T; Aparicio, Samuel; Span, Paul N; Martens, John W M; Knappskog, Stian; Vincent-Salomon, Anne; Børresen-Dale, Anne-Lise; Eyfjörd, Jórunn Erla; Flanagan, Adrienne M; Foster, Christopher; Neal, David E; Cooper, Colin; Eeles, Rosalind; Lakhani, Sunil R; Desmedt, Christine; Thomas, Gilles; Richardson, Andrea L; Purdie, Colin A; Thompson, Alastair M; McDermott, Ultan; Yang, Fengtang; Nik-Zainal, Serena; Campbell, Peter J; Stratton, Michael R

    2015-06-01

    Mitochondrial genomes are separated from the nuclear genome for most of the cell cycle by the nuclear double membrane, intervening cytoplasm, and the mitochondrial double membrane. Despite these physical barriers, we show that somatically acquired mitochondrial-nuclear genome fusion sequences are present in cancer cells. Most occur in conjunction with intranuclear genomic rearrangements, and the features of the fusion fragments indicate that nonhomologous end joining and/or replication-dependent DNA double-strand break repair are the dominant mechanisms involved. Remarkably, mitochondrial-nuclear genome fusions occur at a similar rate per base pair of DNA as interchromosomal nuclear rearrangements, indicating the presence of a high frequency of contact between mitochondrial and nuclear DNA in some somatic cells. Transmission of mitochondrial DNA to the nuclear genome occurs in neoplastically transformed cells, but we do not exclude the possibility that some mitochondrial-nuclear DNA fusions observed in cancer occurred years earlier in normal somatic cells.

  19. Aggregating embryonic but not somatic nuclear transfer embryos increases cloning efficiency in cattle.

    PubMed

    Misica-Turner, Pavla M; Oback, Fleur C; Eichenlaub, Michael; Wells, David N; Oback, Björn

    2007-02-01

    Our objectives were to compare the cellular and molecular effects of aggregating bovine embryonic vs. somatic cell nuclear transfer (ECNT vs. SCNT) embryos and to determine whether aggregation can improve cattle cloning efficiency. We reconstructed cloned embryos from: 1) morula-derived blastomeres, 2) six adult male ear skin fibroblast lines, 3) one fetal female lung fibroblast line (BFF), and 4) two transgenic clonal strains derived from BFF. Embryos were cultured either singularly (1X) or as aggregates of three (3X). In vitro-fertilized (IVF) 1X and 3X embryos served as controls. After aggregation, the in vitro development of ECNT but not that of SCNT or IVF embryos was strongly compromised. The inner cell mass (ICM), total cell (TC) numbers, and ICM:TC ratios significantly increased for all the aggregates. The relative concentration of the key embryonic transcript POU5F1 (or OCT4) did not correlate with these increases, remaining unchanged in the ECNT and IVF aggregates and decreasing significantly in the SCNT aggregates. Overall, the IVF and 3X ECNT but not the 1X ECNT embryos had significantly higher relative POU5F1 levels than the SCNT embryos. High POU5F1 levels correlated with high in vivo survival, while no such correlation was noted for the ICM:TC ratios. Development to weaning was more than doubled in the ECNT aggregates (10/51 or 20% vs. 7/85 or 8% for 3X vs. 1X, respectively; P < 0.05). In contrast, the SCNT and IVF controls showed no improvement in survival. These data reveal striking biological differences between embryonic and somatic clones in response to aggregation.

  20. Production of fertile offspring from oocytes grown in vitro by nuclear transfer in cattle.

    PubMed

    Hirao, Yuji; Naruse, Kenji; Kaneda, Masahiro; Somfai, Tamas; Iga, Kosuke; Shimizu, Manabu; Akagi, Satoshi; Cao, Feng; Kono, Tomohiro; Nagai, Takashi; Takenouchi, Naoki

    2013-09-01

    Because of recent advancements in reproductive technology, oocytes have attained an increasingly enriched value as a unique cell population in the production of offspring. The growing oocytes in the ovary are an immediate potential source that serve this need; however, complete oocyte growth before use is crucial. Our research objective was to create in vitro-grown (IVG) oocytes that would have the ability to perform specialized activities, including nuclear reprogramming, as an alternative to in vivo-grown oocytes. Bovine oocyte-granulosa cell complexes with a mean oocyte diameter of approximately 100 μm were cultured on Millicell membrane inserts, with culture medium supplemented with 4% polyvinylpyrrolidone (molecular weight, 360,000), 20 ng/ml androstenedione, 2 mM hypoxanthine, and 5 ng/ml bone morphogenetic protein 7. Oocyte viability after the 14-day culture period was 95%, and there was a 71% increase in oocyte volume. Upon induction of oocyte maturation, 61% of the IVG oocytes extruded a polar body. Eighty-four percent of the reconstructed IVG oocytes that used cumulus cells as donor cells underwent cleavage, and half of them became blastocysts. DNA methylation analyses of the satellite I and II regions of the blastocysts revealed a similar highly methylated status in the cloned embryos derived from in vivo-grown and IVG oocytes. Finally, one of the nine embryos reconstructed from the IVG oocytes developed into a living calf following embryo transfer. Fertility of the offspring was confirmed. In conclusion, the potential of a proportion of the IVG oocytes was comparable to that of in vivo-grown oocytes.

  1. Protein profiles of bovine placenta derived from somatic cell nuclear transfer.

    PubMed

    Kim, Hong Rye; Kang, Jae Ku; Yoon, Jong Taek; Seong, Hwan Hoo; Jung, Jin Kwan; Lee, Hong Mie; Sik Park, Chang; Jin, Dong Il

    2005-11-01

    Practical application of animal cloning by somatic cell nuclear transfer (SCNT) has been hampered by an extremely low success rate. To address whether placental dysfunction in SCNT causes fetal loss during pregnancy, we have used a global proteomics approach using 2-DE and MS to analyze the differential protein patterns of three placentae from the afterbirth of cases of postnatal death, derived from SCNT of Korean Native cattle, and three normal placentae obtained from the afterbirth of fetuses derived from artificial insemination. Proteins within a pI range of 4.0-7.0 and 6.0-9.0 were analyzed separately by 2-DE in triplicate. A total of approximately 2000 spots were detected in placental 2-DE gels stained with CBB. In the comparison of normal and SCNT samples, 60 spots were identified as differentially expressed proteins, of which 33 spots were up-regulated proteins in SCNT placentae, while 27 spots were down-regulated proteins. Most of the proteins identified in this analysis appeared to be related with protein repair or protection, cytoskeleton, signal transduction, immune system, metabolism, extracellular matrix and remodeling, transcription regulation, cell structure or differentiation and ion transport. One of up-regulated proteins in SCNT was TIMP-2 protein known to be related to extracellular matrix and remodeling during pregnancy. Western blot analysis showed an increased level of TIMP-2 in SCNT placenta compared to normal. Our results revealed composite profiles of key proteins involved in abnormal placenta derived from SCNT, and suggested expression abnormality of these genes in SCNT placenta, resulting in fetal losses following SCNT.

  2. Best Practices for Finite Element Analysis of Spent Nuclear Fuel Transfer, Storage, and Transportation Systems

    SciTech Connect

    Bajwa, Christopher S.; Piotter, Jason; Cuta, Judith M.; Adkins, Harold E.; Klymyshyn, Nicholas A.; Fort, James A.; Suffield, Sarah R.

    2010-08-11

    Storage casks and transportation packages for spent nuclear fuel (SNF) are designed to confine SNF in sealed canisters or casks, provide structural integrity during accidents, and remove decay through a storage or transportation overpack. The transfer, storage, and transportation of SNF in dry storage casks and transport packages is regulated under 10 CFR Part 72 and 10 CFR Part 71, respectively. Finite Element Analysis (FEA) is used with increasing frequency in Safety Analysis Reports and other regulatory technical evaluations related to SNF casks and packages and their associated systems. Advances in computing power have made increasingly sophisticated FEA models more feasible, and as a result, the need for careful review of such models has also increased. This paper identifies best practice recommendations that stem from recent NRC review experience. The scope covers issues common to all commercially available FEA software, and the recommendations are applicable to any FEA software package. Three specific topics are addressed: general FEA practices, issues specific to thermal analyses, and issues specific to structural analyses. General FEA practices covers appropriate documentation of the model and results, which is important for an efficient review process. The thermal analysis best practices are related to cask analysis for steady state conditions and transient scenarios. The structural analysis best practices are related to the analysis of casks and associated payload during standard handling and drop scenarios. The best practices described in this paper are intended to identify FEA modeling issues and provide insights that can help minimize associated uncertainties and errors, in order to facilitate the NRC licensing review process.

  3. Telomere elongation facilitated by trichostatin a in cloned embryos and pigs by somatic cell nuclear transfer.

    PubMed

    Kong, Qingran; Ji, Guangzhen; Xie, Bingteng; Li, Jingyu; Mao, Jian; Wang, Juan; Liu, Shichao; Liu, Lin; Liu, Zhonghua

    2014-06-01

    Telomere attrition and genomic instability are associated with organism aging. Concerns still exist regarding telomere length resetting in cloned embryos and ntES cells, and possibilities of premature aging of cloned animals achieved by somatic cell nuclear transfer (SCNT). Trichostatin A (TSA), a histone deacetylase inhibitor, effectively improves the developmental competence of cloned embryos and animals, and recently contributes to successful generation of human ntES cells by SCNT. To test the function of TSA on resetting telomere length, we analyzed telomeres in cloned blastocysts and pigs following treatment of SCNT embryos with TSA. Here, we show that telomeres of cloned pigs generated by standard SCNT methods are not effectively restored, compared with those of donor cells, however TSA significantly increases telomere lengths in cloned pigs. Telomeres elongate in cloned porcine embryos during early cleavage from one-cell to four-cell stages. Notably, TSA facilitates telomere lengthening of cloned embryos mainly at morula-blastocyst stages. Knockdown of pTert by shRNA in donor cells reduces telomerase activity in cloned blastocysts but does not abrogate telomere elongation in the TSA-treated embryos (p > 0.05). However, genes associated with recombination or telomerase-independent mechanism of alternative lengthening of telomeres (ALT) Rad50 and BLM show increased expression in TSA-treated embryos. These data suggest that TSA may promote telomere elongation of cloned porcine embryos by ALT. Together, TSA can elongate telomeres in cloned embryos and piglets, and this could be one of the mechanisms underlying improved development of cloned embryos and animals treated with TSA. PMID:24510582

  4. Ovine induced pluripotent stem cells are resistant to reprogramming after nuclear transfer.

    PubMed

    German, Sergio D; Campbell, Keith H S; Thornton, Elisabeth; McLachlan, Gerry; Sweetman, Dylan; Alberio, Ramiro

    2015-02-01

    Induced pluripotent stem cells (iPSCs) share similar characteristics of indefinite in vitro growth with embryonic stem cells (ESCs) and may therefore serve as a useful tool for the targeted genetic modification of farm animals via nuclear transfer (NT). Derivation of stable ESC lines from farm animals has not been possible, therefore, it is important to determine whether iPSCs can be used as substitutes for ESCs in generating genetically modified cloned farm animals. We generated ovine iPSCs by conventional retroviral transduction using the four Yamanaka factors. These cells were basic fibroblast growth factor (bFGF)- and activin A-dependent, showed persistent expression of the transgenes, acquired chromosomal abnormalities, and failed to activate endogenous NANOG. Nonetheless, iPSCs could differentiate into the three somatic germ layers in vitro. Because cloning of farm animals is best achieved with diploid cells (G1/G0), we synchronized the iPSCs in G1 prior to NT. Despite the cell cycle synchronization, preimplantation development of iPSC-NT embryos was lower than with somatic cells (2% vs. 10% blastocysts, p<0.01). Furthermore, analysis of the blastocysts produced demonstrated persistent expression of the transgenes, aberrant expression of endogenous SOX2, and a failure to activate NANOG consistently. In contrast, gene expression in blastocysts produced with the parental fetal fibroblasts was similar to those generated by in vitro fertilization. Taken together, our data suggest that the persistent expression of the exogenous factors and the acquisition of chromosomal abnormalities are incompatible with normal development of NT embryos produced with iPSCs. PMID:25513856

  5. Somatic cell nuclear transfer alters peri-implantation trophoblast differentiation in bovine embryos.

    PubMed

    Arnold, Daniel R; Bordignon, Vilceu; Lefebvre, Réjean; Murphy, Bruce D; Smith, Lawrence C

    2006-08-01

    Abnormal placental development limits success in ruminant pregnancies derived from somatic cell nuclear transfer (SCNT), due to reduction in placentome number and consequently, maternal/fetal exchange. In the primary stages of an epithelial-chorial association, the maternal/fetal interface is characterized by progressive endometrial invasion by specialized trophoblast binucleate/giant cells (TGC). We hypothesized that dysfunctional placentation in SCNT pregnancies results from aberration in expression of genes known to be necessary for trophoblast proliferation (Mash2), differentiation (Hand1), and function (IFN-tau and PAG-9). We, therefore, compared the expression of these factors in trophoblast from bovine embryos derived from artificial insemination (AI), in vitro fertilization (IVF), and SCNT prior to (day 17) and following (day 40 of gestation) implantation, as well as TGC densities and function. In preimplantation embryos, Mash2 mRNA was more abundant in SCNT embryos compared to AI, while Hand1 was highest in AI and IVF relative to SCNT embryos. IFN-tau mRNA abundance did not differ among groups. PAG-9 mRNA was undetectable in SCNT embryos, present in IVF embryos and highest in AI embryos. In postimplantation pregnancies, SCNT fetal cotyledons displayed higher Mash2 and Hand1 than AI and IVF tissues. Allelic expression of Mash2 was not different among the groups, which suggests that elevated mRNA expression was not due to altered imprinting status of Mash2. The day 40 SCNT cotyledons had the fewest number of TGC compared to IVF and AI controls. Thus, expression of genes critical to normal placental development is altered in SCNT bovine embryos, and this is expected to cause abnormal trophoblast differentiation and contribute to pregnancy loss.

  6. Vitamin C enhances in vitro and in vivo development of porcine somatic cell nuclear transfer embryos

    SciTech Connect

    Huang, Yongye; Tang, Xiaochun; Xie, Wanhua; Zhou, Yan; Li, Dong; Zhou, Yang; Zhu, Jianguo; Yuan, Ting; Lai, Liangxue; Pang, Daxin; Ouyang, Hongsheng

    2011-07-29

    Highlights: {yields} Report for the first time that vitamin C has a beneficial effect on the development of porcine SCNT embryos. {yields} The level of acH4K5 and Oct4 expression at blastocyst-stage was up-regulated after treatment. {yields} A higher rate of gestation and increased number of piglets born were harvested in the treated group. -- Abstract: The reprogramming of differentiated cells into a totipotent embryonic state through somatic cell nuclear transfer (SCNT) is still an inefficient process. Previous studies revealed that the generation of induced pluripotent stem (iPS) cells from mouse and human fibroblasts could be significantly enhanced with vitamin C treatment. Here, we investigated the effects of vitamin C, to our knowledge for the first time, on the in vitro and in vivo development of porcine SCNT embryos. The rate of blastocyst development in SCNT embryos treated with 50 {mu}g/mL vitamin C 15 h after activation (36.0%) was significantly higher than that of untreated SCNT embryos (11.5%). The enhanced in vitro development rate of vitamin C-treated embryos was associated with an increased acetylation level of histone H4 lysine 5 and higher Oct4, Sox2 and Klf4 expression levels in blastocysts, as determined by real-time PCR. In addition, treatment with vitamin C resulted in an increased pregnancy rate in pigs. These findings suggest that treatment with vitamin C is beneficial for enhancement of the in vitro and in vivo development of porcine SCNT embryos.

  7. Trichostatin A Rescues the Disrupted Imprinting Induced by Somatic Cell Nuclear Transfer in Pigs

    PubMed Central

    Huan, Yanjun; Zhu, Jiang; Huang, Bo; Mu, Yanshuang; Kong, Qingran; Liu, Zhonghua

    2015-01-01

    Imprinting disorders induced by somatic cell nuclear transfer (SCNT) usually lead to the abnormalities of cloned animals and low cloning efficiency. Histone deacetylase inhibitors have been shown to improve gene expression, genomic methylation reprogramming and the development of cloned embryos, however, the imprinting statuses in these treated embryos and during their subsequent development remain poorly studied. In this study, we investigated the dynamics of H19/Igf2 methylation and transcription in porcine cloned embryos treated with trichostatin A (TSA), and examined H19/Igf2 imprinting patterns in cloned fetuses and piglets. Our results showed that compared with the maintenance of H19/Igf2 methylation in fertilized embryos, cloned embryos displayed aberrant H19/Igf2 methylation and lower H19/Igf2 transcripts. When TSA enhanced the development of cloned embryos, the disrupted H19/Igf2 imprinting was largely rescued in these treated embryos, more similar to those detected in fertilized counterparts. Further studies displayed that TSA effectively rescued the disrupted imprinting of H19/Igf2 in cloned fetuses and piglets, prevented the occurrence of cloned fetus and piglet abnormalities, and enhanced the full-term development of cloned embryos. In conclusion, our results demonstrated that aberrant imprinting induced by SCNT led to the abnormalities of cloned fetuses and piglets and low cloning efficiency, and TSA rescued the disrupted imprinting in cloned embryos, fetuses and piglets, and prevented the occurrence of cloned fetus and piglet abnormalities, thereby improving the development of cloned embryos. This study would have important implications in improving cloning efficiency and the health of cloned animals. PMID:25962071

  8. An alternative pluripotent state confers interspecies chimaeric competency.

    PubMed

    Wu, Jun; Okamura, Daiji; Li, Mo; Suzuki, Keiichiro; Luo, Chongyuan; Ma, Li; He, Yupeng; Li, Zhongwei; Benner, Chris; Tamura, Isao; Krause, Marie N; Nery, Joseph R; Du, Tingting; Zhang, Zhuzhu; Hishida, Tomoaki; Takahashi, Yuta; Aizawa, Emi; Kim, Na Young; Lajara, Jeronimo; Guillen, Pedro; Campistol, Josep M; Esteban, Concepcion Rodriguez; Ross, Pablo J; Saghatelian, Alan; Ren, Bing; Ecker, Joseph R; Izpisua Belmonte, Juan Carlos

    2015-05-21

    Pluripotency, the ability to generate any cell type of the body, is an evanescent attribute of embryonic cells. Transitory pluripotent cells can be captured at different time points during embryogenesis and maintained as embryonic stem cells or epiblast stem cells in culture. Since ontogenesis is a dynamic process in both space and time, it seems counterintuitive that these two temporal states represent the full spectrum of organismal pluripotency. Here we show that by modulating culture parameters, a stem-cell type with unique spatial characteristics and distinct molecular and functional features, designated as region-selective pluripotent stem cells (rsPSCs), can be efficiently obtained from mouse embryos and primate pluripotent stem cells, including humans. The ease of culturing and editing the genome of human rsPSCs offers advantages for regenerative medicine applications. The unique ability of human rsPSCs to generate post-implantation interspecies chimaeric embryos may facilitate our understanding of early human development and evolution.

  9. Observation of interspecies 6Li-133Cs Feshbach resonances

    NASA Astrophysics Data System (ADS)

    Repp, M.; Pires, R.; Ulmanis, J.; Heck, R.; Kuhnle, E. D.; Weidemüller, M.; Tiemann, E.

    2013-01-01

    We report on the observation of 19 interspecies Feshbach resonances in an optically trapped ultracold Bose-Fermi mixture of 133Cs and 6Li in the two energetically lowest spin states. We assign the resonances to s- and p-wave molecular channels by a coupled-channels calculation, resulting in an accurate determination of LiCs ground-state potentials. Fits of the resonance position based on the undressed asymptotic bound state model do not provide the same level of accuracy as the coupled-channels calculation. Several broad s-wave resonances provide prospects to create fermionic LiCs molecules with a large dipole moment via Feshbach association followed by stimulated Raman passage. Two of the s-wave resonances overlap with a zero crossing of the Cs scattering length, which offers prospects for the investigation of polarons in an ultracold Li-Cs mixture.

  10. Cytogenetic analysis of experimental interspecies goat-sheep chimera.

    PubMed

    Jaszczak, K; Członkowska, M; Guszkiewicz, A; Parada, R

    1991-01-01

    Chromosomal analysis was carried out on blood lymphocytes, skin fibroblasts, and germinal cells of an interspecies goat-sheep chimera. This chimera was produced by aggregation of blastomeres of goat and sheep embryos. A cell chimerism 54,XX/60,XY was found in blood lymphocytes and skin fibroblasts. At birth the percentage of lymphocytes with karyotype 54,XX (sheep) amounted to 80% and with karyotype 60,XY (goat) to 20%. With age the percentage of lymphocytes with chromosome complement 54,XX increased, so that at 18 months it was 94% sheep and 6% goat. At the same age, in skin fibroblasts the percentage of cells with goat karyotype reached 25%. Analysis of germinal cells showed in spermatogonia the presence of only karyotype 60,XY and in primary spermatocytes of 29 autosomal bivalents and the sex bivalent XY. PMID:2061596

  11. [Isolation and nuclear transfer of ES-like cells colonies derived from embryos being cloning of bovine somatic].

    PubMed

    Dong, Ya-Juan; Bai, Xue-Jin; Li, Jian-Dong; Suzuki, Tatsuyuki

    2003-02-01

    In this experiment, it was designed to carry out proliferous culture of bovine blastocysts(day 7) derived from embryos cloned through bovine somatic cell nuclear transfer, isolating and passaging of ES cells. The cells of blastocysts, which were planted on feeder layer, formed small colonies within 24 h. The nest-shape colonies occurred after culturing for 2-3 days. After the colonies in the same shape were isolated and passaged 4-5 times, many different size colonies with monolayer of multi-cells appeared. The colonies that had been passaged 4-5 times were planted into 4-wells multi-dishes without feeder layer. The colonies with monolayer of multi-cells appeared after 24 h, spread all over the bottom of the dishes, emerged epidermis-like cells that appeared reticulate after 4-7 days. These cells were used as donor cells to carry out nuclear transfer. The results showed that 80% (40/50) of the reconstructed embryos cleaved, 5% (2/40) and 2.5% (1/40) of them developed to the morulaes and blastocyst stage, respectively. It revealed that ES-like cells derived embryos constructed through somatic cell nuclear transfer have the developmental potentials.

  12. A comparison of reproductive characteristics of boars generated by somatic cell nuclear transfer to highly related conventionally produced boars.

    PubMed

    Williams, N E; Walker, S C; Reeves, D E; Sherrer, E; Galvin, J M; Polejaeva, I; Rampacek, G; Benyshek, L; Christenson, R K; Graves, W M; Pratt, S L

    2006-01-01

    This study compares the reproductive performance of boars produced by somatic cell nuclear transfer versus conventional breeding. Two different genotypes were selected for comparison: terminal cross line 1 (TX1) and terminal cross line 2 (TX2). The boars selected for comparison from TX1 were three cloned boars, produced by somatic cell nuclear transfer and the conventionally produced progenitor of the clones. The boars selected for comparison from TX2 were a cloned boar produced by somatic cell nuclear transfer and two conventionally produced half sibling boars that were offspring of the progenitor of the clone. Semen from each boar was collected, extended, evaluated and shipped offsite. Upon arrival, the semen was reevaluated and utilized for artificial insemination of 89 commercial gilts, at least 12 gilts per boar, producing 625 piglets. Pregnancy rates were determined at day 30 and 110 of gestation; and farrowing rate and gestation length were recorded. Differences were observed in some of the semen characteristics analyzed with the clones usually possessing superior semen quality to the control, this likely being a result of age differences amongst the clones and controls. Additionally no differences were noted between the clones and controls (progenitor) or between individual boars within genetic line for pregnancy rates, gestation length or any of the litter parameters examined between the clones and controls. These data further support previous reports with limited numbers that the reproductive capabilities of cloned boars are equal to that of conventionally produced boars.

  13. Effect of epigenetic modification with trichostatin A and S-adenosylhomocysteine on developmental competence and POU5F1-EGFP expression of interspecies cloned embryos in dog.

    PubMed

    Mousai, M; Hosseini, S M; Hajian, M; Jafarpour, F; Asgari, V; Forouzanfar, M; Nasr-Esfahani, M H

    2015-10-01

    Adult canine fibroblasts stably transfected with either cytomegalovirus (CMV) or POU5F1 promoter-driven enhanced green fluorescent protein (EGFP) were used to investigate if pre-treatment of these donor cells with two epigenetic drugs [trichostatin A (TSA), or S-adenosylhomocysteine (SAH)] can improve the efficiency of interspecies somatic cell nuclear transfer (iSCNT). Fluorescence-activated cell sorting (FACS), analyses revealed that TSA, but not SAH, treatment of both transgenic and non-transgenic fibroblasts significantly increased acetylation levels compared with untreated relatives. The expression levels of Bcl2 and P53 were significantly affected in TSA-treated cells compared with untreated cells, whereas SAH treatment had no significant effect on cell apoptosis. Irrespective of epigenetic modification, dog/bovine iSCNT embryos had overall similar rates of cleavage and development to 8-16-cell and morula stages in non-transgenic groups. For transgenic reconstructed embryos, however, TSA and SAH could significantly improve development to 8-16-cell and morula stages compared with control. Even though, irrespective of cell transgenesis and epigenetic modification, none of the iSCNT embryos developed to the blastocyst stage. The iSCNT embryos carrying CMV-EGFP expressed EGFP at all developmental stages (2-cell, 4-cell, 8-16-cell, and morula) without mosaicism, while no POU5F1-EGFP signal was observed in any stage of developing iSCNT embryos irrespective of TSA/SAH epigenetic modifications. These results indicated that bovine oocytes partially remodel canine fibroblasts and that TSA and SAH have marginal beneficial effects on this process.

  14. Effect of epigenetic modification with trichostatin A and S-adenosylhomocysteine on developmental competence and POU5F1-EGFP expression of interspecies cloned embryos in dog.

    PubMed

    Mousai, M; Hosseini, S M; Hajian, M; Jafarpour, F; Asgari, V; Forouzanfar, M; Nasr-Esfahani, M H

    2015-10-01

    Adult canine fibroblasts stably transfected with either cytomegalovirus (CMV) or POU5F1 promoter-driven enhanced green fluorescent protein (EGFP) were used to investigate if pre-treatment of these donor cells with two epigenetic drugs [trichostatin A (TSA), or S-adenosylhomocysteine (SAH)] can improve the efficiency of interspecies somatic cell nuclear transfer (iSCNT). Fluorescence-activated cell sorting (FACS), analyses revealed that TSA, but not SAH, treatment of both transgenic and non-transgenic fibroblasts significantly increased acetylation levels compared with untreated relatives. The expression levels of Bcl2 and P53 were significantly affected in TSA-treated cells compared with untreated cells, whereas SAH treatment had no significant effect on cell apoptosis. Irrespective of epigenetic modification, dog/bovine iSCNT embryos had overall similar rates of cleavage and development to 8-16-cell and morula stages in non-transgenic groups. For transgenic reconstructed embryos, however, TSA and SAH could significantly improve development to 8-16-cell and morula stages compared with control. Even though, irrespective of cell transgenesis and epigenetic modification, none of the iSCNT embryos developed to the blastocyst stage. The iSCNT embryos carrying CMV-EGFP expressed EGFP at all developmental stages (2-cell, 4-cell, 8-16-cell, and morula) without mosaicism, while no POU5F1-EGFP signal was observed in any stage of developing iSCNT embryos irrespective of TSA/SAH epigenetic modifications. These results indicated that bovine oocytes partially remodel canine fibroblasts and that TSA and SAH have marginal beneficial effects on this process. PMID:25314965

  15. Prolonged interval between fusion and activation impairs embryonic development by inducing chromosome scattering and nuclear aneuploidy in pig somatic cell nuclear transfer.

    PubMed

    You, Jinyoung; Song, Kilyoung; Lee, Eunsong

    2010-01-01

    The aim of the present study was to examine the effect of various intervals between electrofusion and activation (FA interval) on the nuclear remodelling and development of somatic cell nuclear transfer (SCNT) embryos in pigs. Reconstructed oocytes were activated at 0 (simultaneous fusion and activation; SFA), 1, 2 and 3 h (delayed activation) after electrofusion; these groups were designated as DA1, DA2 and DA3, respectively. When oocyte nuclear status was examined at 0.5, 1, 2 and 3 h after electrofusion, the incidence of chromosome scattering was increased (P < 0.01) as the FA interval was extended (0.0%, 12.0%, 77.3% and 78.0%, respectively). Extending the FA interval led to an increase (P < 0.01) in the percentage of oocytes containing multiple (>or=3) pseudopronuclei (PPN) (0.0% of SFA; 5.3% of DA1; 21.7% of DA2; and 33.5% of DA3). The development of SCNT embryos to the blastocyst stage was decreased (P < 0.05) in DA2 (5.7%) and DA3 (5.0%) compared with SFA (18.1%) and DA1 (19.5%). Our results demonstrate that extending the FA interval impairs the development of SCNT pig embryos by inducing chromosome scattering and the formation of multiple PPN, which may result in increased nuclear aneuploidy.

  16. Effective Oocyte Vitrification and Survival Techniques for Bovine Somatic Cell Nuclear Transfer.

    PubMed

    Park, Min Jee; Lee, Seung Eun; Kim, Eun Young; Lee, Jun Beom; Jeong, Chang Jin; Park, Se Pill

    2015-06-01

    Bovine somatic cell nuclear transfer (SCNT) using vitrified-thawed (VT) oocytes has been studied; however, the cloning efficiency of these oocytes is not comparable with that of nonvitrified (non-V) fresh oocytes. This study sought to optimize the survival and cryopreservation of VT oocytes for SCNT. Co-culture with feeder cells that had been preincubated for 15 h significantly improved the survival of VT oocytes and their in vitro developmental potential following SCNT in comparison to co-culture with feeder cells that had been preincubated for 2, 5, or 24 h (p<0.05). Spindle assessment via the Oosight Microscopy Imaging System and microtubule staining revealed that vitrified metaphase II oocytes (VT group) were not suitable for SCNT. However, enucleating and/or activating oocytes prior to freezing enhanced their developmental potential and suitability for SCNT. The cloning efficiency of the enucleated-activated-vitrified-thawed (EAVT) group (21.6%) was better than that of the other vitrification groups [enucleated-vitrified-thawed (EVT) group, 13.7%; VT group, 15.0%; p<0.05] and was comparable with that of the non-V group (25.9%). The reactive oxygen species level was significantly lower in the EAVT group than in the other vitrification groups (p<0.05). mRNA levels of maternal genes (ZAR1, BMP15, and NLRP5) and a stress gene (HSF1) were lower in the vitrification groups than in the non-V group (p<0.05), whereas the level of phospho-p44/42 mitogen-activated protein kinase did not differ among the groups. Among the vitrification groups, blastocysts in the EAVT group had the best developmental potential, as judged by their high mRNA expression of developmental potential-related genes (POU5f1, Interferon-tau, and SLC2A5) and their low expression of proapoptotic (CASP3) and stress (Hsp70) genes. This study demonstrates that SCNT using bovine frozen-thawed oocytes can be successfully achieved using optimized vitrification and co-culture techniques.

  17. Effective Oocyte Vitrification and Survival Techniques for Bovine Somatic Cell Nuclear Transfer.

    PubMed

    Park, Min Jee; Lee, Seung Eun; Kim, Eun Young; Lee, Jun Beom; Jeong, Chang Jin; Park, Se Pill

    2015-06-01

    Bovine somatic cell nuclear transfer (SCNT) using vitrified-thawed (VT) oocytes has been studied; however, the cloning efficiency of these oocytes is not comparable with that of nonvitrified (non-V) fresh oocytes. This study sought to optimize the survival and cryopreservation of VT oocytes for SCNT. Co-culture with feeder cells that had been preincubated for 15 h significantly improved the survival of VT oocytes and their in vitro developmental potential following SCNT in comparison to co-culture with feeder cells that had been preincubated for 2, 5, or 24 h (p<0.05). Spindle assessment via the Oosight Microscopy Imaging System and microtubule staining revealed that vitrified metaphase II oocytes (VT group) were not suitable for SCNT. However, enucleating and/or activating oocytes prior to freezing enhanced their developmental potential and suitability for SCNT. The cloning efficiency of the enucleated-activated-vitrified-thawed (EAVT) group (21.6%) was better than that of the other vitrification groups [enucleated-vitrified-thawed (EVT) group, 13.7%; VT group, 15.0%; p<0.05] and was comparable with that of the non-V group (25.9%). The reactive oxygen species level was significantly lower in the EAVT group than in the other vitrification groups (p<0.05). mRNA levels of maternal genes (ZAR1, BMP15, and NLRP5) and a stress gene (HSF1) were lower in the vitrification groups than in the non-V group (p<0.05), whereas the level of phospho-p44/42 mitogen-activated protein kinase did not differ among the groups. Among the vitrification groups, blastocysts in the EAVT group had the best developmental potential, as judged by their high mRNA expression of developmental potential-related genes (POU5f1, Interferon-tau, and SLC2A5) and their low expression of proapoptotic (CASP3) and stress (Hsp70) genes. This study demonstrates that SCNT using bovine frozen-thawed oocytes can be successfully achieved using optimized vitrification and co-culture techniques. PMID:25984830

  18. Rex Rabbit Somatic Cell Nuclear Transfer with In Vitro-Matured Oocytes.

    PubMed

    Liu, Yong; Wang, Huili; Lu, Jinhua; Miao, Yiliang; Cao, Xinyan; Zhang, Ling; Wu, Xiaoqing; Wu, Fengrui; Ding, Biao; Wang, Rong; Luo, Mingjiu; Li, Wenyong; Tan, Jinghe

    2016-06-01

    Somatic cell nuclear transfer (SCNT) requires large numbers of matured oocytes. In vitro-matured (IVM) oocytes have been used in SCNT in many animals. We investigated the use of IVM oocytes in Rex rabbit SCNT using Rex rabbit ovaries obtained from a local abattoir. The meiotic ability of oocytes isolated from follicles of different diameters was studied. Rex rabbit SCNT was optimized for denucleation, activation, and donor cell synchronization. Rex rabbit oocytes grew to the largest diameter (110 μm) when the follicle diameter was 1.0 mm. Oocytes isolated from <0.5-mm follicles lacked the ability to resume meiosis. More than 90% of these oocytes remained in the germinal vesicle (GV) stage after in vitro culture (IVC) for 18 h. Oocytes isolated from >0.7-mm follicles acquired maturation ability. More than 90% of these oocytes matured after IVC for 18 h. The developmental potential of oocytes isolated from >1-mm follicles was greater than that of oocytes isolated from 0.7- to 1.0-mm follicles. The highest activation rates for IVM Rex rabbit oocytes were seen after treatment with 2.5 μM ionomycin for 5 min followed by 2 mM 6-dimethylaminopurine (6-DMAP) and 5 μg/mL cycloheximide (CHX) for 1 h. Ionomycin induced the chromatin of IVM oocytes to protrude from the oocyte surface, promoting denucleation. Fetal fibroblast cells (FFCs) and cumulus cells (CCs) were more suitable for Rex rabbit SCNT than skin fibroblast cells (SFCs) (blastocyst rate was 35.6 ± 2.2% and 38.0 ± 6.0% vs. 19.7 ± 3.1%). The best fusion condition was a 2DC interval for 1 sec, 1.6 kV/cm voltages, and 40 μsec duration in 0.28 M mannitol. In conclusion, the in vitro maturation of Rex rabbit oocytes and SCNT procedures were studied systematically and optimized in this study. PMID:27159389

  19. INTER-SPECIES MODELS FOR ACUTE AQUATIC TOXICITY BASED ON MECHANISM OF ACTION

    EPA Science Inventory

    This presentation will provide interspecies QSARs for acute toxicity to 17 aquatic species, such as fish, snail, tadpole, hydrozoan, crustacean, insect larvae, and bacteria developed using 5,000 toxic effect results for approximately 2400 chemicals.

  20. ESTIMATION OF CHEMICAL TOXICITY TO WILDLIFE SPECIES USING INTERSPECIES CORRELATION MODELS

    EPA Science Inventory

    Ecological risks to wildlife are typically assessed using toxicity data for relataively few species and with limited understanding of differences in species sensitivity to contaminants. Empirical interspecies correlation models were derived from LD50 values for 49 wildlife speci...

  1. Development of Species Sensitivity Distributions for Wildlife Using Interspecies Toxicity Correlation Models

    EPA Science Inventory

    Species sensitivity distributions (SSD) are cumulative distributions of chemical toxicity of multiple species and have had limited application in wildlife risk assessment because of relatively small datasets of wildlife toxicity values. Interspecies correlation estimation (ICE) m...

  2. A beam-powered electric orbital transfer vehicle: The advantages of nuclear electric propulsion without the concerns of nuclear power

    SciTech Connect

    Coomes, E.P.; Dagle, J.E.

    1989-06-01

    The advantages of using electric propulsion systems are well-known in the aerospace community with the most common being its high specific impulse, lower propellant requirements, and lower system mass when compared to conventional chemical propulsion systems. But these advantages may not be enough to overcome the disadvantage of the added mass of the nuclear electric power source. In the past, the lack of suitable electric power systems has been a major drawback to electric propulsion. Current programs will have nuclear electric power systems available by the turn of the century. Coupling this with the resurgence of interest in free space electromagnetic transmission of energy (power beaming) and the advances in energy beam technology, a whole new approach to the use of electric propulsion for an orbital transport vehicle (OTV) is possible. Power beaming allows the on-board nuclear electric power source, the most massive component of the OTV, to be removed from the spacecraft and replaced with a lightweight energy receptor. An OTV based on power beaming could have a pay load fraction in excess of 80% and provide delivery times to geosynchronous earth orbit (GEO) as low as 120 days and require only 200 kWe. 21 refs., 10 figs., 2 tabs.

  3. Production of hGFAP-DsRed transgenic Guangxi Bama mini-pigs via somatic cell nuclear transfer.

    PubMed

    Hu, L L; Lu, Y Q; Xu, H Y; Yang, X G; Lu, S S; Lu, K H

    2015-01-01

    The mini-pig is a useful animal model for human biomedical research due to its physiological similarity to humans and the ease of handling. In order to optimize the efficiency of production of transgenic Bama mini-pigs through somatic cell nuclear transfer (SCNT), we examined the effects of contact inhibition, roscovitine treatment, and serum starvation on the cell cycle synchronization and transgenic cloned embryo development in vivo and in vitro after nuclear transfer. The analysis showed that the rates of G0/G1 stage cells in the contact inhibition (92.11%) and roscovitine treatment groups (89.59%) were significantly higher than in serum starvation group (80.82%). A higher rate of apoptosis was seen in the serum starvation group (14.13%) compared to the contact inhibition and roscovitine treatment groups (6.71 and 2.46% respectively, P < 0.05). There was a significant decrease in blastocyst yield in the serum starvation group (14.19%) compared to the roscovitine treatment and contact inhibition groups (21.31 and 20.32% respectively, P < 0.05). A total of 1070 transgenic cloned embryos derived from the three treatment groups were transferred to surrogate sows; one pregnancy was established and three embryos from the roscovitine treatment group successfully completed gestation. These results indicate that the roscovitine treatment was more effective at synchronizing transgenic kidney cells in Bama mini-pigs and allowed reconstructed embryos to develop to full term. PMID:26662422

  4. Nuclear transfer nTreg model reveals fate-determining TCR-β and novel peripheral nTreg precursors

    PubMed Central

    Ku, Manching; Chang, Shih-En; Hernandez, Julio; Abadejos, Justin R.; Sabouri-Ghomi, Mohsen; Muenchmeier, Niklas J.; Schwarz, Anna; Valencia, Anna M.; Kirak, Oktay

    2016-01-01

    To study the development and function of “natural-arising” T regulatory (nTreg) cells, we developed a novel nTreg model on pure nonobese diabetic background using epigenetic reprogramming via somatic cell nuclear transfer. On RAG1-deficient background, we found that monoclonal FoxP3+ CD4+ Treg cells developed in the thymus in the absence of other T cells. Adoptive transfer experiments revealed that the thymic niche is not a limiting factor in nTreg development. In addition, we showed that the T-cell receptor (TCR) β-chain of our nTreg model was not only sufficient to bias T-cell development toward the CD4 lineage, but we also demonstrated that this TCR β-chain was able to provide stronger TCR signals. This TCR-β–driven mechanism would thus unify former per se contradicting hypotheses of TCR-dependent and -independent nTreg development. Strikingly, peripheral FoxP3− CD4+ T cells expressing the same TCR as this somatic cell nuclear transfer nTreg model had a reduced capability to differentiate into Th1 cells but were poised to differentiate better into induced nTreg cells, both in vitro and in vivo, representing a novel peripheral precursor subset of nTreg cells to which we refer to as pre-nTreg cells. PMID:27044095

  5. Interspecies differences in metabolism of arsenic by cultured primary hepatocytes

    SciTech Connect

    Drobna, Zuzana; Walton, Felecia S.; Harmon, Anne W.; Thomas, David J.; Styblo, Miroslav

    2010-05-15

    Biomethylation is the major pathway for the metabolism of inorganic arsenic (iAs) in many mammalian species, including the human. However, significant interspecies differences have been reported in the rate of in vivo metabolism of iAs and in yields of iAs metabolites found in urine. Liver is considered the primary site for the methylation of iAs and arsenic (+3 oxidation state) methyltransferase (As3mt) is the key enzyme in this pathway. Thus, the As3mt-catalyzed methylation of iAs in the liver determines in part the rate and the pattern of iAs metabolism in various species. We examined kinetics and concentration-response patterns for iAs methylation by cultured primary hepatocytes derived from human, rat, mice, dog, rabbit, and rhesus monkey. Hepatocytes were exposed to [{sup 73}As]arsenite (iAs{sup III}; 0.3, 0.9, 3.0, 9.0 or 30 nmol As/mg protein) for 24 h and radiolabeled metabolites were analyzed in cells and culture media. Hepatocytes from all six species methylated iAs{sup III} to methylarsenic (MAs) and dimethylarsenic (DMAs). Notably, dog, rat and monkey hepatocytes were considerably more efficient methylators of iAs{sup III} than mouse, rabbit or human hepatocytes. The low efficiency of mouse, rabbit and human hepatocytes to methylate iAs{sup III} was associated with inhibition of DMAs production by moderate concentrations of iAs{sup III} and with retention of iAs and MAs in cells. No significant correlations were found between the rate of iAs methylation and the thioredoxin reductase activity or glutathione concentration, two factors that modulate the activity of recombinant As3mt. No associations between the rates of iAs methylation and As3mt protein structures were found for the six species examined. Immunoblot analyses indicate that the superior arsenic methylation capacities of dog, rat and monkey hepatocytes examined in this study may be associated with a higher As3mt expression. However, factors other than As3mt expression may also contribute to

  6. Specific features of external heat and mass transfer in the vibration apparatuses used for regenerating spent fuel from nuclear power plants

    NASA Astrophysics Data System (ADS)

    Sapozhnikov, B. G.; Gorbunova, A. M.; Zelenkova, Yu. O.; Sapozhnikov, G. B.; Shiryaeva, N. P.

    2014-06-01

    We present experimental data on the coefficients of heat and mass transfer for freely floating bodies simulating fragments of cladding and large conglomerates of fuel, as well as on the local coefficients of heat and mass transfer over the bed height, which point to high intensity of heat and mass transfer processes that take place in the elements of vibration apparatuses intended for subjecting spent fuel from nuclear power plants to oxidative recrystallization.

  7. Sensors and nuclear power. Report by the Technology Transfer Sensors Task Team

    SciTech Connect

    Not Available

    1985-06-01

    The existing sensor systems for the basic process parameters in nuclear power plant operation have limitations with respect to accuracy, ease of maintenance and signal processing. These limitations comprise the economy of nuclear power generation. To reduce the costs and improve performance of nuclear power plant fabrication, operation, maintenance and repair we need to advance the sensor technology being applied in the nuclear industry. The economic viability and public acceptance of nuclear power will depend on how well we direct and apply technological advances to the industry. This report was prepared by a team with members representing a wide range of the nuclear industry embracing the university programs, national laboratories, architect engineers and reactor manufacturers. An intensive effort was made to survey current sensor technology, evaluate future trends and determine development needs. This included literature surveys, visits with utilities, universities, laboratories and organizations outside the nuclear industry. Several conferences were attended to take advantage of the access to experts in selected topics and to obtain opinions. Numerous telephone contacts and exchanges by mail supplemented the above efforts. Finally, the broad technical depth of the team members provided the basis for the stimulating working sessions during which this report was organized and drafted.

  8. Summarizing lecture: factors influencing enzymatic H-transfers, analysis of nuclear tunnelling isotope effects and thermodynamic versus specific effects

    PubMed Central

    Marcus, R.A

    2006-01-01

    In the articles in this Discussion, a wide variety of topics are treated, including reorganization energy, initially introduced for electron transfers (‘environmentally assisted tunnelling’), nuclear tunnelling, H/D and C12/C13 kinetic isotope effects (KIEs), the effect of changes of distal and nearby amino acid residues using site-directed mutagenesis, and dynamics versus statistical effects. A coordinate-free form of semi-classical theory is used to examine topics on data such as tunnelling versus ‘over-the-barrier’ paths and temperature and pressure effects on KIEs. The multidimensional semi-classical theory includes classically allowed and classically forbidden transitions. More generally, we address the question of relating kinetic to thermodynamic factors, as in the electron transfer field, so learning about specific versus thermodynamic effects in enzyme catalysis and KIEs. PMID:16873131

  9. Radiation heat transfer calculations for the uranium fuel-containment region of the nuclear light bulb engine.

    NASA Technical Reports Server (NTRS)

    Rodgers, R. J.; Latham, T. S.; Krascella, N. L.

    1971-01-01

    Calculation results are reviewed of the radiant heat transfer characteristics in the fuel and buffer gas regions of a nuclear light bulb engine based on the transfer of energy by thermal radiation from gaseous uranium fuel in a neon vortex, through an internally cooled transparent wall, to seeded hydrogen propellant. The results indicate that the fraction of UV energy incident on the transparent walls increases with increasing power level. For the reference engine power level of 4600 megw, it is necessary to employ space radiators to reject the UV radiated energy absorbed by the transparent walls. This UV energy can be blocked by employing nitric oxide and oxygen seed gases in the fuel and buffer gas regions. However, this results in increased UV absorption in the buffer gas which also requires space radiators to reject the heat load.

  10. CFD Simulations of a Flow Mixing and Heat Transfer Enhancement in an Advanced LWR Nuclear Fuel Assembly

    SciTech Connect

    In, Wang-Kee; Chun, Tae-Hyun; Shin, Chang-Hwan; Oh, Dong-Seok

    2007-07-01

    A computational fluid dynamics (CFD) analysis has been performed to investigate a flow-mixing and heat-transfer enhancement caused by a mixing-vane spacer in a LWR fuel assembly which is a rod bundle. This paper presents the CFD simulations of a flow mixing and heat transfer in a fully heated 5x5 array of a rod bundle with a split-vane and hybrid-vane spacer. The CFD prediction at a low Reynolds number of 42,000 showed a reasonably good agreement of the initial heat transfer enhancement with the measured one for a partially heated experiment using a similar spacer structure. The CFD simulation also predicted the decay rate of a normalized Nusselt number downstream of the split-vane spacer which agrees fairly well with those of the experiment and the correlation. The CFD calculations for the split vane and hybrid vane at the LWR operating conditions(Re = 500,000) predicted hot fuel spots in a streaky structure downstream of the spacer, which occurs due to the secondary flow occurring in an opposite direction near the fuel rod. However, the split-vane and hybrid-vane spacers are predicted to significantly enhance the overall heat transfer of a LWR nuclear fuel assembly. (authors)

  11. Nuclear spectroscopy study of the isotopes populated via multinucleon transfer in the 90Zr + 208Pb reaction

    SciTech Connect

    Ur, C. A.; Corradi, L.; Stefanini, A. M.; Behera, B. R.; Fioretto, E.; Gadea, A.; Latina, A.; Szilner, S.; Beghini, S.; Farnea, E.; Montagnoli, G.; Scarlassara, F.; Haas, F.; Pollarolo, G.

    2006-08-14

    The present work takes advantage of the multinucleon transfer mechanism between heavy reaction partners to study the population pattern of excited nuclear states in near spherical Zirconium isotopes following the 90Zr + 208Pb reaction at an energy closed to the Coulomb barrier. Both the projectile and the target are well known closed shell nuclei offering an optimum situation for clean experimental and theoretical conditions. Total kinetic energy loss (TKEL) distributions were compared with calculations performed with the GRAZING code. The ability to use the TKEL as a selection tool for the states at different excitation energies was shown.

  12. Thermal expansion of UO2+x nuclear fuel rods from a model coupling heat transfer and oxygen diffusion

    SciTech Connect

    Mihaila, Bogden; Zubelewicz, Aleksander; Stan, Marius; Ramirez, Juan

    2008-01-01

    We study the thermal expansion of UO{sub 2+x} nuclear fuel rod in the context of a model coupling heat transfer and oxygen diffusion discussed previously by J.C. Ramirez, M. Stan and P. Cristea [J. Nucl. Mat. 359 (2006) 174]. We report results of simulations performed for steady-state and time-dependent regimes in one-dimensional configurations. A variety of initial- and boundary-value scenarios are considered. We use material properties obtained from previously published correlations or from analysis of previously published data. All simulations were performed using the commercial code COMSOL Multiphysics{sup TM} and are readily extendable to include multidimensional effects.

  13. Inter-species interconnections in acid mine drainage microbial communities

    PubMed Central

    Comolli, Luis R.; Banfield, Jill F.

    2014-01-01

    Metagenomic studies are revolutionizing our understanding of microbes in the biosphere. They have uncovered numerous proteins of unknown function in tens of essentially unstudied lineages that lack cultivated representatives. Notably, few of these microorganisms have been visualized, and even fewer have been described ultra-structurally in their essentially intact, physiologically relevant states. Here, we present cryogenic transmission electron microscope (cryo-TEM) 2D images and 3D tomographic datasets for archaeal species from natural acid mine drainage (AMD) microbial communities. Ultrastructural findings indicate the importance of microbial interconnectedness via a range of mechanisms, including direct cytoplasmic bridges and pervasive pili. The data also suggest a variety of biological structures associated with cell-cell interfaces that lack explanation. Some may play roles in inter-species interactions. Interdependences amongst the archaea may have confounded prior isolation efforts. Overall, the findings underline knowledge gaps related to archaeal cell components and highlight the likely importance of co-evolution in shaping microbial lineages. PMID:25120533

  14. Interspecies tunneling in one-dimensional Bose mixtures

    SciTech Connect

    Pflanzer, Anika C.; Zoellner, Sascha; Schmelcher, Peter

    2010-02-15

    We study the ground-state properties and quantum dynamics of few-boson mixtures with strong interspecies repulsion in one-dimensional traps. If one species localizes at the center, e.g., due to a very large mass compared to the other component, it represents an effective barrier for the latter, and the system can be mapped onto identical bosons in a double well. For weaker localization, the barrier atoms begin to respond to the light component, leading to an induced attraction between the mobile atoms that may even outweigh their bare intraspecies repulsion. To explain the resulting effects, we derive an effective Hubbard model for the lighter species accounting for the back action of the barrier in correction terms to the lattice parameters. Also the tunneling is drastically affected: by varying the degree of localization of the 'barrier' atoms, the dynamics of intrinsically noninteracting bosons can change from Rabi oscillations to effective pair tunneling. For identical fermions (or fermionized bosons), this leads to the tunneling of attractively bound pairs.

  15. An interspecies comparison of the temporomandibular joint disc.

    PubMed

    Kalpakci, K N; Willard, V P; Wong, M E; Athanasiou, K A

    2011-02-01

    The temporomandibular joint (TMJ) disc plays a critical role in normal function of the joint, and many disorders of the TMJ are a result of disc dysfunction. Previous quantitative TMJ characterization studies examined either the human or a specific animal model, but no single study has compared different species, in the belief that differences in joint morphology, function, and diet would be reflected in the material properties of the disc. In this study, we examined topographical biochemical (collagen, glycosaminoglycan, and DNA content) and biomechanical (tensile and compressive) properties of the human TMJ disc, and also discs from the cow, goat, pig, and rabbit. Regional and interspecies variations were identified in all parameters measured, and certain disc characteristics were observed across all species, such as a weak intermediate zone under mediolateral tension. While human discs possessed properties distinct from those of the other species, pig discs were most similar to the human, suggesting that the pig may be a suitable animal model for TMJ bioengineering efforts.

  16. Interspecies Interaction between Pseudomonas aeruginosa and Other Microorganisms

    PubMed Central

    Tashiro, Yosuke; Yawata, Yutaka; Toyofuku, Masanori; Uchiyama, Hiroo; Nomura, Nobuhiko

    2013-01-01

    Microbes interact with each other in multicellular communities and this interaction enables certain microorganisms to survive in various environments. Pseudomonas aeruginosa is a highly adaptable bacterium that ubiquitously inhabits diverse environments including soil, marine habitats, plants and animals. Behind this adaptivity, P. aeruginosa has abilities not only to outcompete others but also to communicate with each other to develop a multispecies community. In this review, we focus on how P. aeruginosa interacts with other microorganisms. P. aeruginosa secretes antimicrobial chemicals to compete and signal molecules to cooperate with other organisms. In other cases, it directly conveys antimicrobial enzymes to other bacteria using the Type VI secretion system (T6SS) or membrane vesicles (MVs). Quorum sensing is a central regulatory system used to exert their ability including antimicrobial effects and cooperation with other microbes. At least three quorum sensing systems are found in P. aeruginosa, Las, Rhl and Pseudomonas quinolone signal (PQS) systems. These quorum-sensing systems control the synthesis of extracellular antimicrobial chemicals as well as interaction with other organisms via T6SS or MVs. In addition, we explain the potential of microbial interaction analysis using several micro devices, which would bring fresh sensitivity to the study of interspecies interaction between P. aeruginosa and other organisms. PMID:23363620

  17. Nutritional consequences of interspecies differences in arginine and lysine metabolism.

    PubMed

    Ball, Ronald O; Urschel, Kristine L; Pencharz, Paul B

    2007-06-01

    Differences in lysine and arginine requirements among various species such as omnivores (humans, pigs, rats, dogs), carnivores (cats), herbivores (rabbits, horses), ruminants (cattle), poultry, and fish, are covered in detail in this article. Although lysine is classified as an indispensable amino acid across species, the classification of arginine as either an indispensable or dispensable amino acid is more ambiguous because of differences among species in rates of de novo arginine synthesis. Because lysine is most often the limiting amino acid in the diet, its requirement has been extensively studied. By use of the ideal protein concept, the requirements of the other indispensable amino acids can be extrapolated from the lysine requirement. The successful use of this concept in pigs is compared with potential application of the ideal protein concept in humans. The current dietary arginine requirement varies widely among species, with ruminants, rabbits, and rats having relatively low requirements and carnivores, fish, and poultry having high requirements. Interspecies differences in metabolic arginine utilization and reasons for different rates of de novo arginine synthesis are reviewed in detail, as these are the primary determinants of the dietary arginine requirement. There is presently no dietary requirement for humans of any age, although this needs to be reassessed, particularly in neonates. A thorough understanding of the factors contributing to the lysine and arginine requirements in different species will be useful in our understanding of human amino acid requirements.

  18. Importance of toxicokinetics for interspecies variation in sensitivity to chemicals.

    PubMed

    Nyman, Anna-Maija; Schirmer, Kristin; Ashauer, Roman

    2014-05-20

    Interspecies variation in sensitivity to synthetic chemicals can be orders of magnitude large. Species traits causing the variation can be related to toxicokinetics (uptake, distribution, biotransformation, elimination) or toxicodynamics (interaction with biological target sites). We present an approach to systematically measure and model the contribution of uptake, biotransformation, internal distribution, and elimination kinetics toward species sensitivity differences. The aim is to express sensitivity as target tissue specific, internal lethal concentrations. A case study with the pesticides diazinon, imidacloprid, and propiconazole and the aquatic invertebrates Gammarus pulex, Gammarus fossarum, and Lymnaea stagnalis illustrates the approach. L. stagnalis accumulates more pesticides than Gammaridae when measured in whole organisms but less in target tissues such as the nervous system. Toxicokinetics, i.e. biotransformation and distribution, explain the higher tolerance of L. stagnalis to the insecticide diazinon when compared to Gammaridae. L. stagnalis was again more tolerant to the other neurotoxicant imidacloprid; however, the difference in sensitivity could not be explained by toxicokinetics alone, indicating the importance of toxicodynamic differences. Sensitivity to propiconazole was comparable among all species and, when expressed as internal lethal concentrations, falls in the range of baseline toxicity.

  19. Conventional and Bimodal Nuclear Thermal Rocket (NTR) Artificial Gravity Mars Transfer Vehicle Concepts

    NASA Technical Reports Server (NTRS)

    Borowski, Stanley K.; McCurdy, David R.; Packard, Thomas W.

    2014-01-01

    A variety of countermeasures have been developed to address the debilitating physiological effects of "zero-gravity" (0-g) experienced by cosmonauts and astronauts during their approximately 0.5-1.2 year long stays in LEO (Low Earth Orbit). Longer interplanetary flights, combined with possible prolonged stays in Mars orbit, could subject crewmembers to up to approximately 2.5 years of weightlessness. In view of known and recently diagnosed problems associated with 0-g, an artificial gravity spacecraft offers many advantages and may indeed be an enabling technology for human flights to Mars. A number of important human factors must be taken into account in selecting the rotation radius, rotation rate, and orientation of the habitation module or modules. These factors include the gravity gradient effect, radial and tangential Coriolis forces, along with cross-coupled acceleration effects. Artificial gravity (AG) Mars transfer vehicle (MTV) concepts are presented that utilize both conventional NTR, as well as, enhanced "bimodal" nuclear thermal rocket (BNTR) propulsion. The NTR is a proven technology that generates high thrust and has a specific impulse (I (sub sp)) capability of approximately 900 s - twice that of today's best chemical rockets. The AG/MTV concepts using conventional NTP carry twin cylindrical "ISS-type" habitation modules with their long axes oriented either perpendicular or parallel to the longitudinal spin axis of the MTV and utilize photovoltaic arrays (PVAs) for spacecraft power. The twin habitat modules are connected to a central operations hub located at the front of the MTV via two pressurized tunnels that provide the rotation radius for the habitat modules. For the BNTR AG/MTV option, each engine has its own "closed" secondary helium-xenon gas loop and Brayton rotating unit that can generate tens of kilowatts (kW (sub e)) of spacecraft electrical power during the mission coast phase eliminating the need for large PVAs. A single inflatable

  20. Engineering design elements of a two-phase thermosyphon to transfer nuclear thermal energy to a hydrogen plant

    NASA Astrophysics Data System (ADS)

    Sabharwall, Piyush

    Two hydrogen production processes, both powered by Next Generation Nuclear Plant (NGNP), are currently under investigation at the Idaho National Laboratory. The first is high-temperature steam electrolysis utilizing both heat and electricity and the second is thermo-chemical production through the sulfur-iodine process primarily utilizing heat. Both processes require high temperature (>850°C) for enhanced efficiency; temperatures indicative of NGNP. Safety and licensing mandates prudently dictate that the NGNP and the hydrogen production facility be physically isolated, perhaps requiring separation of over 100m. There are several options to transferring multi-megawatt thermal power over such a distance. One option is simply to produce only electricity, transfer by wire to the hydrogen plant, and then reconvert the electric energy to heat via Joule or induction heating. Electrical transport, however, suffers energy losses of 60-70% due to the thermal to electric conversion inherent in the Brayton cycle. A second option is thermal energy transport via a single-phase forced convection loop where a fluid is mechanically pumped between heat exchangers at the nuclear and hydrogen plants. High temperatures, however, present unique materials and pumping challenges. Single phase, low pressure helium is an attractive option for NGNP, but is not suitable for a single purpose facility dictated to hydrogen production because low pressure helium requires higher pumping power and makes the process very inefficient. A third option is two-phase heat transfer utilizing a high temperature thermosyphon. Heat transport occurs via evaporation and condensation, and the heat transport fluid is re-circulated by gravitational force. Thermosyphon has the capability to transport heat at high rates over appreciable distances, virtually isothermally and without any requirement for external pumping devices. For process heat, intermediate heat exchangers (IHX) are desired to transfer heat from

  1. Establishment of pregnancy after the transfer of nuclear transfer embryos produced from the fusion of argali (Ovis ammon) nuclei into domestic sheep (Ovis aries) enucleated oocytes.

    PubMed

    White, K L; Bunch, T D; Mitalipov, S; Reed, W A

    1999-01-01

    Cloning mammalian species from cell lines of adult animals has been demonstrated. Aside from its importance for cloning multiple copies of genetically valuable livestock, cloning now has the potential to salvage endangered or even extinct species. The aim of this study was to investigate the effect of the bovine and domestic (Ovis aries) ovine oocyte cytoplasm on the nucleus of an established cell line from an endangered argali wild sheep (Ovis ammon) after nuclear transplantation. A fibroblast cell line was established from skin biopsies from an adult argali ram from the People's Republic of China. Early karyotype analysis of cells between 3-6 passages revealed a normal diploid chromosome number of 56. The argali karyotype consisted of 2 pairs of biarmed and 25 pairs of acrocentric autosomes, a large acrocentric and minute biarmed Y. Bovine ovaries were collected from a local abattoir, oocytes aspirated, and immediately placed in maturation medium consisting of M-199 containing 10% fetal bovine serum, 100 IU/mL penicillin, 100 microg/mL streptomycin, 0.5 microg/mL follicle-stimulating hormone (FSH), 5.0 microg/mL luetinizing hormone (LH) and 1.0 microg/mL estradiol. Ovine (O. aries) oocytes were collected at surgery 25 hours postonset of estrus from the oviducts of superovulated donor animals. All cultures were carried out at 39 degrees C in a humidified atmosphere of 5% CO2 and air. In vitro matured MII bovine oocytes were enucleated 16-20 hours after onset of maturation and ovine oocytes within 2-3 hours after collection. Enucleation was confirmed using Hoechst 33342 and UV light. The donor argali cells were synchronized in G0-G1 phase by culturing in Dulbecco's modified Eagle's medium (DMEM) plus 0.5% fetal bovine serum for 5-10 days. Fusion of nuclear donor cell to an enucleated oocyte (cytoplast) to produce nuclear transfer (NT) embryos was induced by 2 electric pulses of 1.4 kV/cm for 30 microsc. Fused NT embryos were activated after 24 hours of maturation

  2. The transfer of strontium-90 and caesium-137 to milk in a dairy herd grazing near a major nuclear installation.

    PubMed

    Sumerling, T J; Dodd, N J; Green, N

    1984-03-01

    A field investigation of the transfer of artificially produced radionuclides in the pasture--cow--milk pathway has been made at a farm close to the nuclear fuel reprocessing installation at Sellafield on the north-west coast of England. This paper reports results from analyses of samples collected during 1981, reports transfers coefficients for 90Sr and 137Cs from various types of feed to milk, and discusses factors that affect the transfer of these radionuclides. It is shown that during 1981 a large proportion of the 90Sr and 137Cs consumed by cattle grazing near Sellafield was derived from activity deposited in previous years. Transfer coefficients to milk, Fm, have been derived which are within the ranges of those observed in tracer and fallout studies. There are significant seasonal changes in transfer. For 90Sr, values of Fm between 9 X 10(-4)d 1(-1) and 4 X 10(-3)d 1(-1) have been obtained. It is concluded that this large range arises because daily intakes of 90Sr by the herd during the winter months are lower (by a factor of about 3) than intakes during the summer months and that the concentration of 90Sr in milk is not in equilibrium with intake, that is, the concentration of 90Sr in milk is maintained both by recent intakes and by remobilisation of activity that has been accumulated in bone from earlier intakes. For 137Cs, values of Fm between 3 X 10(-3)d 1(-1) and 9 X 10(-3)d 1(-1) have been obtained. It is concluded that this range most probably occurs because during the summer months, when the cows are grazing, a substantial proportion of the 137Cs intake is associated with soil on the surface of herbage and that, in this form, the 137Cs is less available for uptake from the digestive tract of the cows.

  3. Comparable frequencies of coding mutations and loss of imprinting in human pluripotent cells derived by nuclear transfer and defined factors.

    PubMed

    Johannesson, Bjarki; Sagi, Ido; Gore, Athurva; Paull, Daniel; Yamada, Mitsutoshi; Golan-Lev, Tamar; Li, Zhe; LeDuc, Charles; Shen, Yufeng; Stern, Samantha; Xu, Nanfang; Ma, Hong; Kang, Eunju; Mitalipov, Shoukhrat; Sauer, Mark V; Zhang, Kun; Benvenisty, Nissim; Egli, Dieter

    2014-11-01

    The recent finding that reprogrammed human pluripotent stem cells can be derived by nuclear transfer into human oocytes as well as by induced expression of defined factors has revitalized the debate on whether one approach might be advantageous over the other. Here we compare the genetic and epigenetic integrity of human nuclear-transfer embryonic stem cell (NT-ESC) lines and isogenic induced pluripotent stem cell (iPSC) lines, derived from the same somatic cell cultures of fetal, neonatal, and adult origin. The two cell types showed similar genome-wide gene expression and DNA methylation profiles. Importantly, NT-ESCs and iPSCs had comparable numbers of de novo coding mutations, but significantly more than parthenogenetic ESCs. As iPSCs, NT-ESCs displayed clone- and gene-specific aberrations in DNA methylation and allele-specific expression of imprinted genes. The occurrence of these genetic and epigenetic defects in both NT-ESCs and iPSCs suggests that they are inherent to reprogramming, regardless of derivation approach.

  4. MicroRNA-34c Expression in Donor Cells Influences the Early Development of Somatic Cell Nuclear Transfer Bovine Embryos

    PubMed Central

    Wang, Bo; Wang, Yongsheng; Zhang, Man; Du, Yue; Zhang, Yijun; Xing, Xupeng; Zhang, Lei; Su, JianMin

    2014-01-01

    Abstract The essence of the reprogramming activity of somatic cell nuclear transfer (SCNT) embryos is to produce normal fertilized embryos. However, reprogramming of somatic cells is not as efficient as the reprogramming of sperm. In this report, we describe the effect of an inducible, specific miR-34 microRNA expression in donor cells that enables a similar level of sperm:transgene expression on the early development of SCNT embryos. Our results showed that donor cells with doxycycline (dox)-induced miR-34c expression for the preparation of SCNT embryos resulted in altered developmental rates, histone modification (H3K9ac and H3K4me3), and extent of apoptosis. The cleavage rate and blastocyst formation of the induced nuclear transfer (NT) group were significantly increased. The immunofluorescence signal of H3K9ac in embryos in the induced NT group significantly increased in two-cell- and eight-cell-stage embryos; that of H3K4me3 increased significantly in eight-cell-stage embryos. Although significant differences in staining signals of apoptosis were not detected between groups, lower apoptosis levels were observed in the induced NT group. In conclusion, miR-34c expression induced by dox treatment enhances the developmental potential of SCNT embryos, modifies the epigenetic status, and changes blastocyst quality. PMID:25437869

  5. Generation of embryonic stem cells from mouse adipose-tissue derived cells via somatic cell nuclear transfer.

    PubMed

    Qin, Yiren; Qin, Jilong; Zhou, Chikai; Li, Jinsong; Gao, Wei-Qiang

    2015-01-01

    Somatic cells can be reprogrammed into embryonic stem cells (ESCs) by nuclear transfer (NT-ESCs), or into induced pluripotent stem cells (iPSCs) by the "Yamanaka method." However, recent studies have indicated that mouse and human iPSCs are prone to epigenetic and transcriptional aberrations, and that NT-ESCs correspond more closely to ESCs derived from in vitro fertilized embryos than iPSCs. In addition, the procedure of NT-ESCs does not involve gene modification. Demonstration of generation of NT-ESCs using an easily-accessible source of adult cell types would be very important. Adipose tissue is a source of readily accessible donor cells and can be isolated from both males and females at different ages. Here we report that NT-ESCs can be generated from adipose tissue-derived cells (ADCs). At morphological, mRNA and protein levels, these NT-ESCs show classic ESC colonies, exhibit alkaline phosphatase (AP) activity, and display normal diploid karyotypes. Importantly, these cells express pluripotent markers including Oct4, Sox2, Nanog and SSEA-1. Furthermore, they can differentiate in vivo into various types of cells from 3 germinal layers by teratoma formation assays. This study demonstrates for the first time that ESCs can be generated from the adipose tissue by somatic cell nuclear transfer (SCNT) and suggests that ADCs can be a new donor-cell type for potential therapeutic cloning.

  6. Generation of embryonic stem cells from mouse adipose-tissue derived cells via somatic cell nuclear transfer

    PubMed Central

    Qin, Yiren; Qin, Jilong; Zhou, Chikai; Li, Jinsong; Gao, Wei-Qiang

    2015-01-01

    Somatic cells can be reprogrammed into embryonic stem cells (ESCs) by nuclear transfer (NT-ESCs), or into induced pluripotent stem cells (iPSCs) by the “Yamanaka method.” However, recent studies have indicated that mouse and human iPSCs are prone to epigenetic and transcriptional aberrations, and that NT-ESCs correspond more closely to ESCs derived from in vitro fertilized embryos than iPSCs. In addition, the procedure of NT-ESCs does not involve gene modification. Demonstration of generation of NT-ESCs using an easily-accessible source of adult cell types would be very important. Adipose tissue is a source of readily accessible donor cells and can be isolated from both males and females at different ages. Here we report that NT-ESCs can be generated from adipose tissue-derived cells (ADCs). At morphological, mRNA and protein levels, these NT-ESCs show classic ESC colonies, exhibit alkaline phosphatase (AP) activity, and display normal diploid karyotypes. Importantly, these cells express pluripotent markers including Oct4, Sox2, Nanog and SSEA-1. Furthermore, they can differentiate in vivo into various types of cells from 3 germinal layers by teratoma formation assays. This study demonstrates for the first time that ESCs can be generated from the adipose tissue by somatic cell nuclear transfer (SCNT) and suggests that ADCs can be a new donor-cell type for potential therapeutic cloning. PMID:25692793

  7. Robotics and nuclear power. Report by the Technology Transfer Robotics Task Team

    SciTech Connect

    Not Available

    1985-06-01

    A task team was formed at the request of the Department of Energy to evaluate and assess technology development needed for advanced robotics in the nuclear industry. The mission of these technologies is to provide the nuclear industry with the support for the application of advanced robotics to reduce nuclear power generating costs and enhance the safety of the personnel in the industry. The investigation included robotic and teleoperated systems. A robotic system is defined as a reprogrammable, multifunctional manipulator designed to move materials, parts, tools, or specialized devices through variable programmed motions for the performance of a variety of tasks. A teleoperated system includes an operator who remotely controls the system by direct viewing or through a vision system.

  8. Factors Determining the Efficiency of Porcine Somatic Cell Nuclear Transfer: Data Analysis with Over 200,000 Reconstructed Embryos.

    PubMed

    Liu, Tianbin; Dou, Hongwei; Xiang, Xi; Li, Lin; Li, Yong; Lin, Lin; Pang, Xinzhi; Zhang, Yijie; Chen, Yu; Luan, Jing; Xu, Ying; Yang, Zhenzhen; Yang, Wenxian; Liu, Huan; Li, Feida; Wang, Hui; Yang, Huanming; Bolund, Lars; Vajta, Gabor; Du, Yutao

    2015-12-01

    Data analysis in somatic cell nuclear transfer (SCNT) research is usually limited to several hundreds or thousands of reconstructed embryos. Here, we report mass results obtained with an established and consistent porcine SCNT system (handmade cloning [HMC]). During the experimental period, 228,230 reconstructed embryos and 82,969 blastocysts were produced. After being transferred into 656 recipients, 1070 piglets were obtained. First, the effects of different types of donor cells, including fetal fibroblasts (FFs), adult fibroblasts (AFs), adult preadipocytes (APs), and adult blood mesenchymal (BM) cells, were investigated on the further in vitro and in vivo development. Compared to adult donor cells (AFs, APs, BM cells, respectively), FF cells resulted in a lower blastocyst/reconstructed embryo rate (30.38% vs. 37.94%, 34.65%, and 34.87%, respectively), but a higher overall efficiency on the number of piglets born alive per total blastocysts transferred (1.50% vs. 0.86%, 1.03%, and 0.91%, respectively) and a lower rate of developmental abnormalities (10.87% vs. 56.57%, 24.39%, and 51.85%, respectively). Second, recloning was performed with cloned adult fibroblasts (CAFs) and cloned fetal fibroblasts (CFFs). When CAFs were used as the nuclear donor, fewer developmental abnormalities and higher overall efficiency were observed compared to AFs (56.57% vs. 28.13% and 0.86% vs. 1.59%, respectively). However, CFFs had an opposite effect on these parameters when compared with CAFs (94.12% vs. 10.87% and 0.31% vs. 1.50%, respectively). Third, effects of genetic modification on the efficiency of SCNT were investigated with transgenic fetal fibroblasts (TFFs) and gene knockout fetal fibroblasts (KOFFs). Genetic modification of FFs increased developmental abnormalities (38.96% and 25.24% vs. 10.87% for KOFFs, TFFs, and FFs, respectively). KOFFs resulted in lower overall efficiency compared to TFFs and FFs (0.68% vs. 1.62% and 1.50%, respectively). In conclusion, this is the

  9. Nuclear and Q{sup 2} dependence of quaselastic (e,e{prime}p) scattering at large momentum transfer

    SciTech Connect

    Jackson, H.E.; Geesaman, D.F.; Jones, C.E.

    1995-08-01

    An experiment was completed at the Stanford Linear Accelerator Center in which measurements of the (e,e{prime}p) coincidence quasielastic cross section in nuclei were extended to the largest possible Q{sup 2} attainable with the Nuclear Physics Injector and the End Station A spectrometers. Coincidence measurements of the quasielastic (e,e{prime}p) cross section were made on nuclei from carbon to gold in the Q{sup 2} range of 1-7 (GeV/c){sup 2}. Several papers describing the results were published or submitted. Analysis of the data is in its final stages. In summary, the cross section for quasielastic {sup 12}C(e,e{prime}p) scattering was measured at momentum transfer Q{sup 2}=1, 3, 5, and 6.8 (GeV/c){sup 2}. The results are consistent with scattering from a single nucleon as the dominant process. The nuclear transparency is obtained and compared with theoretical calculations that incorporate color transparency effects. No significant rise of the transparency with Q{sup 2} is observed. Cross sections were reported for the reaction {sup 2}H(e,e{prime}p)n for momentum transfers in the range 1.2 {<=}Q{sup 2}{<=}6.8 (GeV/c){sup 2} and for missing momenta from 0 to 250 MeV/c. The longitudinal-transverse interference structure function was separated at Q{sup 2}=1.5 (GeV/c){sup 2}. The observables were compared to calculations performed in nonrelativistic and relativistic frameworks. The data are best described by a fully relativistic calculation. The A-dependence of the quasielastic A(e,e{prime}p) reaction was studied with {sup 2}H, C, Fe, and Au nuclei at momentum transfers Q{sup 2}=1, 3, 5, and 6.8 (GeV/c){sup 2}. The nuclear transparency T A,Q{sup 2}, a measure of the average probability that the struck proton escapes from the nucleu A without interaction, was extracted. Several calculations predict a significant increase in T with momentum transfer, a phenomenon known as color transparency. No significant rise within errors is seen for any of the nuclei studied.

  10. Nuclear fragmentation energy and momentum transfer distributions in relativistic heavy-ion collisions

    NASA Technical Reports Server (NTRS)

    Khandelwal, Govind S.; Khan, Ferdous

    1989-01-01

    An optical model description of energy and momentum transfer in relativistic heavy-ion collisions, based upon composite particle multiple scattering theory, is presented. Transverse and longitudinal momentum transfers to the projectile are shown to arise from the real and absorptive part of the optical potential, respectively. Comparisons of fragment momentum distribution observables with experiments are made and trends outlined based on our knowledge of the underlying nucleon-nucleon interaction. Corrections to the above calculations are discussed. Finally, use of the model as a tool for estimating collision impact parameters is indicated.

  11. Interspecies variations in Bordetella catecholamine receptor gene regulation and function.

    PubMed

    Brickman, Timothy J; Suhadolc, Ryan J; Armstrong, Sandra K

    2015-12-01

    Bordetella bronchiseptica can use catecholamines to obtain iron from transferrin and lactoferrin via uptake pathways involving the BfrA, BfrD, and BfrE outer membrane receptor proteins, and although Bordetella pertussis has the bfrD and bfrE genes, the role of these genes in iron uptake has not been demonstrated. In this study, the bfrD and bfrE genes of B. pertussis were shown to be functional in B. bronchiseptica, but neither B. bronchiseptica bfrD nor bfrE imparted catecholamine utilization to B. pertussis. Gene fusion analyses found that expression of B. bronchiseptica bfrA was increased during iron starvation, as is common for iron receptor genes, but that expression of the bfrD and bfrE genes of both species was decreased during iron limitation. As shown previously for B. pertussis, bfrD expression in B. bronchiseptica was also dependent on the BvgAS virulence regulatory system; however, in contrast to the case in B. pertussis, the known modulators nicotinic acid and sulfate, which silence Bvg-activated genes, did not silence expression of bfrD in B. bronchiseptica. Further studies using a B. bronchiseptica bvgAS mutant expressing the B. pertussis bvgAS genes revealed that the interspecies differences in bfrD modulation are partly due to BvgAS differences. Mouse respiratory infection experiments determined that catecholamine utilization contributes to the in vivo fitness of B. bronchiseptica and B. pertussis. Additional evidence of the in vivo importance of the B. pertussis receptors was obtained from serologic studies demonstrating pertussis patient serum reactivity with the B. pertussis BfrD and BfrE proteins. PMID:26371128

  12. Interspecies Variations in Bordetella Catecholamine Receptor Gene Regulation and Function

    PubMed Central

    Brickman, Timothy J.; Suhadolc, Ryan J.

    2015-01-01

    Bordetella bronchiseptica can use catecholamines to obtain iron from transferrin and lactoferrin via uptake pathways involving the BfrA, BfrD, and BfrE outer membrane receptor proteins, and although Bordetella pertussis has the bfrD and bfrE genes, the role of these genes in iron uptake has not been demonstrated. In this study, the bfrD and bfrE genes of B. pertussis were shown to be functional in B. bronchiseptica, but neither B. bronchiseptica bfrD nor bfrE imparted catecholamine utilization to B. pertussis. Gene fusion analyses found that expression of B. bronchiseptica bfrA was increased during iron starvation, as is common for iron receptor genes, but that expression of the bfrD and bfrE genes of both species was decreased during iron limitation. As shown previously for B. pertussis, bfrD expression in B. bronchiseptica was also dependent on the BvgAS virulence regulatory system; however, in contrast to the case in B. pertussis, the known modulators nicotinic acid and sulfate, which silence Bvg-activated genes, did not silence expression of bfrD in B. bronchiseptica. Further studies using a B. bronchiseptica bvgAS mutant expressing the B. pertussis bvgAS genes revealed that the interspecies differences in bfrD modulation are partly due to BvgAS differences. Mouse respiratory infection experiments determined that catecholamine utilization contributes to the in vivo fitness of B. bronchiseptica and B. pertussis. Additional evidence of the in vivo importance of the B. pertussis receptors was obtained from serologic studies demonstrating pertussis patient serum reactivity with the B. pertussis BfrD and BfrE proteins. PMID:26371128

  13. Interspecies variations in Bordetella catecholamine receptor gene regulation and function.

    PubMed

    Brickman, Timothy J; Suhadolc, Ryan J; Armstrong, Sandra K

    2015-12-01

    Bordetella bronchiseptica can use catecholamines to obtain iron from transferrin and lactoferrin via uptake pathways involving the BfrA, BfrD, and BfrE outer membrane receptor proteins, and although Bordetella pertussis has the bfrD and bfrE genes, the role of these genes in iron uptake has not been demonstrated. In this study, the bfrD and bfrE genes of B. pertussis were shown to be functional in B. bronchiseptica, but neither B. bronchiseptica bfrD nor bfrE imparted catecholamine utilization to B. pertussis. Gene fusion analyses found that expression of B. bronchiseptica bfrA was increased during iron starvation, as is common for iron receptor genes, but that expression of the bfrD and bfrE genes of both species was decreased during iron limitation. As shown previously for B. pertussis, bfrD expression in B. bronchiseptica was also dependent on the BvgAS virulence regulatory system; however, in contrast to the case in B. pertussis, the known modulators nicotinic acid and sulfate, which silence Bvg-activated genes, did not silence expression of bfrD in B. bronchiseptica. Further studies using a B. bronchiseptica bvgAS mutant expressing the B. pertussis bvgAS genes revealed that the interspecies differences in bfrD modulation are partly due to BvgAS differences. Mouse respiratory infection experiments determined that catecholamine utilization contributes to the in vivo fitness of B. bronchiseptica and B. pertussis. Additional evidence of the in vivo importance of the B. pertussis receptors was obtained from serologic studies demonstrating pertussis patient serum reactivity with the B. pertussis BfrD and BfrE proteins.

  14. Imaging of Amide Proton Transfer and Nuclear Overhauser Enhancement in Ischemic Stroke with Corrections for Competing Effects

    PubMed Central

    Li, Hua; Zu, Zhongliang; Zaiss, Moritz; Khan, Imad S.; Singer, Robert; Gochberg, Daniel F.; Bachert, Peter; Gore, John C.; Xu, Junzhong

    2015-01-01

    Chemical exchange saturation transfer (CEST) potentially provides the ability to detect small solute pools through indirect measurements of attenuated water signals. However, CEST effects may be diluted by various competing effects such as non-specific magnetization transfer (MT) and asymmetric MT effects, water longitudinal relaxation (T1), and direct water saturation (RF spillover). In the current study, CEST images were acquired in rats following ischemic stroke and analyzed by comparing the reciprocals of the CEST signals at three different saturation offsets. This combined approach corrects the above competing effects and provides a more robust signal metric sensitive specifically to proton exchange rate constant. The corrected amide proton transfer (APT) data show greater differences between the ischemic and the contralateral (non-ischemic) hemispheres. By contrast, corrected nuclear Overhauser enhancements (NOEs) around −3.5 ppm from water change over time in both hemispheres, indicating whole-brain changes that have not been reported before. This study may help better understand the contrast mechanisms of APT and NOE imaging in ischemic stroke, and also establishes a framework for future stroke measurements using CEST imaging with spillover-, MT- and T1-corrections. PMID:25483870

  15. No differences in sheep somatic cell nuclear transfer outcomes using serum-starved or actively growing donor granulosa cells.

    PubMed

    Peura, T T; Hartwich, K M; Hamilton, H M; Walker, S K

    2003-01-01

    The aim of this study was to compare serum-starved and non-starved donor cells in sheep nuclear transfer with a special emphasis on cloning outcomes. Sheep oocytes, derived either in vivo or in vitro, were fused with cultured serum-starved or actively growing adult granulosa cells. Resulting blastocysts were transferred to recipients fresh or after vitrification, and subsequent pregnancies followed to term. Donor cell treatment did not significantly affect preimplantation development, pregnancy rates, fetal loss or neonate survival rates. Of 22 lambs born, ten survived the immediate perinatal period but all succumbed at various timepoints within the first few weeks of life. The results of the study suggest that the use of serum-starved cells offers no advantages or disadvantages to cloning outcomes. Neither were significant differences in outcomes observed when using either in vivo- or in vitro-derived oocytes or embryos transferred fresh or after vitrification. Yet, these results continue to highlight problems associated with somatic cell cloning as indicated by offspring mortality. It remains unclear whether the high offspring mortality in the current study was related to species, associated with the cell lines used or the result of other causes. PMID:12921702

  16. Transfer of elements relevant to nuclear fuel cycle from soil to boreal plants and animals in experimental meso- and microcosms.

    PubMed

    Tuovinen, Tiina S; Kasurinen, Anne; Häikiö, Elina; Tervahauta, Arja; Makkonen, Sari; Holopainen, Toini; Juutilainen, Jukka

    2016-01-01

    Uranium (U), cobalt (Co), molybdenum (Mo), nickel (Ni), lead (Pb), thorium (Th) and zinc (Zn) occur naturally in soil but their radioactive isotopes can also be released into the environment during the nuclear fuel cycle. The transfer of these elements was studied in three different trophic levels in experimental mesocosms containing downy birch (Betula pubescens), narrow buckler fern (Dryopteris carthusiana) and Scandinavian small-reed (Calamagrostis purpurea ssp. Phragmitoides) as producers, snails (Arianta arbostorum) as herbivores, and earthworms (Lumbricus terrestris) as decomposers. To determine more precisely whether the element uptake of snails is mainly via their food (birch leaves) or both via soil and food, a separate microcosm experiment was also performed. The element uptake of snails did not generally depend on the presence of soil, indicating that the main uptake route was food, except for U, where soil contact was important for uptake when soil U concentration was high. Transfer of elements from soil to plants was not linear, i.e. it was not correctly described by constant concentration ratios (CR) commonly applied in radioecological modeling. Similar nonlinear transfer was found for the invertebrate animals included in this study: elements other than U were taken up more efficiently when element concentration in soil or food was low. PMID:26363398

  17. A multi-species based taxonomic microarray reveals interspecies hybridization and introgression in Saccharomyces cerevisiae

    PubMed Central

    Muller, Ludo A. H.; McCusker, John H.

    2009-01-01

    A multi-species based taxonomic microarray targeting coding sequences of diverged orthologous genes in Saccharomyces cerevisiae, S. paradoxus, S. mikatae, S. bayanus, S. kudriavzevii, Naumovia castellii, Lachancea kluyveri and Candida glabrata was designed to allow identification of isolates of these species and their interspecies hybrids. Analysis of isolates of several Saccharomyces species and interspecies hybrids demonstrated the ability of the microarray to differentiate these yeasts on the basis of their specific hybridization patterns. Subsequent analysis of 183 supposed S. cerevisiae isolates of various ecological and geographical backgrounds revealed one misclassified S. bayanus or S. uvarum isolate and four aneuploid interspecies hybrids, one between S. cerevisiae and S. bayanus and three between S. cerevisiae and S. kudriavzevii. Furthermore, this microarray design allowed the detection of multiple introgressed S. paradoxus DNA fragments in the genomes of three different S. cerevisiae isolates. These results show the power of multi-species based microarrays as taxonomic tools for the identification of species and interspecies hybrids, and their ability to provide a more detailed characterization of interspecies hybrids and recombinants. PMID:19054123

  18. Radiocesium transfer from hillslopes to the Pacific Ocean after the Fukushima Nuclear Power Plant accident: A review.

    PubMed

    Evrard, Olivier; Laceby, J Patrick; Lepage, Hugo; Onda, Yuichi; Cerdan, Olivier; Ayrault, Sophie

    2015-10-01

    The devastating tsunami triggered by the Great East Japan Earthquake on March 11, 2011 inundated the Fukushima Dai-ichi Nuclear Power Plant (FDNPP) resulting in a loss of cooling and a series of explosions releasing the largest quantity of radioactive material into the atmosphere since the Chernobyl nuclear accident. Although 80% of the radionuclides from this accidental release were transported over the Pacific Ocean, 20% were deposited over Japanese coastal catchments that are subject to frequent typhoons. Among the radioisotopes released during the FDNPP accident, radiocesium ((134)Cs and (137)Cs) is considered the most serious current and future health risk for the local population. The goal of this review is to synthesize research relevant to the transfer of FDNPP derived radiocesium from hillslopes to the Pacific Ocean. After radiocesium fallout deposition on vegetation and soils, the contamination may remain stored in forest canopies, in vegetative litter on the ground, or in the soil. Once radiocesium contacts soil, it is quickly and almost irreversibly bound to fine soil particles. The kinetic energy of raindrops instigates the displacement of soil particles, and their bound radiocesium, which may be mobilized and transported with overland flow. Soil erosion is one of the main processes transferring particle-bound radiocesium from hillslopes through rivers and streams, and ultimately to the Pacific Ocean. Accordingly this review will summarize results regarding the fundamental processes and dynamics that govern radiocesium transfer from hillslopes to the Pacific Ocean published in the literature within the first four years after the FDNPP accident. The majority of radiocesium is reported to be transported in the particulate fraction, attached to fine particles. The contribution of the dissolved fraction to radiocesium migration is only relevant in base flows and is hypothesized to decline over time. Owing to the hydro-meteorological context of the

  19. Radiocesium transfer from hillslopes to the Pacific Ocean after the Fukushima Nuclear Power Plant accident: A review.

    PubMed

    Evrard, Olivier; Laceby, J Patrick; Lepage, Hugo; Onda, Yuichi; Cerdan, Olivier; Ayrault, Sophie

    2015-10-01

    The devastating tsunami triggered by the Great East Japan Earthquake on March 11, 2011 inundated the Fukushima Dai-ichi Nuclear Power Plant (FDNPP) resulting in a loss of cooling and a series of explosions releasing the largest quantity of radioactive material into the atmosphere since the Chernobyl nuclear accident. Although 80% of the radionuclides from this accidental release were transported over the Pacific Ocean, 20% were deposited over Japanese coastal catchments that are subject to frequent typhoons. Among the radioisotopes released during the FDNPP accident, radiocesium ((134)Cs and (137)Cs) is considered the most serious current and future health risk for the local population. The goal of this review is to synthesize research relevant to the transfer of FDNPP derived radiocesium from hillslopes to the Pacific Ocean. After radiocesium fallout deposition on vegetation and soils, the contamination may remain stored in forest canopies, in vegetative litter on the ground, or in the soil. Once radiocesium contacts soil, it is quickly and almost irreversibly bound to fine soil particles. The kinetic energy of raindrops instigates the displacement of soil particles, and their bound radiocesium, which may be mobilized and transported with overland flow. Soil erosion is one of the main processes transferring particle-bound radiocesium from hillslopes through rivers and streams, and ultimately to the Pacific Ocean. Accordingly this review will summarize results regarding the fundamental processes and dynamics that govern radiocesium transfer from hillslopes to the Pacific Ocean published in the literature within the first four years after the FDNPP accident. The majority of radiocesium is reported to be transported in the particulate fraction, attached to fine particles. The contribution of the dissolved fraction to radiocesium migration is only relevant in base flows and is hypothesized to decline over time. Owing to the hydro-meteorological context of the

  20. DNA methylation patterns in tissues from mid-gestation bovine foetuses produced by somatic cell nuclear transfer show subtle abnormalities in nuclear reprogramming

    PubMed Central

    2010-01-01

    Background Cloning of cattle by somatic cell nuclear transfer (SCNT) is associated with a high incidence of pregnancy failure characterized by abnormal placental and foetal development. These abnormalities are thought to be due, in part, to incomplete re-setting of the epigenetic state of DNA in the donor somatic cell nucleus to a state that is capable of driving embryonic and foetal development to completion. Here, we tested the hypothesis that DNA methylation patterns were not appropriately established during nuclear reprogramming following SCNT. A panel of imprinted, non-imprinted genes and satellite repeat sequences was examined in tissues collected from viable and failing mid-gestation SCNT foetuses and compared with similar tissues from gestation-matched normal foetuses generated by artificial insemination (AI). Results Most of the genomic regions examined in tissues from viable and failing SCNT foetuses had DNA methylation patterns similar to those in comparable tissues from AI controls. However, statistically significant differences were found between SCNT and AI at specific CpG sites in some regions of the genome, particularly those associated with SNRPN and KCNQ1OT1, which tended to be hypomethylated in SCNT tissues. There was a high degree of variation between individuals in methylation levels at almost every CpG site in these two regions, even in AI controls. In other genomic regions, methylation levels at specific CpG sites were tightly controlled with little variation between individuals. Only one site (HAND1) showed a tissue-specific pattern of DNA methylation. Overall, DNA methylation patterns in tissues of failing foetuses were similar to apparently viable SCNT foetuses, although there were individuals showing extreme deviant patterns. Conclusion These results show that SCNT foetuses that had developed to mid-gestation had largely undergone nuclear reprogramming and that the epigenetic signature at this stage was not a good predictor of whether the

  1. Experimental study on the heat transfer characteristics of a nuclear reactor containment wall cooled by gravitationally falling water

    NASA Astrophysics Data System (ADS)

    Pasek, Ari D.; Umar, Efrison; Suwono, Aryadi; Manalu, Reinhard E. E.

    2012-06-01

    Gravitationally falling water cooling is one of mechanism utilized by a modern nuclear Pressurized Water Reactor (PWR) for its Passive Containment Cooling System (PCCS). Since the cooling is closely related to the safety, water film cooling characteristics of the PCCS should be studied. This paper deals with the experimental study of laminar water film cooling on the containment model wall. The influences of water mass flow rate and wall heat rate on the heat transfer characteristic were studied. This research was started with design and assembly of a containment model equipped with the water cooling system, and calibration of all measurement devices. The containment model is a scaled down model of AP 1000 reactor. Below the containment steam is generated using electrical heaters. The steam heated the containment wall, and then the temperatures of the wall in several positions were measure transiently using thermocouples and data acquisition. The containment was then cooled by falling water sprayed from the top of the containment. The experiments were done for various wall heat rate and cooling water flow rate. The objective of the research is to find the temperature profile along the wall before and after the water cooling applied, prediction of the water film characteristic such as means velocity, thickness and their influence to the heat transfer coefficient. The result of the experiments shows that the wall temperatures significantly drop after being sprayed with water. The thickness of water film increases with increasing water flow rate and remained constant with increasing wall heat rate. The heat transfer coefficient decreases as film mass flow rate increase due to the increases of the film thickness which causes the increasing of the thermal resistance. The heat transfer coefficient increases slightly as the wall heat rate increases. The experimental results were then compared with previous theoretical studied.

  2. Diffusion of the Interspecies Electron Carriers H2 and Formate in Methanogenic Ecosystems and Its Implications in the Measurement of Km for H2 or Formate Uptake

    PubMed Central

    Boone, David R.; Johnson, Richard L.; Liu, Yitai

    1989-01-01

    We calculated the potential H2 and formate diffusion between microbes and found that at H2 concentrations commonly found in nature, H2 could not diffuse rapidly enough to dispersed methanogenic cells to account for the rate of methane synthesis but formate could. Our calculations were based on individual organisms dispersed in the medium, as supported by microscopic observations of butyrate-degrading cocultures. We isolated an axenic culture of Syntrophomonas wolfei and cultivated it on butyrate in syntrophic coculture with Methanobacterium formicicum; during growth the H2 concentration was 63 nM (10.6 Pa). S. wolfei contained formate dehydrogenase activity (as does M. formicicum), which would allow interspecies formate transfer in that coculture. Thus, interspecies formate transfer may be the predominant mechanism of syntrophy. Our diffusion calculations also indicated that H2 concentration at the cell surface of H2-consuming organisms was low but increased to approximately the bulk-fluid concentration at a distance of about 10 μm from the surface. Thus, routine estimation of kinetic parameters would greatly overestimate the Km for H2 or formate. PMID:16347966

  3. High Power Nuclear Electric Propulsion (NEP) for Cargo and Propellant Transfer Missions in Cislunar Space

    NASA Technical Reports Server (NTRS)

    Falck, Robert D.; Borowski, Stanley K.

    2003-01-01

    The performance of Nuclear Electric Propulsion (NEP) in transporting cargo and propellant from Low Earth Orbit (LEO) to the first Earth-Moon Lagrange point (EML1) is examined. The baseline NEP vehicle utilizes a fission reactor system with Brayton power conversion for electric power generation to power multiple liquid hydrogen magnetoplasmadynamic (MPD) thrusters. Vehicle characteristics and performance levels are based on technology availability in a fifteen to twenty year timeframe. Results of numerical trajectory analyses are also provided.

  4. Sensitizing solid state nuclear magnetic resonance of dilute nuclei by spin-diffusion assisted polarization transfers.

    PubMed

    Lupulescu, Adonis; Frydman, Lucio

    2011-10-01

    Recent years have witnessed efforts geared at increasing the sensitivity of NMR experiments, by relying on the suitable tailoring and exploitation of relaxation phenomena. These efforts have included the use of paramagnetic agents, enhanced (1)H-(1)H incoherent and coherent transfers processes in 2D liquid state spectroscopy, and homonuclear (13)C-(13)C spin diffusion effects in labeled solids. The present study examines some of the opportunities that could open when exploiting spontaneous (1)H-(1)H spin-diffusion processes, to enhance relaxation and to improve the sensitivity of dilute nuclei in solid state NMR measurements. It is shown that polarization transfer experiments executed under sufficiently fast magic-angle-spinning conditions, enable a selective polarization of the dilute low-γ spins by their immediate neighboring protons. Repolarization of the latter can then occur during the time involved in monitoring the signal emitted by the low-γ nuclei. The basic features involved in the resulting approach, and its potential to improve the effective sensitivity of solid state NMR measurements on dilute nuclei, are analyzed. Experimental tests witness the advantages that could reside from utilizing this kind of approach over conventional cross-polarization processes. These measurements also highlight a number of limitations that will have to be overcome for transforming selective polarization transfers of this kind into analytical methods of choice.

  5. Core gas dynamics and heat transfer analyses of the COMET nuclear thermal rocket engine

    SciTech Connect

    Moore, R.L.; Bennett, R.G. )

    1992-01-01

    There has been a renewed interest in nuclear space propulsion for both the Space Exploration Initiative and military missions. The conical multiple element thruster (COMET) nuclear-thermal rocket engine discussed in this paper is one of several designs that have been proposed as an alternative to the pellet-bed reactor, which the US Air Force has been developing for various military space missions. COMET was designed to have a thrust of 75,000 lb[sub f] and a specific impulse of 900 to 1000 s. The nuclear reactor subsystem consists of 37 fuel assemblies. The core geometry is conical, as is the shape of each individual assembly. A typical fuel assembly, consists of a conical hot flow channel surrounded by a conical fueled region. The fuel region contains [approx] 500 conical fuel wafers, which are each 0.5 mm thick. Each wafer is separated from the adjacent wafers by a 0.1-mm coolant channel, resulting in a fueled region porosity of 20%. Surrounding the fuel region is an annular inlet flow channel. The hydrogen propellant enters the inlet flow channel at the upper end of the fuel assembly at a temperature of [approx] 300 K. As the hydrogen flows down the inlet channel, it is diverted into the narrow fuel coolant channels and heated to [approx] 3150 K as it exits the fuel. The work reported here involved modeling the flow through the conical coolant channels and the hot flow duct of COMET fuel assemblies.

  6. Inter-species entanglement of Bose-Bose mixtures trapped in optical lattices

    NASA Astrophysics Data System (ADS)

    (王 巍, Wei Wang; Penna, Vittorio; Capogrosso-Sansone, Barbara

    2016-06-01

    In the present work we discuss inter-species entanglement in Bose-Bose mixtures trapped in optical lattices. This work is motivated by the observation that, in the presence of a second component, the MI lobe shifts differently on the hole- and particle-side with respect to the Mott lobe of the single species system (Guglielmino et al 2010 Phys. Rev. A 82 021601; Capogrosso-Sansone et al 2011 Laser Phys. 21 1443). We use perturbation theory, formulated in a Hilbert space decomposed by means of lattice symmetries, in order to show that the nonuniform shift of the Mott lobe is a manifestation of inter-species entanglement which differs in the lowest excited states to remove and add a particle. Our results indicate that inter-species entanglement in mixtures can provide a new perspective in understanding quantum phase transitions. To validate our approach, we compare our results from perturbation theory with quantum Monte Carlo simulations.

  7. Induced pluripotent stem-induced cells show better constitutive heterochromatin remodeling and developmental potential after nuclear transfer than their parental cells.

    PubMed

    Liu, Zichuan; Wan, Haifeng; Wang, Eryao; Zhao, Xiaoyang; Ding, Chenhui; Zhou, Shuya; Li, Tianda; Shuai, Ling; Feng, Chunjing; Yu, Yang; Zhou, Qi; Beaujean, Nathalie

    2012-11-01

    Recently, reprogramming of somatic cells from a differentiated to pluripotent state by overexpression of specific external transcription factors has been accomplished. It has been widely speculated that an undifferentiated state may make donor cells more efficient for nuclear transfer. To test this hypothesis, we derived induced pluripotent stem cells (iPS cells) from several somatic cell lines: mouse embryonic fibroblast (MEF), adult tail tip fibroblast (TTF), and brain neural stem cells (NSCs). Three dimensional (3D)-fluorescent in situ hybridization (FISH) and quantitative-FISH (Q-FISH) were then used to evaluate constitutive (pericentric and telomeric) heterochromatin organization in these iPS cells and in their parental differentiated cells. Here, we show that important nuclear remodeling and telomeres rejuvenation occur in these iPS cells regardless of their parental origin. When we used these cells as donors for nuclear transfer, we produced live-born cloned mice at much higher rates with the iPS-induced cells than with the parental cell lines. Interestingly, we noticed that developmental potential after nuclear transfer could be correlated with telomere length of the donor cells. Altogether, our findings suggest that constitutive heterochromatin organization from differentiated somatic cells can be reprogrammed to the pluripotent state by induction of iPS cells, which in turn support nuclear transfer procedure quite efficiently. PMID:22657835

  8. Corrigendum to "Coupled thermochemical, isotopic evolution and heat transfer simulations in highly irradiated UO2 nuclear fuel"

    NASA Astrophysics Data System (ADS)

    Piro, M. H. A.; Banfield, J.; Clarno, K.; Simunovic, S.; Besmann, T. M.; Lewis, B. J.; Thompson, W. T.

    2016-09-01

    Figs. 7-9 in "Coupled thermochemical, isotopic evolution and heat transfer simulations in highly irradiated UO2 nuclear fuel" [1] have a consistent error corresponding to the relative proportions of iodine. Reported concentrations of iodine in the original manuscript are approximately ten times higher than expected, and are comparable in atomic proportions to cesium. One would expect that the amount of cesium would be about one order of magnitude greater than iodine based on the difference in fission yields of 235U and 239Pu. A practical consequence of this error would affect the predicted quantity and chemical composition of iodine on the fuel surface, which is related to iodine-induced stress corrosion cracking [2].

  9. Generation of hypoxanthine phosphoribosyltransferase gene knockout rabbits by homologous recombination and gene trapping through somatic cell nuclear transfer.

    PubMed

    Yin, Mingru; Jiang, Weihua; Fang, Zhenfu; Kong, Pengcheng; Xing, Fengying; Li, Yao; Chen, Xuejin; Li, Shangang

    2015-11-02

    The rabbit is a common animal model that has been employed in studies on various human disorders, and the generation of genetically modified rabbit lines is highly desirable. Female rabbits have been successfully cloned from cumulus cells, and the somatic cell nuclear transfer (SCNT) technology is well established. The present study generated hypoxanthine phosphoribosyltransferase (HPRT) gene knockout rabbits using recombinant adeno-associated virus-mediated homologous recombination and SCNT. Gene trap strategies were employed to enhance the gene targeting rates. The male and female gene knockout fibroblast cell lines were derived by different strategies. When male HPRT knockout cells were used for SCNT, no live rabbits were obtained. However, when female HPRT(+/-) cells were used for SCNT, live, healthy rabbits were generated. The cloned HPRT(+/-) rabbits were fertile at maturity. We demonstrate a new technique to produce gene-targeted rabbits. This approach may also be used in the genetic manipulation of different genes or in other species.

  10. Bovine somatic cell nuclear transfer using recipient oocytes recovered by ovum pick-up: effect of maternal lineage of oocyte donors.

    PubMed

    Brüggerhoff, Katja; Zakhartchenko, Valeri; Wenigerkind, Hendrik; Reichenbach, Horst-Dieter; Prelle, Katja; Schernthaner, Wolfgang; Alberio, Ramiro; Küchenhoff, Helmut; Stojkovic, Miodrag; Brem, Gottfried; Hiendleder, Stefan; Wolf, Eckhard

    2002-02-01

    The efficiency of bovine nuclear transfer using recipient oocytes recovered by ultrasound-guided follicle aspiration (ovum pick-up [OPU]) was investigated. Oocyte donors were selected from 2 distinct maternal lineages (A and B) differing in 11 nucleotide positions of the mitochondrial DNA control region. A total of 1342 cumulus-oocyte complexes (COCs) were recovered. The numbers of total COCs and class I/II COCs recovered from donors of lineage A were higher (P < 0.001) than those obtained from lineage B. Follicle aspiration once per week yielded a higher (P < 0.001) total number of COCs per session than aspiration twice per week, whereas the reproduction status of donors (heifer vs. cow) had no effect on OPU results. Of the 1342 oocytes recovered, 733 (55%) were successfully matured in vitro and used for nuclear transfer. Fusion was achieved in 550 (75%) karyoplast-cytoplast complexes (KCCs), resulting in 277 (50%) cleaved embryos on Day 3. On Day 7 of culture, 84 transferable embryos (15% based on fused KCCs) were obtained. After 38 transfers (10 single, 22 double, and 6 triple transfers), 9 recipients (8 double and 1 triple transfer) were diagnosed as pregnant on Day 28, corresponding to a pregnancy rate of 24%. The proportion of transferable embryos on Day 7 was significantly (P < 0.05) influenced by maternal lineage of oocyte donors and by the frequency of follicle aspiration. Our study demonstrates the feasibility of generating nuclear transfer embryos with defined cytoplasmic background. These will be valuable tools to experimentally dissect the effects of nuclear and cytoplasmic components on embryonic, fetal, and postnatal development.

  11. Improvement of mouse cloning using nuclear transfer-derived embryonic stem cells and/or histone deacetylase inhibitor.

    PubMed

    Wakayama, Sayaka; Wakayama, Teruhiko

    2010-01-01

    Nuclear transfer-derived ES (ntES) cell lines can be established from somatic cell nuclei with a relatively high success rate. Although ntES cells have been shown to be equivalent to ES cells, there are ethical objections concerning human cells, such as the use of fresh oocyte donation from young healthy woman. In contrast, the use of induced pluripotent stem (iPS) cells for cloning poses few ethical problems and is a relatively easy technique compared with nuclear transfer. Therefore, although there are several reports proposing the use of ntES cells as a model of regenerative medicine, the use of these cells in preliminary medical research is waning. However, in theory, 5 to 10 donor cells can establish one ntES cell line and, once established, these cells will propagate indefinitely. These cells can be used to generate cloned animals from ntES cell lines using a second round of NT. Even in infertile and "unclonable" strains of mice, we can generate offspring from somatic cells by combining cloning with ntES technology. Moreover, cloned offspring can be generated potentially even from the nuclei of dead bodies or freeze-dried cells via ntES cells, such as from an extinct frozen animal. Currently, only the ntES technology is available for this purpose, because all other techniques, including iPS cell derivation, require significant numbers of living donor cells. This review describes how to improve the efficiency of cloning, the establishment of clone-derived embryonic stem cells and further applications.

  12. In vitro development of porcine transgenic nuclear-transferred embryos derived from newborn Guangxi Bama mini-pig kidney fibroblasts.

    PubMed

    Liu, Hongbo; Lv, Peiru; Zhu, Xiangxing; Wang, Xianwei; Yang, Xiaogan; Zuo, Erwei; Lu, Yangqing; Lu, Shengsheng; Lu, Kehuan

    2014-10-01

    Porcine transgenic cloning has potential applications for improving production traits and for biomedical research purposes. To produce a transgenic clone, kidney fibroblasts from a newborn Guangxi Bama mini-pig were isolated, cultured, and then transfected with red and green fluorescent protein genes using lipofectamine for nuclear transfer. The results of the present study show that the kidney fibroblasts exhibited excellent proliferative capacity and clone-like morphology, and were adequate for generation of somatic cell nuclear transfer (SCNT)-derived embryos, which was confirmed by their cleavage activity and blastocyst formation rate of 70.3% and 7.9%, respectively. Cells transfected with red fluorescent protein genes could be passed more than 35 times. Transgenic embryos cloned with fluorescent or blind enucleation methods were not significantly different with respect to cleavage rates (92.5% vs. 86.8%, p > 0.05) and blastocyst-morula rates (26.9% vs. 34.0%, p > 0.05), but were significantly different with respect to blastocyst rates (3.0% vs. 13.2%, p < 0.05). Cleavage (75.3%, 78.5% vs. 78.0%, p > 0.05), blastocyst (14.1%, 16.1% vs. 23.1%, p > 0.05) and morula/blastocyst rates (43.5%, 47.0% vs. 57.6%, p > 0.05) were not significantly different between the groups of transgenic cloned embryos, cloned embryos, and parthenogenetic embryos. This indicates that long-time screening by G418 caused no significant damage to kidney fibroblasts. Thus, kidney fibroblasts represent a promising new source for transgenic SCNT, and this work lays the foundation for the production of genetically transformed cloned Guangxi Bama mini-pigs. PMID:24879084

  13. Photoprotection and triplet energy transfer in higher plants: the role of electronic and nuclear fluctuations.

    PubMed

    Cupellini, Lorenzo; Jurinovich, Sandro; Prandi, Ingrid G; Caprasecca, Stefano; Mennucci, Benedetta

    2016-04-28

    Photosynthetic organisms employ several photoprotection strategies to avoid damage due to the excess energy in high light conditions. Among these, quenching of triplet chlorophylls by neighboring carotenoids (Cars) is fundamental in preventing the formation of singlet oxygen. Cars are able to accept the triplets from chlorophylls by triplet energy transfer (TET). We have here studied TET rates in CP29, a minor light-harvesting complex (LHC) of the Photosystem II in plants. A fully atomistic strategy combining classical molecular dynamics of the LHC in its natural environment with a hybrid time-dependent density functional theory/polarizable MM description of the TET is used. We find that the structural fluctuations of the pigment-protein complex can largely enhance the transfer rates with respect to those predicted using the crystal structure, reducing the triplet quenching times in the subnanosecond scale. These findings add a new perspective for the interpretation of the photoprotection function and its relation with structural motions of the LHC.

  14. A Multi-Dimensional Heat Transfer Model of a Tie-Tube and Hexagonal Fuel Element for Nuclear Thermal Propulsion

    NASA Technical Reports Server (NTRS)

    Gomez, C. F.; Mireles, O. R.; Stewart, E.

    2016-01-01

    The Space Capable Cryogenic Thermal Engine (SCCTE) effort considers a nuclear thermal rocket design based around a Low-Enriched Uranium (LEU) design fission reactor. The reactor core is comprised of bundled hexagonal fuel elements that directly heat hydrogen for expansion in a thrust chamber and hexagonal tie-tubes that house zirconium hydride moderator mass for the purpose of thermalizing fast neutrons resulting from fission events. Created 3D steady state Hex fuel rod model with 1D flow channels. Hand Calculation were used to set up initial conditions for fluid flow. The Hex Fuel rod uses 1D flow paths to model the channels using empirical correlations for heat transfer in a pipe. Created a 2-D axisymmetric transient to steady state model using the CFD turbulent flow and Heat Transfer module in COMSOL. This model was developed to find and understand the hydrogen flow that might effect the thermal gradients axially and at the end of the tie tube where the flow turns and enters an annulus. The Hex fuel rod and Tie tube models were made based on requirements given to us by CSNR and the SCCTE team. The models helped simplify and understand the physics and assumptions. Using pipe correlations reduced the complexity of the 3-D fuel rod model and is numerically more stable and computationally more time-efficient compared to the CFD approach. The 2-D axisymmetric tie tube model can be used as a reference "Virtual test model" for comparing and improving 3-D Models.

  15. Production of Bovine Embryos and Calves Cloned by Nuclear Transfer Using Mesenchymal Stem Cells from Amniotic Fluid and Adipose Tissue.

    PubMed

    da Silva, Carolina Gonzales; Martins, Carlos Frederico; Cardoso, Tereza Cristina; da Cunha, Elisa Ribeiro; Bessler, Heidi Christina; Martins, George Henrique Lima; Pivato, Ivo; Báo, Sônia Nair

    2016-04-01

    The less differentiated the donor cells are used in nuclear transfer (NT), the more easily are they reprogrammed by the recipient cytoplasm. In this context, mesenchymal stem cells (MSCs) appear as an alternative to donor nuclei for NT. The amniotic fluid and adipose tissue are sources of MSCs that have not been tested for the production of cloned embryos in cattle. The objective of this study was to isolate, characterize, and use MSCs derived from amniotic fluid (MSC-AF) and adipose tissue (MSC-AT) to produce cloned calves. Isolation of MSC-AF was performed using in vivo ultrasound-guided transvaginal amniocentesis, and MSC-AT were isolated by explant culture. Cellular phenotypic and genotypic characterization by flow cytometry, immunohistochemistry, and RT-PCR were performed, as well as induction in different cell lineages. The NT was performed using MSC-AF and MSC-AT as nuclear donors. The mesenchymal markers of MSC were expressed in bovine MSC-AF and MSC-AT cultures, as evidenced by flow cytometry, immunohistochemistry, and RT-PCR. When induced, these cells differentiated into osteocytes, chondrocytes, and adipocytes. Embryo production was similar between the cell types, and two calves were born. The calf from MSC-AT was born healthy, and this fact opens a new possibility of using this type of cell to produce cloned cattle by NT. PMID:27055630

  16. Rapamycin treatment during in vitro maturation of oocytes improves embryonic development after parthenogenesis and somatic cell nuclear transfer in pigs

    PubMed Central

    Lee, Joohyeong; Park, Jong-Im; Yun, Jung Im; Lee, Yongjin; Yong, Hwanyul; Lee, Seung Tae; Park, Choon-Keun; Hyun, Sang-Hwan; Lee, Geun-Shik

    2015-01-01

    This study was conducted to investigate the effects of rapamycin treatment during in vitro maturation (IVM) on oocyte maturation and embryonic development after parthenogenetic activation (PA) and somatic cell nuclear transfer (SCNT) in pigs. Morphologically good (MGCOCs) and poor oocytes (MPCOCs) were untreated or treated with 1 nM rapamycin during 0-22 h, 22-42 h, or 0-42 h of IVM. Rapamycin had no significant effects on nuclear maturation and blastocyst formation after PA of MGCOCs. Blastocyst formation after PA was significantly increased by rapamycin treatment during 22-42 h and 0-42 h (46.6% and 46.5%, respectively) relative to the control (33.3%) and 0-22 h groups (38.6%) in MPCOCs. In SCNT, blastocyst formation tended to increase in MPCOCs treated with rapamycin during 0-42 h of IVM relative to untreated oocytes (20.3% vs. 14.3%, 0.05 < p < 0.1), while no improvement was observed in MGCOCs. Gene expression analysis revealed that transcript abundance of Beclin 1 and microtubule-associated protein 1 light chain 3 mRNAs was significantly increased in MPCOCs by rapamycin relative to the control. Our results demonstrated that autophagy induction by rapamycin during IVM improved developmental competence of oocytes derived from MPCOCs. PMID:25797293

  17. [The radioecology of the grapevine. 1. The transfer of nuclear weapons fallout from the soil into wine].

    PubMed

    Hellmuth, K H; Wagner, A; Fischer, E

    1989-04-01

    In a field investigation (1983-1985) comprising eight places of the most important viticultural regions in the Federal Republic of Germany, the contents of the radionuclides tritium (3H), carbon-14 (14C), strontium-90 (90Sr), and cesium-137 (137Cs) in air, soils, leaves of the vine, grapes and wine were measured and site-specific transfer factors were calculated. Data concerning soil parameters, climatic conditions, cultivation and vinification were collected. The tritium content of all samples was 10 Bq/l water of combustion, independent of location and year. The specific activity of 14C in the atmosphere and in biological material was 0.22 Bq/g carbon, independent of site and year. 90Sr contents of soils fluctuated between 0.7 and 3.5 Bq/kg dry matter. The mean content of leaves was 2 Bq/kg fresh material, of grapes 0.035 Bq/kg and of wine 0.008 Bq/l. 137Cs content of soils fluctuated between 1.3 and 7.9 Bq/kg dry matter. The mean content of leaves was 0.098 Bq/kg fresh material, of grapes 0.021 Bq/kg and of wine 0.0085 Bq/l. A relation between transfer of radionuclides and soil parameters and between the contents of grapes and wine was not recognizable. While cultivar-specific differences were not observed in grapes, red wines contained somewhat more 137Cs than white wines. Transfer factors soil grapes were 0.027 for 90Sr and 0.0057 for 137Cs. Site-specific influences such as soil parameters, climate, cultivation, vinification and differences between years led to a relatively small fluctuation of values. An influence of the nuclear power station Neckarwestheim has not been found in any of the radionuclides.

  18. Impact of different sources of donor cells upon the nuclear transfer efficiency in Chinese indigenous Meishan pig.

    PubMed

    Hua, Z; Xu, G; Liu, X; Bi, Y; Xiao, H; Hua, W; Li, L; Zhang, L; Ren, H; Zheng, X

    2016-01-01

    Somatic cell nuclear transfer (SCNT) is currently the most efficient and precise method to generate genetically tailored pig models for both agricultural and biomedical research. However, its efficiency is crucially dependent on the source of nuclear donor cells. In this study, we compared the cloning efficiency by using three lines of donor cells that are derived from fetal, newborn and adult fibroblasts of Chinese indigenous Meishan pig. We showed that cleavage rate and blastocyst formation rate of the reconstructed embryos were not significantly different between the fetal (80.7% and 15.6%) and newborn ear skin (77.5% and 12.3%) fibroblast groups (p>0.05), but in both groups these indices were significantly higher than that found in the adult ear skin (70.5% and 8.8%; p<0.05). Reconstructed embryos derived from fetal, newborn, and adult ear skin fibroblasts were transferred to four surrogates, respectively. For the fetal, newborn, and adult ear skin fibroblasts, the number of pregnancies were two (50.0%), two (50.0%), and one (25.0%), respectively, and the number of deliveries were two (50.0%), one (25.0%), and zero (0.0%), respectively. Seven and two cloned piglets were obtained from the fetal and newborn ear skin fibroblasts respectively, while no piglets were obtained from the adult ear skin fibroblasts. Two cloned piglets from the newborn ear skin fibroblasts died shortly after birth because of neonatal asphyxia caused by dystocia. The birth weights of the piglets derived from the fetal and newborn ear skin fibroblasts were 1230.5 and 1310.0 g, respectively, which were statistically insignificant (p>0.05), but both were significantly higher than that of the control groups (p<0.05). Microsatellite analyses demonstrated that the genotypes of all cloned piglets were identical to their donor cells. Therefore, cloned pigs were successfully produced using two sources of donor cells isolated from the fetal and newborn ear skin fibroblasts of Meishan piglet, and

  19. Impact of different sources of donor cells upon the nuclear transfer efficiency in Chinese indigenous Meishan pig.

    PubMed

    Hua, Z; Xu, G; Liu, X; Bi, Y; Xiao, H; Hua, W; Li, L; Zhang, L; Ren, H; Zheng, X

    2016-01-01

    Somatic cell nuclear transfer (SCNT) is currently the most efficient and precise method to generate genetically tailored pig models for both agricultural and biomedical research. However, its efficiency is crucially dependent on the source of nuclear donor cells. In this study, we compared the cloning efficiency by using three lines of donor cells that are derived from fetal, newborn and adult fibroblasts of Chinese indigenous Meishan pig. We showed that cleavage rate and blastocyst formation rate of the reconstructed embryos were not significantly different between the fetal (80.7% and 15.6%) and newborn ear skin (77.5% and 12.3%) fibroblast groups (p>0.05), but in both groups these indices were significantly higher than that found in the adult ear skin (70.5% and 8.8%; p<0.05). Reconstructed embryos derived from fetal, newborn, and adult ear skin fibroblasts were transferred to four surrogates, respectively. For the fetal, newborn, and adult ear skin fibroblasts, the number of pregnancies were two (50.0%), two (50.0%), and one (25.0%), respectively, and the number of deliveries were two (50.0%), one (25.0%), and zero (0.0%), respectively. Seven and two cloned piglets were obtained from the fetal and newborn ear skin fibroblasts respectively, while no piglets were obtained from the adult ear skin fibroblasts. Two cloned piglets from the newborn ear skin fibroblasts died shortly after birth because of neonatal asphyxia caused by dystocia. The birth weights of the piglets derived from the fetal and newborn ear skin fibroblasts were 1230.5 and 1310.0 g, respectively, which were statistically insignificant (p>0.05), but both were significantly higher than that of the control groups (p<0.05). Microsatellite analyses demonstrated that the genotypes of all cloned piglets were identical to their donor cells. Therefore, cloned pigs were successfully produced using two sources of donor cells isolated from the fetal and newborn ear skin fibroblasts of Meishan piglet, and

  20. Nuclear donor cell lines considerably influence cloning efficiency and the incidence of large offspring syndrome in bovine somatic cell nuclear transfer.

    PubMed

    Liu, J; Wang, Y; Su, J; Luo, Y; Quan, F; Zhang, Y

    2013-08-01

    Total five ear skin fibroblast lines (named F1, F2, F3, F4 and F5) from different newborn Holstein cows have been used as nuclear donor cells for producing cloned cows by somatic cell nuclear transfer (SCNT). The effects of these cell lines on both in vitro and in vivo developmental rates of cloned embryos, post-natal survivability and incidence of large offspring syndrome (LOS) were examined in this study. We found that the different cell lines possessed the same capacity to support pre-implantation development of cloned embryos, the cleavage and blastocyst formation rates ranged from 80.2 ± 0.9 to 84.5 ± 2.5% and 28.5 ± 0.9 to 33.3 ± 1.4%, respectively. However, their capacities to support the in vivo development of SCNT embryos showed significant differences (p < 0.05). The pregnancy rates at 90 and 240 day were significantly lower in groups F2 (4.9% and 3.3%) and F3 (5.4% and 5.4%) compared to groups F1 (23.3% and 16.3%), F4 (25.7% and 18.6%) and F5 (25.9% and 19.8%) (p < 0.05). The cloning efficiency was significantly higher in group F5 than those in group F1, F2, F3 and F4 (9.3% vs 4.1%, 1.2%, 2.0% and 5.0%, respectively, p < 0.05). Moreover, large offspring syndrome (LOS) incidence in group F5 was significantly lower than those in other groups (p < 0.05). All cloned offspring from cell line F1, F2, F3 and F4 showed LOS and gestation length delay, while all cloned offspring from F5 showed normal birthweight and gestation length. We concluded that the nuclear donor cell lines have significant impact on the in vivo development of cloned embryos and the incidence of LOS in cloned calves.

  1. Genetic evidence for hybridisation between Haemonchus contortus and Haemonchus placei in natural field populations and its implications for interspecies transmission of anthelmintic resistance.

    PubMed

    Chaudhry, Umer; Redman, Elizabeth M; Abbas, Muhammad; Muthusamy, Raman; Ashraf, Kamran; Gilleard, John S

    2015-02-01

    Genetic hybridisation between parasitic nematode species has potentially important consequences. It could lead to the introgression of genes between species including those involved in pathogenicity, host specificity, transmission and drug resistance. It could also complicate diagnosis and control. However, there are few compelling examples of its occurrence in parasites in the field. Haemonchus contortus and Haemonchus placei are two closely related parasitic nematode species that predominantly infect small ruminants and cattle, respectively. They are capable of experimental hybridisation when adult worms of each species are transplanted into the same individual host. Given that co-infection occurs in both small ruminants and cattle, there is potential for hybridisation in the field. However, this has not been definitively demonstrated and its extent is unknown. We investigated the occurrence of co-infection and interspecies hybridisation in H. contortus and H. placei in field populations from small ruminants from Pakistan and southern India using a number of independent genetic markers. Haemonchus contortus and H. placei co-infections were common in Pakistan but not in southern India where H. placei appeared to be absent in small ruminant hosts. In the former region, a number of worms were identified that were heterozygous for fixed, species-specific rDNA internal transcribed spacer 2 (ITS-2) single nucleotide polymorphisms. Genotyping of these ITS-2 heterozygotes with an additional four nuclear markers conclusively demonstrated them to be F1 interspecies hybrids. Mitochondrial NADH dehydrogenase subunit 4 haplotype analysis demonstrated that four of the hybrid worms had a H. placei maternal parent and one had a H. contortus maternal parent showing that hybridisation could occur in either direction. Interestingly, one of these hybrids contained an H. contortus isotype-1 β-tubulin benzimidazole resistance allele, suggesting there is a potential for interspecies

  2. Transferred nuclear Overhauser effect analyses of membrane-bound enkephalin analogues by sup 1 H nuclear magnetic resonance: Correlation between activities and membrane-bound conformations

    SciTech Connect

    Milon, Alain; Miyazawa, Tatsuo; Higashijima, Tsutomu )

    1990-01-09

    Leu-enkephalin, (D-Ala{sup 2})Leu-enkephalin, and (D-Ala{sup 2})Leu-enkephalinamide (agonists) and (L-Ala{sup 2})Leu-enkephalin (inactive analogue) bind to lipid bilayer consisting of phosphatidylcholine and phosphatidylserine. The conformations that these compounds assume, once bound to perdeuterated phospholipid bilayer, have been shown to be unique, as shown by the transferred nuclear Overhauser effect (TRNOE) of {sup 1}H NMR spectroscopy. In addition, their location in the bilayer was analyzed by TRNOE in the presence of spin-labeled phospholipids. These analyses showed a clear relationship between the activity and the peptide-membrane interaction. The three active peptides, when bound to membranes, adopt the same conformation, characterized by a type II{prime} {beta}-turn around Gly{sup 3}-Phe and a {gamma}-turn around Gly{sup 2} (or D-Ala{sup 2}). The inactive analogue, (L-Ala{sup 2})Leu-enkephalin, displayed a completely different TRNOE pattern corresponding to a different conformation in the membrane-bound state. The tyrosine residue of the active compounds is not inserted into the interior of membrane, but it is inserted into the bilayer for the L-Ala{sup 2} analogue. According to these results, (L-Ala{sup 2})Leu-enkephalin may be explained to be inactive because the mode of binding to the membranes is different from that of active compounds.

  3. USE OF INTERSPECIES CORRELATION ESTIMATIONS TO PREDICT HC5'S BASED ON QSAR

    EPA Science Inventory

    Dyer, S.D., S. Belanger, J. Chaney, D. Versteeg and F. Mayer. In press. Use of Interspecies Correlation Estimations to predict HC5's Based on QSARs (Abstract). To be presented at the SETAC Europe 14th Annual Meeting: Environmental Science Solution: A Pan-European Perspective, 18-...

  4. WEB-BASED INTERSPECIES CORRELATION ESTIMATION (WEB-ICE) FOR ACUTE TOXICITY: USER MANUAL V2

    EPA Science Inventory

    Predictive toxicological models are integral to environmental risk Assessment where data for most species are limited. Web-based Interspecies Correlation Estimation (Web-ICE) models are least square regressions that predict acute toxicity (LC50/LD50) of a chemical to a species, ...

  5. USE OF INTERSPECIES CORRELATION ESTIMATIONS TO PREDICT HC5'S BASED ON MINIMAL DATA

    EPA Science Inventory

    Dyer, S., S. Belanger, J. Chaney, D. Versteeg and F. Mayer. In press. Use of Interspecies Correlation Estimations to Predict HC5's Based on Minimal Data (Abstract). To be presented at the SETAC Fourth World Congress, 14-18 November 2004, Portland, OR. 1 p. (ERL,GB R1013).

  6. Being Nature: Interspecies Articulation as a Species-Specific Practice of Relating to Environment

    ERIC Educational Resources Information Center

    Rautio, Pauliina

    2013-01-01

    Rather than categorically teaching us ways to be less anthropocentric, environmental education could be about educating us of the ways in which we already are nature as human animals. In this paper, one species-specific practice of human relating to environment--interspecies articulation--is argued as one way of being nature. Interspecies…

  7. Interspecies extrapolation based on the RepDose database--a probabilistic approach.

    PubMed

    Escher, Sylvia E; Batke, Monika; Hoffmann-Doerr, Simone; Messinger, Horst; Mangelsdorf, Inge

    2013-04-12

    Repeated dose toxicity studies from the RepDose database (DB) were used to determine interspecies differences for rats and mice. NOEL (no observed effect level) ratios based on systemic effects were investigated for three different types of exposure: inhalation, oral food/drinking water and oral gavage. Furthermore, NOEL ratios for local effects in inhalation studies were evaluated. On the basis of the NOEL ratio distributions, interspecies assessment factors (AF) are evaluated. All data sets were best described by a lognormal distribution. No difference was seen between inhalation and oral exposure for systemic effects. Rats and mice were on average equally sensitive at equipotent doses with geometric mean (GM) values of 1 and geometric standard deviation (GSD) values ranging from 2.30 to 3.08. The local AF based on inhalation exposure resulted in a similar distribution with GM values of 1 and GSD values between 2.53 and 2.70. Our analysis confirms former analyses on interspecies differences, including also dog and human data. Furthermore it supports the principle of allometric scaling according to caloric demand in the case that body doses are applied. In conclusion, an interspecies distribution animal/human with a GM equal to allometric scaling and a GSD of 2.5 was derived.

  8. Demonstration of the Web-based Interspecies Correlation Estimation (Web-ICE) modeling application

    EPA Science Inventory

    The Web-based Interspecies Correlation Estimation (Web-ICE) modeling application is available to the risk assessment community through a user-friendly internet platform (http://epa.gov/ceampubl/fchain/webice/). ICE models are log-linear least square regressions that predict acute...

  9. Investigation of Interspecies Interactions within Marine Micromonosporaceae Using an Improved Co-Culture Approach

    PubMed Central

    Adnani, Navid; Vazquez-Rivera, Emmanuel; Adibhatla, Srikar N.; Ellis, Gregory A.; Braun, Doug R.; Bugni, Tim S.

    2015-01-01

    With respect to bacterial natural products, a significant outcome of the genomic era was that the biosynthetic potential in many microorganisms surpassed the number of compounds isolated under standard laboratory growth conditions, particularly among certain members in the phylum Actinobacteria. Our group, as well as others, investigated interspecies interactions, via co-culture, as a technique to coax bacteria to produce novel natural products. While co-culture provides new opportunities, challenges exist and questions surrounding these methods remain unanswered. In marine bacteria, for example, how prevalent are interspecies interactions and how commonly do interactions result in novel natural products? In an attempt to begin to answer basic questions surrounding co-culture of marine microorganisms, we have tested both antibiotic activity-based and LC/MS-based methods to evaluate Micromonosporaceae secondary metabolite production in co-culture. Overall, our investigation of 65 Micromonosporaceae led to the identification of 12 Micromonosporaceae across three genera that produced unique metabolites in co-culture. Our results suggest that interspecies interactions were prevalent between marine Micromonosporaceae and marine mycolic acid-containing bacteria. Furthermore, our approach highlights a sensitive and rapid method for investigating interspecies interactions in search of novel antibiotics, secondary metabolites, and genes. PMID:26404321

  10. Development of Algal Interspecies Correlation Estimation Models for Chemical Hazard Assessment

    EPA Science Inventory

    Web-based Interspecies Correlation Estimation (ICE) is an application developed to predict the acute toxicity of a chemical from 1 species to another taxon. Web-ICE models use the acute toxicity value for a surrogate species to predict effect values for other species, thus potent...

  11. Augmenting Species Diversity in Water Quality Criteria Derivation using Interspecies Correlation Models

    EPA Science Inventory

    The specific requirements for taxa diversity of the 1985 guidelines have limited the number of ambient water quality criteria (AWQC) developed for aquatic life protection. The EPA developed the Web-based Interspecies Correlation Estimation (Web-ICE) tool to allow extrapolation of...

  12. Genetic analysis of Group A rotaviruses: evidence for interspecies transmission of rotavirus genes.

    PubMed

    Palombo, Enzo A

    2002-01-01

    Rotaviruses are the major cause of severe gastroenteritis in young children and animals. The rotavirus genome is composed of eleven segments of double-stranded RNA and can undergo genetic reassortment during mixed infections, leading to progeny viruses with novel or atypical phenotypes. There are numerous descriptions of rotavirus strains isolated from human and animals that share genetic and antigenic features of viruses from heterologous species. In many cases, genetic analysis by hybridization has clearly demonstrated the genetic relatedness of gene segments to those from viruses isolated from different species. Together with the observation that some virus strains appear to have been transmitted to a different species as a whole genome constellation, these data suggest that interspecies transmission occurs naturally, albeit at low frequencies. Although interspecies transmission has not been documented directly, there is an increasing number of reports of atypical rotaviruses that are apparently derived from transmission between: humans, cats and dogs; humans and cattle; humans and pigs; pigs and cattle; and pigs and horses. Interspecies evolutionary relationships are supported by phylogenetic analysis of rotavirus genes from different species. The emergence of novel strains derived from interspecies transmission has implications for the design and implementation of successful human rotavirus vaccine strategies. PMID:11928984

  13. Generation of nuclear hybrids overcoming the natural barrier of incompatibility: transfer of nuclei from Lentinula edodes into protoplasts of Coriolus versicolor.

    PubMed

    Kim, C; Choi, E C; Kim, B K

    2000-02-01

    Heterokaryotic nuclear hybrids overcoming the natural barriers of incompatibility have been studied in basidiomycetes. To produce these nuclear hybrids between incompatible mushrooms, which have several potent pharmacological effects, nuclear transfer was performed between Lentinula edodes and Coriolus versicolor. Nuclei from serine auxotrophs of Lentinula edodes, LE207 (Ser-) were transferred into the protoplasts of arginine auxotrophs of Coriolus versicolor, CV17 (Arg-), using 30% polyethylene glycol 4000 in 10 mM CaCl2-glycine solution (pH 8.0). Nuclear transfer progenies were selected by nutritional complementation on minimal media supplemented with 0.6 M sucrose. The progenies were classified based on colony morphology to L. edodes-like, C. versicolor-like and non-parental type. Most of the progenies grew slower than either parent. The number of nuclei per cell was similar but the DNA content varied between progenies. The isozyme patterns of nuclear hybrids resembled either of the parent profiles or showed a mixed profile. PMID:10728662

  14. Effective donor cell fusion conditions for production of cloned dogs by somatic cell nuclear transfer.

    PubMed

    Park, JungEun; Oh, HyunJu; Hong, SoGun; Kim, MinJung; Kim, GeonA; Koo, OkJae; Kang, SungKeun; Jang, Goo; Lee, ByeongChun

    2011-03-01

    As shown by the birth of the first cloned dog 'Snuppy', a protocol to produce viable cloned dogs has been reported. In order to evaluate optimum fusion conditions for improving dog cloning efficiency, in vivo matured oocytes were reconstructed with adult somatic cells from a female Pekingese using different fusion conditions. Fusion with needle vs chamber methods, and with low vs high pulse strength was compared by evaluating fusion rate and in vivo development of canine cloned embryos. The fusion rates in the high voltage groups were significantly higher than in the low voltage groups regardless of fusion method (83.5 vs 66.1% for the needle fusion method, 67.4 vs 37.9% for the fusion chamber method). After embryo transfer, one each pregnancy was detected after using the needle fusion method with high and low voltage and in the chamber fusion method with high voltage, whereas no pregnancy was detected using the chamber method with low voltage. However, only the pregnancy from the needle fusion method with high voltage was maintained to term and one healthy puppy was delivered. The results of the present study demonstrated that two DC pulses of 3.8 to 4.0 kV/cm for 15 μsec using the needle fusion method were the most effective method for the production of cloned dogs under the conditions of this experiment.

  15. A magnetization-transfer nuclear magnetic resonance study of the folding of staphylococcal nuclease

    SciTech Connect

    Evans, P.A.; Kautz, R.A.; Fox, R.O.; Dobson, C.M. )

    1989-01-10

    The equilibrium between alternative folded states of a globular protein, staphylococcal nuclease, has been investigated by using {sup 1}H NMR. Magnetization-transfer experiments have revealed the existence of a related structural heterogeneity of the unfolded state, and quantitative analysis of a series of these experiments has permitted the kinetics of folding and interconversion of the different states to be explored. A model based on cis/trans isomerism at the peptide bond preceding Pro-117 has been developed to account for the results. This model, recently supported by a protein-engineering experiment has been used to interpret the kinetic data, providing insight into the nature of the folding processes. The predominance of the cis-proline form in the folded state is shown to derive from a large favorable enthalpy term resulting from more effective overall folding interactions. The kinetics of folding and isomerization are shown to occur on similar time scales, such that more than one pathway between two states may be significant. It has been possible, however, to compare the direct folding and unfolding rates within the cis- and trans-proline-containing populations, with results suggesting that the specific stabilization of the cis peptide bond is effective only at a late stage in the folding process.

  16. Hyperpolarization of 29Si by Resonant Nuclear Spin Transfer from Optically Hyperpolarized 31P Donors

    NASA Astrophysics Data System (ADS)

    Dluhy, Phillip; Salvail, Jeff; Saeedi, Kamyar; Thewalt, Mike; Simons, Stephanie

    2014-03-01

    Recent developments in nanomedicine have allowed nanoparticles of silicon containing hyperpolarized 29Si to be imaged in vivo using magnetic resonance imaging. The extremely long relaxation times and isotropy of the Si lattice make polarized 29Si isotopes ideal for these sorts of imaging methods. However, one of the major difficulties standing in the path of widespread adoption of these techniques is the slow rate at which the 29Si is hyperpolarized and the limited maximum hyperpolarization achievable. In this talk, I will describe an effective method for hyperpolarization of the 29Si isotopes using resonant optical pumping of the donor bound exciton transitions to polarize the 31P donor nuclei, and a choice of static magnetic field that conserves energy during spin flip flops between donor nuclear and 29Si spins to facilitate diffusion of this polarization. Using this method, we are able to polarize greater than 10% of the 29Si centers in 64 hours without seeing saturation of the 29Si polarization.

  17. Interspecies acetate transfer influences the extent of anaerobic benzoate degradation by syntrophic consortia

    SciTech Connect

    Warikoo, V.; McInerney, M.J.; Suflita, J.M.

    1997-03-01

    Benzoate degradation by an anaerobic, syntrophic bacterium, strain SB, in coculture with Desulfovibrio strain G-11 reached a threshold value which depended on the amount of acetate added, and ranged from about 2.5 to 29.9 {mu}M. Increasing acetate concentrations also uncompetitively inhibited benzoate degradation. The apparent V{sub max} and K{sub m} for benzoate degradation decreased with increasing acetate concentration, but the benzoate degradation capacity (V{sub max}/K{sub m}) of cell suspensions remained comparable. The addition of an acetate-using bacterium to cocultures after the threshold was reached resulted in the degradation of benzoate to below the detection limit. Mathematical simulations showed that the benzoate threshold was not predicted by the inhibitory effect of acetate on benzoate degradation kinetics. With nitrate instead of sulfate as the terminal electron acceptor, no benzoate threshold was observed in the presence of 20 mM acetate even though the degradation capacity was lower with nitrate than with sulfate. When strain SB was grown with a hydrogen-using partner that had a 5-fold lower hydrogen utilization capacity, a 5 to 9-fold lower the benzoate degradation capacity was observed compared to SB/G-11 cocultures. The Gibb`s free energy for benzoate degradation was less negative in cell suspensions with threshold compared to those without threshold. These studies showed that the threshold was not a function of the inhibition of benzoate degradation capacity by acetate, or the toxicity of the undissociated form of acetate. Rather a critical or minimal Gibb`s free energy may exist where thermodynamic constraints preclude further benzoate degradation.

  18. Interspecies Acetate Transfer Influences the Extent of Anaerobic Benzoate Degradation by Syntrophic Consortia

    PubMed Central

    Warikoo, V.; McInerney, M. J.; Robinson, J. A.; Suflita, J. M.

    1996-01-01

    Benzoate degradation by an anaerobic, syntrophic bacterium, strain SB, in coculture with Desulfovibrio sp. strain G-11 reached a threshold value which depended on the amount of acetate added and ranged from about 2.5 to 29.9 (mu)M. Increasing acetate concentrations also uncompetitively inhibited benzoate degradation. The apparent V(infmax) and apparent K(infm) for benzoate degradation decreased with increasing acetate concentration, but the benzoate degradation capacities (V(infmax)/K(infm)) of cell suspensions remained comparable. The addition of an acetate-using bacterium to cocultures after the threshold was reached resulted in the degradation of benzoate to below the detection limit. Mathematical simulations showed that the benzoate threshold was not predicted by the inhibitory effect of acetate on benzoate degradation kinetics. With nitrate instead of sulfate as the terminal electron acceptor, no benzoate threshold was observed in the presence of 20 mM acetate even though the kinetics of benzoate degradation were slower with nitrate rather than sulfate as the electron acceptor. When strain SB was grown with Desulfovibrio sp. strain DG2 that had a fourfold-lower V(infmax) for hydrogen use than strain G-11, the V(infmax) for benzoate degradation was 37-fold lower than that of strain SB-G-11 cocultures. The Gibb's free energy for benzoate degradation was less negative in cell suspensions with a threshold than in suspensions without a threshold. These studies showed that the threshold was not a function of the inhibition of benzoate degradation by acetate or the toxicity of the undissociated form of acetate. Rather, a critical or minimal Gibb's free energy may exist where thermodynamic constraints preclude further benzoate degradation. PMID:16535215

  19. Heat Transfer Salts for Nuclear Reactor Systems - Chemistry Control, Corrosion Mitigation, and Modeling

    SciTech Connect

    Anderson, Mark; Sridharan, Kumar; Morgan, Dane; Peterson, Per; Calderoni, Pattrick; Scheele, Randall; Casekka, Andrew; McNamara, Bruce

    2015-01-22

    The concept of a molten salt reactor has existed for nearly sixty years. Previously all work was done during a large collaborative effort at Oak Ridge National Laboratory, culminating in a research reactor which operated for 15,000 hours without major error. This technical success has garnished interest in modern, high temperature, reactor schemes. Research using molten fluoride salts for nuclear applications requires a steady supply of high grade molten salts. There is no bulk supplier of research grade fluoride salts in the world, so a facility which could provide all the salt needed for testing at the University of Wisconsin had to be produced. Two salt purification devices were made for this purpose, a large scale purifier, and a small scale purifier, each designed to clean the salts from impurities and reduce their corrosion potential. As of now, the small scale has performed with flibe salt, hydrogen, and hydrogen fluoride, yielding clean salt. This salt is currently being used in corrosion testing facilities at the Massachusetts Institute of Technology and the University of Wisconsin. Working with the beryllium based salts requires extensive safety measures and health monitoring to prevent the development of acute or chronic beryllium disease, two pulmonary diseases created by an allergic reaction to beryllium in the lungs. Extensive health monitoring, engineering controls, and environment monitoring had to be set up with the University of Wisconsin department of Environment, Health and Safety. The hydrogen fluoride required for purification was also an extreme health hazard requiring thoughtful planning and execution. These dangers have made research a slow and tedious process. Simple processes, such as chemical handling and clean-up, can take large amounts of ingenuity and time. Other work has complemented the experimental research at Wisconsin to advance high temperature reactor goals. Modeling work has been performed in house to re

  20. Significant improvement in cloning efficiency of an inbred miniature pig by histone deacetylase inhibitor treatment after somatic cell nuclear transfer.

    PubMed

    Zhao, Jianguo; Ross, Jason W; Hao, Yanhong; Spate, Lee D; Walters, Eric M; Samuel, Melissa S; Rieke, August; Murphy, Clifton N; Prather, Randall S

    2009-09-01

    The National Institutes of Health (NIH) miniature pig was developed specifically for xenotransplantation and has been extensively used as a large-animal model in many other biomedical experiments. However, the cloning efficiency of this pig is very low (<0.2%), and this has been an obstacle to the promising application of these inbred swine genetics for biomedical research. It has been demonstrated that increased histone acetylation in somatic cell nuclear transfer (SCNT) embryos, by applying a histone deacetylase (HDAC) inhibitor such as trichostatin A (TSA), significantly enhances the developmental competence in several species. However, some researchers also reported that TSA treatment had various detrimental effects on the in vitro and in vivo development of the SCNT embryos. Herein, we report that treatment with 500 nM 6-(1,3-dioxo-1H, 3H-benzo[de]isoquinolin-2-yl)-hexanoic acid hydroxyamide (termed scriptaid), a novel HDAC inhibitor, significantly enhanced the development of SCNT embryos to the blastocyst stage when NIH inbred fetal fibroblast cells (FFCs) were used as donors compared with the untreated group (21% vs. 9%, P < 0.05). Scriptaid treatment resulted in eight pregnancies from 10 embryo transfers (ETs) and 14 healthy NIH miniature pigs from eight litters, while no viable piglets (only three mummies) were obtained from nine ETs in the untreated group. Thus, scriptaid dramatically increased the cloning efficiency when using inbred genetics from 0.0% to 1.3%. In contrast, scriptaid treatment decreased the blastocyst rate in in vitro fertilization embryos (from 37% to 26%, P < 0.05). In conclusion, the extremely low cloning efficiency in the NIH miniature pig may be caused by its inbred genetic background and can be improved by alteration of genomic histone acetylation patterns.

  1. Effect of radiocesium transfer on ambient dose rate in forest environments affected by the Fukushima Nuclear Power Plant accident

    NASA Astrophysics Data System (ADS)

    Kato, H.

    2015-12-01

    We investigated the transfer of canopy-intercepted radiocesium to the forest floor during 3 years following the Fukushima Daiichi Nuclear Power Plant accident. The cesium-137 (Cs-137) contents in throughfall, stemflow, and litterfall were monitored in two coniferous stands (plantation of Japanese cedar) and a deciduous broad-leaved forest stand (Japanese oak with red pine). We also measured the ambient dose rate (ADR) at different heights in the forest using a survey meter and a portable Ge gamma-ray detector. Total Cs-137 deposition flux from the canopy to forest floor for the mature cedar, young cedar, and the mixed broad-leaved stands were 166 kBq/m2, 174 kBq/m2, and 60 kBq/m2, respectively. These values correspond to 38%, 40% and 13% of total atmospheric input after the accident. The ambient dose rate in forest exhibited height dependency and its vertical distribution varied with forest type and stand age. The ambient dose rate showed an exponential decrease with time for all the forest sites, however the decreasing trend differed depending on the height of dose measurement and forest type. The ambient dose rate at the canopy (approx. 10 m-height) decreased faster than that expected from physical decay of the two radiocesium isotopes, whereas those at the forest floor varied between the three forest stands. The radiocesium deposition via throughfall seemed to increase ambient dose rate during the first 200 days after the accident, however there was no clear relationship between litterfall and ambient dose rate since 400 days after the accident. These data suggested that the ambient dose rate in forest environment varied both spatially and temporally reflecting the transfer of radiocesium from canopy to forest floor. However, further monitoring investigation and analysis are required to determine the effect of litterfall on long-term trend of ambient dose rate in forest environments.

  2. Post-fusion treatment with MG132 increases transcription factor expression in somatic cell nuclear transfer embryos in pigs.

    PubMed

    You, Jinyoung; Lee, Joohyeong; Kim, Jinyoung; Park, Junhong; Lee, Eunsong

    2010-02-01

    The objective of this study was to examine the effect of post-fusion treatment of somatic cell nuclear transfer (SCNT) oocytes with the proteasomal inhibitor MG132 on maturation promoting factor (MPF) activity, nuclear remodeling, embryonic development, and gene expression of cloned pig embryos. Immediately after electrofusion, SCNT oocytes were treated with MG132 and/or caffeine for 2 hr, vanadate for 0.5 hr, or vanadate for 0.5 hr followed by MG132 for 1.5 hr. Of the MG132 concentrations tested (0-5 microM), the 1 microM concentration showed a higher rate of blastocyst formation (25.9%) than 0 (14.2%), 0.5 (16.9%), and 5 microM (16.9%). Post-fusion treatment with MG132, caffeine, and both MG132 and caffeine improved blastocyst formation (22.1%, 21.4%, and 24.4%, respectively), whereas vanadate treatment inhibited blastocyst formation (6.5%) compared to the control (11.1%). When examined 2 hr after fusion and 1 hr after activation, MPF activity remained at a higher (P < 0.05) level in SCNT oocytes that were treated post-fusion with caffeine and/or MG132, but it was decreased by vanadate. The rate of oocytes showing premature chromosome condensation was not altered by MG132 but was decreased by vanadate treatment. In addition, formation of single pronuclei was increased by MG132 compared to control and vanadate treatment. MG132-treated embryos showed increased expression of POU5F1, DPPA2, DPPA3, DPPA5, and NDP52l1 genes compared to control embryos. Our results demonstrate that post-fusion treatment of SCNT oocytes with MG132 prevents MPF degradation and increases expression of transcription factors in SCNT embryos, which are necessary for normal development of SCNT embryos.

  3. Production of a healthy calf by somatic cell nuclear transfer without micromanipulators and carbon dioxide incubators using the Handmade Cloning (HMC) and the Submarine Incubation System (SIS).

    PubMed

    Vajta, Gábor; Bartels, Paul; Joubert, Jennifer; de la Rey, Morné; Treadwell, Robert; Callesen, Henrik

    2004-11-01

    The aim of this work was to investigate the minimum technical requirements for production of live offspring with somatic cell nuclear transfer. The experiment was performed in a field type laboratory without micromanipulators and carbon dioxide incubators. All long-term incubations were performed in the Submarine Incubation System (SIS) using various gas mixtures. The somatic cell culture was established from ear biopsy of a 9-year-old Holstein cow. Nuclear transfer was performed using the Handmade Cloning (HMC) technique. Zona-free oocytes were randomly bisected by hand with a disposable blade and a stereomicroscope. Cytoplast were selected using Hoechst staining and a fluorescent microscope. After a two-step fusion embryos were activated with calcium ionophore and dimethylaminopurine. Embryos were cultured in microwells (WOWs) in SOFaaci medium supplemented with 5% cattle serum. In two consecutive experiments, six blastocysts were produced from 52 reconstructed embryos. On Day 7, five blastocysts were transferred into synchronized recipients. All three recipients became pregnant but two pregnancies aborted at 6 and 7 months, respectively. A heifer calf weighing 27 kg was delivered at term by Caesarean section from the third pregnancy. The healthy 6-month-old heifer, the first cloned animal of Africa, is living evidence that nuclear transfer technology may be successfully used under basic laboratory conditions.

  4. Somatic cell nuclear transfer in buffalos: effect of the fusion and activation protocols and embryo culture system on preimplantation embryo development.

    PubMed

    Simon, Liz; Veerapandian, C; Balasubramanian, S; Subramanian, A

    2006-01-01

    The present study was conducted primarily to evaluate several factors that affect the nuclear transfer programme in water buffalos, in which relatively few studies have been performed. Embryos reconstructed with quiescent fetal fibroblasts and metaphase II cytoplasts were matured for 24 h, and activation was found to be comparatively better than in those matured for 30 h. A significantly higher proportion of embryos fused (52.0 +/- 1.9) and cleaved (51.2 +/- 1.7) when the couplets were fused 4-6 h before activation than when fused and activated simultaneously (46.5 +/- 1.6 and 44.5 +/- 2.0, respectively). Development of nuclear transfer embryos to the blastocyst stage (4.8 +/- 2.2) was supported by a commercially available sequential medium, and cleavage (76.5 +/- 2.8) was significantly higher in this medium compared with cleavage in TCM-199 with oviduct epithelial cell coculture (45.6 +/- 1.5) and synthetic oviduct fluid (21.8 +/- 6.6). Of the 16 cloned embryos transferred, none resulted in pregnancy. The present study demonstrates that optimal numbers of cloned buffalo blastocysts can be obtained from oocytes matured for 24 h, fused 3-4 h before activation and cultured in a commercially available sequential media (G1/G2), thus providing further information to enable successful nuclear transfer in buffalos.

  5. Improved cloning efficiency and developmental potential in bovine somatic cell nuclear transfer with the oosight imaging system.

    PubMed

    Kim, Eun Young; Park, Min Jee; Park, Hyo Young; Noh, Eun Ji; Noh, Eun Hyung; Park, Kyoung Sik; Lee, Jun Beom; Jeong, Chang Jin; Riu, Key Zung; Park, Se Pill

    2012-08-01

    In somatic cell nuclear transfer (SCNT) procedures, exquisite enucleation of the recipient oocyte is critical to cloning efficiency. The purpose of this study was to compare the effects of two enucleation systems, Hoechst staining and UV irradiation (hereafter, irradiation group) and Oosight imaging (hereafter, Oosight group), on the in vitro production of bovine SCNT embryos. In the Oosight group, the apoptotic index (2.8 ± 0.5 vs. 7.3 ± 1.2) was lower, and the fusion rate (75.6% vs. 62.9%), cleavage rate (78.0% vs. 63.7%), blastocyst rate (40.2% vs. 29.2%), and total cell number (128.3±4.8 vs. 112.2 ± 7.6) were higher than those in the irradiation group (all p<0.05). The overall efficiency after SCNT was twice as high in the Oosight group as that in the irradiation group (p<0.05). The relative mRNA expression levels of Oct4, Nanog, Interferon-tau, and Dnmt3A were higher and those of Caspase-3 and Hsp70 were lower in the Oosight group compared with the irradiation group (p<0.05). This is the first report to show the positive effect of the Oosight imaging system on molecular gene expression in the SCNT embryo. The Oosight imaging system may become the preferred choice for enucleation because it is less detrimental to the developmental potential of bovine SCNT embryos.

  6. Cloning adult farm animals: a review of the possibilities and problems associated with somatic cell nuclear transfer.

    PubMed

    Edwards, J L; Schrick, F N; McCracken, M D; van Amstel, S R; Hopkins, F M; Welborn, M G; Davies, C J

    2003-08-01

    In 1997, Wilmut et al. announced the birth of Dolly, the first ever clone of an adult animal. To date, adult sheep, goats, cattle, mice, pigs, cats and rabbits have been cloned using somatic cell nuclear transfer. The ultimate challenge of cloning procedures is to reprogram the somatic cell nucleus for development of the early embryo. The cell type of choice for reprogramming the somatic nucleus is an enucleated oocyte. Given that somatic cells are easily obtained from adult animals, cultured in the laboratory and then genetically modified, cloning procedures are ideal for introducing specific genetic modifications in farm animals. Genetic modification of farm animals provides a means of studying genes involved in a variety of biological systems and disease processes. Moreover, genetically modified farm animals have created a new form of 'pharming' whereby farm animals serve as bioreactors for production of pharmaceuticals or organ donors. A major limitation of cloning procedures is the extreme inefficiency for producing live offspring. Dolly was the only live offspring produced after 277 attempts. Similar inefficiencies for cloning adult animals of other species have been described by others. Many factors related to cloning procedures and culture environment contribute to the death of clones, both in the embryonic and fetal periods as well as during neonatal life. Extreme inefficiencies of this magnitude, along with the fact that death of the surrogate may occur, continue to raise great concerns with cloning humans.

  7. Stochastic anomaly of methylome but persistent SRY hypermethylation in disorder of sex development in canine somatic cell nuclear transfer

    PubMed Central

    Jeong, Young-Hee; Lu, Hanlin; Park, Chi-Hun; Li, Meiyan; Luo, Huijuan; Kim, Joung Joo; Liu, Siyang; Ko, Kyeong Hee; Huang, Shujia; Hwang, In Sung; Kang, Mi Na; Gong, Desheng; Park, Kang Bae; Choi, Eun Ji; Park, Jung Hyun; Jeong, Yeon Woo; Moon, Changjong; Hyun, Sang-Hwan; Kim, Nam Hyung; Jeung, Eui-Bae; Yang, Huanming; Hwang, Woo Suk; Gao, Fei

    2016-01-01

    Somatic cell nuclear transfer (SCNT) provides an excellent model for studying epigenomic reprogramming during mammalian development. We mapped the whole genome and whole methylome for potential anomalies of mutations or epimutations in SCNT-generated dogs with XY chromosomal sex but complete gonadal dysgenesis, which is classified as 78, XY disorder of sex development (DSD). Whole genome sequencing revealed no potential genomic variations that could explain the pathogenesis of DSD. However, extensive but stochastic anomalies of genome-wide DNA methylation were discovered in these SCNT DSD dogs. Persistent abnormal hypermethylation of the SRY gene was observed together with its down-regulated mRNA and protein expression. Failure of SRY expression due to hypermethylation was further correlated with silencing of a serial of testis determining genes, including SOX9, SF1, SOX8, AMH and DMRT1 in an early embryonic development stage at E34 in the XYDSD gonad, and high activation of the female specific genes, including FOXL2, RSPO1, CYP19A1, WNT4, ERα and ERβ, after one postnatal year in the ovotestis. Our results demonstrate that incomplete demethylation on the SRY gene is the driving cause of XYDSD in these XY DSD dogs, indicating a central role of epigenetic regulation in sex determination. PMID:27501986

  8. Trichostatin A specifically improves the aberrant expression of transcription factor genes in embryos produced by somatic cell nuclear transfer.

    PubMed

    Inoue, Kimiko; Oikawa, Mami; Kamimura, Satoshi; Ogonuki, Narumi; Nakamura, Toshinobu; Nakano, Toru; Abe, Kuniya; Ogura, Atsuo

    2015-01-01

    Although mammalian cloning by somatic cell nuclear transfer (SCNT) has been established in various species, the low developmental efficiency has hampered its practical applications. Treatment of SCNT-derived embryos with histone deacetylase (HDAC) inhibitors can improve their development, but the underlying mechanism is still unclear. To address this question, we analysed gene expression profiles of SCNT-derived 2-cell mouse embryos treated with trichostatin A (TSA), a potent HDAC inhibitor that is best used for mouse cloning. Unexpectedly, TSA had no effect on the numbers of aberrantly expressed genes or the overall gene expression pattern in the embryos. However, in-depth investigation by gene ontology and functional analyses revealed that TSA treatment specifically improved the expression of a small subset of genes encoding transcription factors and their regulatory factors, suggesting their positive involvement in de novo RNA synthesis. Indeed, introduction of one of such transcription factors, Spi-C, into the embryos at least partially mimicked the TSA-induced improvement in embryonic development by activating gene networks associated with transcriptional regulation. Thus, the effects of TSA treatment on embryonic gene expression did not seem to be stochastic, but more specific than expected, targeting genes that direct development and trigger zygotic genome activation at the 2-cell stage. PMID:25974394

  9. Stochastic anomaly of methylome but persistent SRY hypermethylation in disorder of sex development in canine somatic cell nuclear transfer.

    PubMed

    Jeong, Young-Hee; Lu, Hanlin; Park, Chi-Hun; Li, Meiyan; Luo, Huijuan; Kim, Joung Joo; Liu, Siyang; Ko, Kyeong Hee; Huang, Shujia; Hwang, In Sung; Kang, Mi Na; Gong, Desheng; Park, Kang Bae; Choi, Eun Ji; Park, Jung Hyun; Jeong, Yeon Woo; Moon, Changjong; Hyun, Sang-Hwan; Kim, Nam Hyung; Jeung, Eui-Bae; Yang, Huanming; Hwang, Woo Suk; Gao, Fei

    2016-01-01

    Somatic cell nuclear transfer (SCNT) provides an excellent model for studying epigenomic reprogramming during mammalian development. We mapped the whole genome and whole methylome for potential anomalies of mutations or epimutations in SCNT-generated dogs with XY chromosomal sex but complete gonadal dysgenesis, which is classified as 78, XY disorder of sex development (DSD). Whole genome sequencing revealed no potential genomic variations that could explain the pathogenesis of DSD. However, extensive but stochastic anomalies of genome-wide DNA methylation were discovered in these SCNT DSD dogs. Persistent abnormal hypermethylation of the SRY gene was observed together with its down-regulated mRNA and protein expression. Failure of SRY expression due to hypermethylation was further correlated with silencing of a serial of testis determining genes, including SOX9, SF1, SOX8, AMH and DMRT1 in an early embryonic development stage at E34 in the XY(DSD) gonad, and high activation of the female specific genes, including FOXL2, RSPO1, CYP19A1, WNT4, ERα and ERβ, after one postnatal year in the ovotestis. Our results demonstrate that incomplete demethylation on the SRY gene is the driving cause of XY(DSD) in these XY DSD dogs, indicating a central role of epigenetic regulation in sex determination. PMID:27501986

  10. Analysis of ENPP2 in the Uterine Endometrium of Pigs Carrying Somatic Cell Nuclear Transfer Cloned Embryos

    PubMed Central

    Seo, Heewon; Choi, Yohan; Yu, Inkyu; Shim, Jangsoo; Lee, Chang-Kyu; Hyun, Sang-Hwan; Lee, Eunsong; Ka, Hakhyun

    2013-01-01

    Somatic cell nuclear transfer (SCNT) is a useful tool for animal cloning, but the efficiency of producing viable offspring by SCNT is very low. To improve this efficiency in the production of cloned pigs, it is critical to understand the interactions between uterine function and cloned embryos during implantation. Lysophosphatidic acid (LPA) is a lipid mediator that plays an important role in the establishment of pregnancy in pigs; however, LPA production in the uterine endometrium of pigs carrying SCNT-cloned conceptuses has not been determined. Therefore, we investigated expression of ENPP2, an LPA-generating enzyme, in the uterine endometrium of gilts with conceptuses derived from SCNT during the implantation period. Uterine endometrial tissue and uterine flushing were obtained from gilts carrying SCNT-derived conceptuses and from gilts carrying conceptuses resulting from natural mating on d 12 of pregnancy. Our results demonstrated no difference in the level of ENPP2 mRNA expression in the uterine endometrium between gilts carrying SCNT-derived conceptuses and gilts carrying naturally-conceived conceptuses, but secretion of ENPP2 protein into the uterine lumen did decrease significantly in pigs with SCNT-derived conceptuses. These results indicate that expression and secretion of ENPP2, which are critical for appropriate LPA production and successful pregnancy, are dysregulated in the uterine endometrium of pigs carrying SCNT-derived conceptuses. PMID:25049907

  11. An ideal oocyte activation protocol and embryo culture conditions for somatic cell nuclear transfer using sheep oocytes.

    PubMed

    Patel, Hiren; Chougule, Shruti; Chohan, Parul; Shah, Naval; Bhartiya, Deepa

    2014-10-01

    Pluripotent stem cells are possibly the best candidates for regenerative medicine, and somatic cell nuclear transfer (SCNT) is one of the viable options to make patient-specific embryonic stem cells. Till date efficacy of SCNT embryos is very low and requires further improvement like ideal oocyte activation and in vitro culture system. The aim of the present study was to evaluate ideal oocyte activation using different stimulation protocols and to study the effect of cumulus co-culture conditions on embryo development. Results demonstrate that between electric stimulation and chemical stimulation using calcium ionomycin and ionophore, best oocyte activation was obtained using calcium ionomycin (5 microM for 5 min) which resulted in 83% cleavage followed by 7% of early blastocyst which further increased to 15% when a cumulus bed was also introduced during embryo culture. Sequential modified Charles Rosenkrans 2 (mCR2) medium was used for embryo culture in which glucose levels were increased from 1 mM to 5 mM from Day 3 onwards. SCNT using cumulus cells as donor somatic cell, calcium ionomycin to activate the reconstructed oocyte and embryo culture on a cumulus bed in sequential mCR2 medium, resulted in the development of 6% embryos to early blastocyst stage. Such technological advances will make SCNT a viable option to make patient-specific pluripotent stem cell lines in near future.

  12. Generation of porcine fetal fibroblasts expressing the tetracycline-inducible Cas9 gene by somatic cell nuclear transfer.

    PubMed

    Liu, Guoqian; Liu, Kai; Wei, Hengxi; Li, Li; Zhang, Shouquan

    2016-09-01

    Cas9 endonuclease, from so-called clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated (Cas) systems of Streptococcus pyogenes, type II functions as an RNA-guided endonuclease and edits the genomes of prokaryotic and eukaryotic organisms, including deletion and insertion by DNA double‑stranded break repair mechanisms. In previous studies, it was observed that Cas9, with a genome‑scale lentiviral single‑guide RNA library, could be applied to a loss‑of‑function genetic screen, although the loss‑of‑function genes have yet to be verified in vitro and this approach has not been used in porcine cells. Based on these observations, lentiviral Cas9 was used to infect porcine primary fibroblasts to achieve cell colonies carrying Cas9 endonuclease. Subsequently, porcine fetal fibroblasts expressing the tetracycline‑inducible Cas9 gene were generated by somatic cell nuclear transfer, and three 30 day transgenic porcine fetal fibroblasts (PFFs) were obtained. Polymerase chain reaction (PCR), reverse transcription‑PCR and western blot analysis indicated that the PFFs were Cas9‑positive. In addition, one of the three integrations was located near to known functional genes in the PFF1 cell line, whereas neither of the integrations was located in the PFF1 or PFF2 cell lines. It was hypothesized that these transgenic PFFs may be useful for conditional genomic editing in pigs, and for generating ideal modified porcine models. PMID:27430306

  13. Generation of porcine fetal fibroblasts expressing the tetracycline-inducible Cas9 gene by somatic cell nuclear transfer

    PubMed Central

    Liu, Guoqian; Liu, Kai; Wei, Hengxi; Li, Li; Zhang, Shouquan

    2016-01-01

    Cas9 endonuclease, from so-called clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated (Cas) systems of Streptococcus pyogenes, type II functions as an RNA-guided endonuclease and edits the genomes of prokaryotic and eukaryotic organisms, including deletion and insertion by DNA double-stranded break repair mechanisms. In previous studies, it was observed that Cas9, with a genome-scale lentiviral single-guide RNA library, could be applied to a loss-of-function genetic screen, although the loss-of-function genes have yet to be verified in vitro and this approach has not been used in porcine cells. Based on these observations, lentiviral Cas9 was used to infect porcine primary fibroblasts to achieve cell colonies carrying Cas9 endonuclease. Subsequently, porcine fetal fibroblasts expressing the tetracycline-inducible Cas9 gene were generated by somatic cell nuclear transfer, and three 30 day transgenic porcine fetal fibroblasts (PFFs) were obtained. Polymerase chain reaction (PCR), reverse transcription-PCR and western blot analysis indicated that the PFFs were Cas9-positive. In addition, one of the three integrations was located near to known functional genes in the PFF1 cell line, whereas neither of the integrations was located in the PFF1 or PFF2 cell lines. It was hypothesized that these transgenic PFFs may be useful for conditional genomic editing in pigs, and for generating ideal modified porcine models. PMID:27430306

  14. Dynamic aspects of antibody:oligosaccharide complexes characterized by molecular dynamics simulations and saturation transfer difference nuclear magnetic resonance.

    PubMed

    Theillet, François-Xavier; Frank, Martin; Vulliez-Le Normand, Brigitte; Simenel, Catherine; Hoos, Sylviane; Chaffotte, Alain; Bélot, Frédéric; Guerreiro, Catherine; Nato, Farida; Phalipon, Armelle; Mulard, Laurence A; Delepierre, Muriel

    2011-12-01

    Carbohydrates are likely to maintain significant conformational flexibility in antibody (Ab):carbohydrate complexes. As demonstrated herein for the protective monoclonal Ab (mAb) F22-4 recognizing the Shigella flexneri 2a O-antigen (O-Ag) and numerous synthetic oligosaccharide fragments thereof, the combination of molecular dynamics simulations and nuclear magnetic resonance saturation transfer difference experiments, supported by physicochemical analysis, allows us to determine the binding epitope and its various contributions to affinity without using any modified oligosaccharides. Moreover, the methods used provide insights into ligand flexibility in the complex, thus enabling a better understanding of the Ab affinities observed for a representative set of synthetic O-Ag fragments. Additionally, these complementary pieces of information give evidence to the ability of the studied mAb to recognize internal as well as terminal epitopes of its cognate polysaccharide antigen. Hence, we show that an appropriate combination of computational and experimental methods provides a basis to explore carbohydrate functional mimicry and receptor binding. The strategy may facilitate the design of either ligands or carbohydrate recognition domains, according to needed improvements of the natural carbohydrate:receptor properties. PMID:21610193

  15. Histone Demethylase Expression Enhances Human Somatic Cell Nuclear Transfer Efficiency and Promotes Derivation of Pluripotent Stem Cells.

    PubMed

    Chung, Young Gie; Matoba, Shogo; Liu, Yuting; Eum, Jin Hee; Lu, Falong; Jiang, Wei; Lee, Jeoung Eun; Sepilian, Vicken; Cha, Kwang Yul; Lee, Dong Ryul; Zhang, Yi

    2015-12-01

    The extremely low efficiency of human embryonic stem cell (hESC) derivation using somatic cell nuclear transfer (SCNT) limits its potential application. Blastocyst formation from human SCNT embryos occurs at a low rate and with only some oocyte donors. We previously showed in mice that reduction of histone H3 lysine 9 trimethylation (H3K9me3) through ectopic expression of the H3K9me3 demethylase Kdm4d greatly improves SCNT embryo development. Here we show that overexpression of a related H3K9me3 demethylase KDM4A improves human SCNT, and that, as in mice, H3K9me3 in the human somatic cell genome is an SCNT reprogramming barrier. Overexpression of KDM4A significantly improves the blastocyst formation rate in human SCNT embryos by facilitating transcriptional reprogramming, allowing efficient derivation of SCNT-derived ESCs using adult Age-related Macular Degeneration (AMD) patient somatic nuclei donors. This conserved mechanistic insight has potential applications for improving SCNT in a variety of contexts, including regenerative medicine. PMID:26526725

  16. Dynamic aspects of antibody:oligosaccharide complexes characterized by molecular dynamics simulations and saturation transfer difference nuclear magnetic resonance.

    PubMed

    Theillet, François-Xavier; Frank, Martin; Vulliez-Le Normand, Brigitte; Simenel, Catherine; Hoos, Sylviane; Chaffotte, Alain; Bélot, Frédéric; Guerreiro, Catherine; Nato, Farida; Phalipon, Armelle; Mulard, Laurence A; Delepierre, Muriel

    2011-12-01

    Carbohydrates are likely to maintain significant conformational flexibility in antibody (Ab):carbohydrate complexes. As demonstrated herein for the protective monoclonal Ab (mAb) F22-4 recognizing the Shigella flexneri 2a O-antigen (O-Ag) and numerous synthetic oligosaccharide fragments thereof, the combination of molecular dynamics simulations and nuclear magnetic resonance saturation transfer difference experiments, supported by physicochemical analysis, allows us to determine the binding epitope and its various contributions to affinity without using any modified oligosaccharides. Moreover, the methods used provide insights into ligand flexibility in the complex, thus enabling a better understanding of the Ab affinities observed for a representative set of synthetic O-Ag fragments. Additionally, these complementary pieces of information give evidence to the ability of the studied mAb to recognize internal as well as terminal epitopes of its cognate polysaccharide antigen. Hence, we show that an appropriate combination of computational and experimental methods provides a basis to explore carbohydrate functional mimicry and receptor binding. The strategy may facilitate the design of either ligands or carbohydrate recognition domains, according to needed improvements of the natural carbohydrate:receptor properties.

  17. Generation of GGTA1 biallelic knockout pigs via zinc-finger nucleases and somatic cell nuclear transfer.

    PubMed

    Bao, Lei; Chen, HaiDe; Jong, UiMyong; Rim, CholHo; Li, WenLing; Lin, XiJuan; Zhang, Dan; Luo, Qiong; Cui, Chun; Huang, HeFeng; Zhang, Yan; Xiao, Lei; Fu, ZhiXin

    2014-02-01

    Genetically modified pigs are valuable models of human disease and donors of xenotransplanted organs. Conventional gene targeting in pig somatic cells is extremely inefficient. Zinc-finger nuclease (ZFN) technology has been shown to be a powerful tool for efficiently inducing mutations in the genome. However, ZFN-mediated targeting in pigs has rarely been achieved. Here, we used ZFNs to knock out the porcine α-1, 3-galactosyl-transferase (GGTA1) gene, which generates Gal epitopes that trigger hyperacute immune rejection in pig-to-human transplantation. Primary pig fibroblasts were transfected with ZFNs targeting the coding region of GGTA1. Eighteen mono-allelic and four biallelic knockout cell clones were obtained after drug selection with efficiencies of 23.4% and 5.2%, respectively. The biallelic cells were used to produce cloned pigs via somatic cell nuclear transfer (SCNT). Three GGTA1 null piglets were born, and one knockout primary fibroblast cell line was established from a cloned fetus. Gal epitopes on GGTA1 null pig cells were completely eliminated from the cell membrane. Functionally, GGTA1 knockout cells were protected from complement-mediated immune attacks when incubated with human serum. This study demonstrated that ZFN is an efficient tool in creating gene-modified pigs. GGTA1 null pigs and GGTA1 null fetal fibroblasts would benefit research and pig-to-human transplantation. PMID:24430555

  18. Comparison of milk produced by cows cloned by nuclear transfer with milk from non-cloned cows.

    PubMed

    Walsh, Marie K; Lucey, John A; Govindasamy-Lucey, Selvarani; Pace, Marvin M; Bishop, Michael D

    2003-01-01

    Cloning technologies, including embryo splitting and nuclear transfer, were introduced into dairy cattle breeding in the early 1980s. With the recent worldwide attention on the cloning of sheep ("Dolly") and cows ("Gene"), the potential food safety concerns for food products derived from cloned animals needs to be addressed. There has been no study of the composition of milk produced by cloned cows. In this preliminary study, we evaluated the composition of milk from 15 lactating non-embryonic cell cloned cows and six non-cloned lactating cows over a single season. The cloned cows came from five unique genetic lines and three distinct breeds. Milk samples were analyzed for total solids, fat, fatty acid profile, lactose, protein and compared to non-cloned and literature values. Gross chemical composition of milk from cloned cows was similar to that of the non-cloned cows and literature values. Our results lead us to conclude that there are no obvious differences in milk composition produced from cloned cows compared to non-cloned cows. PMID:14588139

  19. Cloning adult farm animals: a review of the possibilities and problems associated with somatic cell nuclear transfer.

    PubMed

    Edwards, J L; Schrick, F N; McCracken, M D; van Amstel, S R; Hopkins, F M; Welborn, M G; Davies, C J

    2003-08-01

    In 1997, Wilmut et al. announced the birth of Dolly, the first ever clone of an adult animal. To date, adult sheep, goats, cattle, mice, pigs, cats and rabbits have been cloned using somatic cell nuclear transfer. The ultimate challenge of cloning procedures is to reprogram the somatic cell nucleus for development of the early embryo. The cell type of choice for reprogramming the somatic nucleus is an enucleated oocyte. Given that somatic cells are easily obtained from adult animals, cultured in the laboratory and then genetically modified, cloning procedures are ideal for introducing specific genetic modifications in farm animals. Genetic modification of farm animals provides a means of studying genes involved in a variety of biological systems and disease processes. Moreover, genetically modified farm animals have created a new form of 'pharming' whereby farm animals serve as bioreactors for production of pharmaceuticals or organ donors. A major limitation of cloning procedures is the extreme inefficiency for producing live offspring. Dolly was the only live offspring produced after 277 attempts. Similar inefficiencies for cloning adult animals of other species have been described by others. Many factors related to cloning procedures and culture environment contribute to the death of clones, both in the embryonic and fetal periods as well as during neonatal life. Extreme inefficiencies of this magnitude, along with the fact that death of the surrogate may occur, continue to raise great concerns with cloning humans. PMID:12846674

  20. Proteomic analysis of the major cellular proteins of bovine trophectoderm cell lines derived from IVP, parthenogenetic, and nuclear transfer embryos

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Nuclear cloning of cattle is currently very inefficient in terms of the production and survival of nuclear cloned calves. Because of the great promise of using nuclear cloning technology in farm animals to genetically improve their production and quality traits, nuclear cloning in cattle, and other...

  1. Potential of adipose-derived mesenchymal stem cells and skeletal muscle-derived satellite cells for somatic cell nuclear transfer mediated transgenesis in Arbas Cashmere goats.

    PubMed

    Ren, Yu; Wu, Haiqing; Ma, Yuzhen; Yuan, Jianlong; Liang, Hao; Liu, Dongjun

    2014-01-01

    Somatic cell nuclear transfer is used to generate genetic models for research and new, genetically modified livestock varieties. Goat fetal fibroblast cells (gFFCs) are the predominant nuclear donors in Cashmere goat transgenic cloning, but have disadvantages. We evaluated the potential of goat adipose-derived mesenchymal stem cells (gADSCs) and goat skeletal muscle-derived satellite cells (gMDSCs) for somatic cell nuclear transfer, evaluating their proliferation, pluripotency, transfection efficiency and capacity to support full term development of embryos after additive gene transfer or homologous recombination. gADSCs and gMDSCs were isolated by enzyme digestion and differentiated into neurocytes, myotube cells and insulin-producing cells. Neuron-specific enolase, fast muscle myosin and insulin expression were determined by immunohistochemistry. Following somatic cell nuclear transfer with donor cells derived from gADSCs, gMDSCs and gFFCs, transfection and cloning efficiencies were compared. Red fluorescent protein levels were determined by quantitative PCR and western blotting. 5-Methylcytosine, H4K5, H4K12 and H3K18 were determined immunohistochemically. gADSCs and gMDSCs were maintained in culture for up to 65 passages, whereas gFFCs could be passaged barely more than 15 times. gADSCs and gMDSCs had higher fluorescent colony forming efficiency and greater convergence (20%) and cleavage (10%) rates than gFFCs, and exhibited differing H4K5 histone modification patterns after somatic cell nuclear transfer and in vitro cultivation. After transfection with a pDsRed2-1 expression plasmid, the integrated exogenous genes did not influence the pluripotency of gADSCs-pDsRed2-1 or gMDSCs-pDsRed2-1. DsRed2 mRNA expression by cloned embryos derived from gADSCs-pDsRed2-1 or gMDSCs-pDsRed2-1 was more than twice that of gFFCs-pDsRed2-1 embryos (P<0.01). Pregnancy rates of gADSCs-pDsRed2-1 and gMDSCs-pDsRed2-1 recipients were higher than those of gFFCs-pDsRed2-1 recipients (P

  2. Potential of adipose-derived mesenchymal stem cells and skeletal muscle-derived satellite cells for somatic cell nuclear transfer mediated transgenesis in Arbas Cashmere goats.

    PubMed

    Ren, Yu; Wu, Haiqing; Ma, Yuzhen; Yuan, Jianlong; Liang, Hao; Liu, Dongjun

    2014-01-01

    Somatic cell nuclear transfer is used to generate genetic models for research and new, genetically modified livestock varieties. Goat fetal fibroblast cells (gFFCs) are the predominant nuclear donors in Cashmere goat transgenic cloning, but have disadvantages. We evaluated the potential of goat adipose-derived mesenchymal stem cells (gADSCs) and goat skeletal muscle-derived satellite cells (gMDSCs) for somatic cell nuclear transfer, evaluating their proliferation, pluripotency, transfection efficiency and capacity to support full term development of embryos after additive gene transfer or homologous recombination. gADSCs and gMDSCs were isolated by enzyme digestion and differentiated into neurocytes, myotube cells and insulin-producing cells. Neuron-specific enolase, fast muscle myosin and insulin expression were determined by immunohistochemistry. Following somatic cell nuclear transfer with donor cells derived from gADSCs, gMDSCs and gFFCs, transfection and cloning efficiencies were compared. Red fluorescent protein levels were determined by quantitative PCR and western blotting. 5-Methylcytosine, H4K5, H4K12 and H3K18 were determined immunohistochemically. gADSCs and gMDSCs were maintained in culture for up to 65 passages, whereas gFFCs could be passaged barely more than 15 times. gADSCs and gMDSCs had higher fluorescent colony forming efficiency and greater convergence (20%) and cleavage (10%) rates than gFFCs, and exhibited differing H4K5 histone modification patterns after somatic cell nuclear transfer and in vitro cultivation. After transfection with a pDsRed2-1 expression plasmid, the integrated exogenous genes did not influence the pluripotency of gADSCs-pDsRed2-1 or gMDSCs-pDsRed2-1. DsRed2 mRNA expression by cloned embryos derived from gADSCs-pDsRed2-1 or gMDSCs-pDsRed2-1 was more than twice that of gFFCs-pDsRed2-1 embryos (P<0.01). Pregnancy rates of gADSCs-pDsRed2-1 and gMDSCs-pDsRed2-1 recipients were higher than those of gFFCs-pDsRed2-1 recipients (P

  3. Potential of Adipose-Derived Mesenchymal Stem Cells and Skeletal Muscle-Derived Satellite Cells for Somatic Cell Nuclear Transfer Mediated Transgenesis in Arbas Cashmere Goats

    PubMed Central

    Yuan, Jianlong; Liang, Hao; Liu, Dongjun

    2014-01-01

    Somatic cell nuclear transfer is used to generate genetic models for research and new, genetically modified livestock varieties. Goat fetal fibroblast cells (gFFCs) are the predominant nuclear donors in Cashmere goat transgenic cloning, but have disadvantages. We evaluated the potential of goat adipose-derived mesenchymal stem cells (gADSCs) and goat skeletal muscle-derived satellite cells (gMDSCs) for somatic cell nuclear transfer, evaluating their proliferation, pluripotency, transfection efficiency and capacity to support full term development of embryos after additive gene transfer or homologous recombination. gADSCs and gMDSCs were isolated by enzyme digestion and differentiated into neurocytes, myotube cells and insulin-producing cells. Neuron-specific enolase, fast muscle myosin and insulin expression were determined by immunohistochemistry. Following somatic cell nuclear transfer with donor cells derived from gADSCs, gMDSCs and gFFCs, transfection and cloning efficiencies were compared. Red fluorescent protein levels were determined by quantitative PCR and western blotting. 5-Methylcytosine, H4K5, H4K12 and H3K18 were determined immunohistochemically. gADSCs and gMDSCs were maintained in culture for up to 65 passages, whereas gFFCs could be passaged barely more than 15 times. gADSCs and gMDSCs had higher fluorescent colony forming efficiency and greater convergence (20%) and cleavage (10%) rates than gFFCs, and exhibited differing H4K5 histone modification patterns after somatic cell nuclear transfer and in vitro cultivation. After transfection with a pDsRed2-1 expression plasmid, the integrated exogenous genes did not influence the pluripotency of gADSCs–pDsRed2-1 or gMDSCs–pDsRed2-1. DsRed2 mRNA expression by cloned embryos derived from gADSCs–pDsRed2-1 or gMDSCs–pDsRed2-1 was more than twice that of gFFCs–pDsRed2-1 embryos (P<0.01). Pregnancy rates of gADSCs–pDsRed2-1 and gMDSCs–pDsRed2-1 recipients were higher than those of gFFCs–pDsRed2

  4. Streptococcus pneumoniae R6 interspecies transformation: genetic analysis of penicillin resistance determinants and genome-wide recombination events.

    PubMed

    Sauerbier, Julia; Maurer, Patrick; Rieger, Martin; Hakenbeck, Regine

    2012-11-01

    Interspecies gene transfer has been implicated as the major driving force for the evolution of penicillin resistance in Streptococcus pneumoniae. Genomic alterations of S. pneumoniae R6 introduced during four successive transformations with DNA of the high-level penicillin-resistant Streptococcus mitis B6 with beta-lactam selection have now been determined and the contribution of genes to high resistance levels was analysed genetically. Essential for high level resistance to penicillins of the transformant CCCB was the combination of murM(B) (6) and the 3' region of pbp2b(B) (6) . Sequences of both genes were detected in clinical isolates of S. pneumoniae, confirming the participation of S. mitis in the global gene pool of beta-lactam resistance determinants. The S. mitis PBP1b gene which contains an authentic stop codon within the transpeptidase domain is now shown to contribute only marginal to resistance, but it is possible that the presence of its transglycosylase domain is important in the context of cognate PBPs. The genome sequence of CCCB revealed 36 recombination events, including deletion and acquisition of genes and repeat elements. A total of 78 genes were affected representing 67 kb or 3.3% of the genome, documenting extensive alterations scattered throughout the genome.

  5. Quantile-Quantile Plots:. AN Approach for the Inter-Species Comparison of Promoter Architecture in Eukaryotes

    NASA Astrophysics Data System (ADS)

    Feldmeier, Kaspar; Kilian, Joachim; Harter, Klaus; Wanke, Dierk; Berendzen, Kenneth W.

    2011-01-01

    Regulatory non-coding DNA is important to drive gene transcription and thereby influence mRNA and consequently protein abundance. Therefore, biologists and bioinformation scientists aim to extract meaningful information from these sequence regions, in particular upstream regulation regions called promoters, and conclude on regulatory sequence function. While some approaches have been successful for single genes or a single genome, it is an open question whether information on promoter function can readily be transferred between different species. Thus, it is useful for biologists to know more about the general structure and composition of promoters including the occurrence of cisregulatory DNA-elements (CREs) to be able to compare promoter architecture between organisms. To approach this task, we utilized the fully sequenced genomes of the plant model organisms: mouse-ear cress (Arabidopsis thaliana), western balsam poplar (Populus trichocarpa), Sorghum bicolor and rice (Oryza sativa). For the interspecies comparison we made use of quantile-quantile (QQ)-plots of the variances of hexanucleotides or known functional CREs of core-promoter regions. Here, we suggest that the differences in promoter architecture correlate with the sizes of the intergenic space, i.e. regions, in which the promoters are located. In contrast, analysis of CREs is hampered by the general lack of well characterized transcription factor-CRE-relationships.

  6. Development and investigations of compact heat-transfer equipment for a nuclear power station equipped with a high-temperature gas-cooled reactor

    NASA Astrophysics Data System (ADS)

    Golovko, V. F.; Dmitrieva, I. V.; Kodochigov, N. G.; Bykh, O. A.

    2013-07-01

    The project of a nuclear power station the reactor coolant system of which includes a high-temperature gas-cooled reactor combined with a gas-turbine energy conversion unit supposes the use of high-efficient gas-cycle-based heat-transfer equipment. An analysis aimed at selecting the optimal heat-transfer surfaces is presented together with the results from their calculated and experimental investigation. The design features of recuperators arranged integrally with end and intermediate coolers and placed in a vertical sealed high-pressure vessel of limited sizes are considered.

  7. Comparison of Species Sensitivity Distributions Derived from Interspecies Correlation Models to Distributions used to Derive Water Quality Criteria

    EPA Science Inventory

    Species sensitivity distributions (SSD) require a large number of measured toxicity values to define a chemical’s toxicity to multiple species. This investigation comprehensively evaluated the accuracy of SSDs generated from toxicity values predicted from interspecies correlation...

  8. Influence of Taxonomic Relatedness and Chemical Mode of Action in Acute Interspecies Estimation Models for Aquatic species

    EPA Science Inventory

    Ecological risks to aquatic organisms are typically assessed using toxicity data for relatively few species and with limited understanding of relative species sensitivity. We developed a comprehensive set of interspecies correlation estimation (ICE) models for aquatic organisms a...

  9. Interspecies systems biology uncovers metabolites affecting C. elegans gene expression and life history traits.

    PubMed

    Watson, Emma; MacNeil, Lesley T; Ritter, Ashlyn D; Yilmaz, L Safak; Rosebrock, Adam P; Caudy, Amy A; Walhout, Albertha J M

    2014-02-13

    Diet greatly influences gene expression and physiology. In mammals, elucidating the effects and mechanisms of individual nutrients is challenging due to the complexity of both the animal and its diet. Here, we used an interspecies systems biology approach with Caenorhabditis elegans and two of its bacterial diets, Escherichia coli and Comamonas aquatica, to identify metabolites that affect the animal's gene expression and physiology. We identify vitamin B12 as the major dilutable metabolite provided by Comamonas aq. that regulates gene expression, accelerates development, and reduces fertility but does not affect lifespan. We find that vitamin B12 has a dual role in the animal: it affects development and fertility via the methionine/S-Adenosylmethionine (SAM) cycle and breaks down the short-chain fatty acid propionic acid, preventing its toxic buildup. Our interspecies systems biology approach provides a paradigm for understanding complex interactions between diet and physiology.

  10. Inter-species embryos and human clones: issues of free movement and gestation.

    PubMed

    Laing, Jacqueline A

    2009-03-01

    The United Kingdom's Human Fertilisation and Embryology Bill, introduced into Parliament on the 8th of November 2007 contains a number of controversial proposals inter alia expressly permitting the creation of inter-species embryos for research and destruction and increasing the scope for human cloning also for destructive research. It is supposed that there ought not to be a blanket ban on the creation of human clones, hybrids, cybrids and chimeras because these embryos are valuable for research purposes. The prohibition on the gestation of non-permitted embryos and interspecies embryos is used to generate confidence that embryos with compromised origins would not be gestated and reared. The argument outlined here demonstrates how uncertain are any legal prohibitions on gestation. Accordingly, the practical import of the distinction between compromised embryos for research and the same for live birth is equally dubious. The legislation would not, on this analysis, supply effective controls over this reproductive technology.

  11. Interspecies Systems Biology Uncovers Metabolites Affecting C. elegans Gene Expression and Life History Traits

    PubMed Central

    Watson, Emma; MacNeil, Lesley T.; Ritter, Ashlyn D.; Yilmaz, L. Safak; Rosebrock, Adam P.; Caudy, Amy A.; Walhout, Albertha J. M.

    2014-01-01

    SUMMARY Diet greatly influences gene expression and physiology. In mammals, elucidating the effects and mechanisms of individual nutrients is challenging due to the complexity of both the animal and its diet. Here we used an interspecies systems biology approach with Caenorhabditis elegans and two if its bacterial diets, Escherichia coli and Comamonas aquatica, to identify metabolites that affect the animal’s gene expression and physiology. We identify vitamin B12 as the major dilutable metabolite provided by Comamonas aq. that regulates gene expression, accelerates development and reduces fertility, but does not affect lifespan. We find that vitamin B12 has a dual role in the animal: it affects development and fertility via the methionine/S-Adenosylmethionine (SAM) cycle and breaks down the short-chain fatty acid propionic acid preventing its toxic buildup. Our interspecies systems biology approach provides a paradigm for understanding complex interactions between diet and physiology. PMID:24529378

  12. Formation of interspecies fusants of Agaricus bisporus and Agaricus bitorquis mushroom by protoplast fusion.

    PubMed

    Singh, Rana Inder; Aarti, Kanojiya; Singh, Sandhu Sardul

    2007-12-01

    Interspecies fusants are formed between Agaricus bisporus and Agaricus bitorquis by protoplast fusion technique. Protoplasts were isolated and regenerated by using Novozyme 234 lytic enzyme. Twenty slow growing isolates were separated from the protoplast regenerated colonies, which were assumed as homokaryons (putative homokaryons). These twenty isolates were subjected to growth rate, colony morphology and spawn run studies for screening of true homokaryons. Antifungal markers were developed for selection of fusants. PMID:23100692

  13. Quantifying inter-species differences in contractile function through biophysical modelling.

    PubMed

    Tøndel, Kristin; Land, Sander; Niederer, Steven A; Smith, Nicolas P

    2015-03-01

    Animal models and measurements are frequently used to guide and evaluate clinical interventions. In this context, knowledge of inter-species differences in physiology is crucial for understanding the limitations and relevance of animal experimental assays for informing clinical applications. Extensive effort has been put into studying the structure and function of cardiac contractile proteins and how differences in these translate into the functional properties of muscles. However, integrating this knowledge into a quantitative description, formalising and highlighting inter-species differences both in the kinetics and in the regulation of physiological mechanisms, remains challenging. In this study we propose and apply a novel approach for the quantification of inter-species differences between mouse, rat and human. Assuming conservation of the fundamental physiological mechanisms underpinning contraction, biophysically based computational models are fitted to simulate experimentally recorded phenotypes from multiple species. The phenotypic differences between species are then succinctly quantified as differences in the biophysical model parameter values. This provides the potential of quantitatively establishing the human relevance of both animal-based experimental and computational models for application in a clinical context. Our results indicate that the parameters related to the sensitivity and cooperativity of calcium binding to troponin C and the activation and relaxation rates of tropomyosin/crossbridge binding kinetics differ most significantly between mouse, rat and human, while for example the reference tension, as expected, shows only minor differences between the species. Hence, while confirming expected inter-species differences in calcium sensitivity due to large differences in the observed calcium transients, our results also indicate more unexpected differences in the cooperativity mechanism. Specifically, the decrease in the unbinding rate of

  14. Space transfer concepts and analysis for exploration missions. Implementation plan and element description document (draft final). Volume 5: Nuclear electric propulsion vehicle

    NASA Technical Reports Server (NTRS)

    1991-01-01

    The nuclear electric propulsion (NEP) concept design developed in support of the Space Transfer Concepts and Analysis for Exploration Missions (STCAEM) study is presented. The evolution of the NEP concept is described along with the requirements, guidelines, and assumptions for the design. Operating modes and options are defined and a systems description of the vehicle is presented. Artificial gravity configuration options and space and ground support systems are discussed. Finally, an implementation plan is presented which addresses technology needs, schedules, facilities and costs.

  15. Vitamin C supplementation enhances compact morulae formation but reduces the hatching blastocyst rate of bovine somatic cell nuclear transfer embryos.

    PubMed

    Li, Qian; Wang, Yong-Sheng; Wang, Li-Jun; Zhang, Hui; Li, Rui-Zhe; Cui, Chen-Chen; Li, Wen-Zhe; Zhang, Yong; Jin, Ya-Ping

    2014-08-01

    Vitamin C, an antioxidant that reduces reactive oxygen species (ROS) in cells, is capable of significantly improving the developmental competence of porcine and mouse somatic cell nuclear transfer (SCNT) embryos, both in vitro and in vivo. In the present study, the effects of vitamin C on the developmental competence of bovine SCNT embryos were investigated. The results indicated that vitamin C (40 μg/mL) positively affected the scavenging of intracellular ROS, cleavage rate at 24 h (76.67 vs. 68.26%, p<0.05), compact morulae formation (60.83 vs. 51.30%, p<0.05), and the blastomere apoptosis index (3.70 ± 1.41 vs. 4.43% ± 1.65, p<0.05) of bovine SCNT embryos. However, vitamin C supplementation did not significantly affect the blastocyst formation rate and proportion of inner cell mass over total cells per blastocyst on day 7. Moreover, vitamin C supplementation obviously impaired the total cell numbers per blastocyst (97.20 ± 11.35 vs. 88.57 ± 10.43, p<0.05) on day 7 and the hatching blastocysts formation rate on day 9 (26.51 vs. 50.65%, p<0.05) compared with that of the untreated group. Vitamin C supplementation preferentially improved the viability of bovine SCNT embryos prior to the blastocyst stage, but did not enhance the formation and quality of blastocysts in vitro. In conclusion, the effect of vitamin C on the development of bovine SCNT embryos is complex, and vitamin C is not a suitable antioxidant chemical for the in vitro culture of bovine SCNT embryos.

  16. Identification of animals produced by somatic cell nuclear transfer using DNA methylation in the retrotransposon-like 1 promoter.

    PubMed

    Couldrey, Christine; Maclean, Paul; Wells, David N

    2014-12-01

    Public perception of somatic cell nuclear transfer (SCNT) in the production of agricultural animals is surrounded by fear, which is exacerbated by the inability to differentiate animals generated by SCNT from those generated by natural mating or artificial insemination (AI). Unfortunately, the DNA sequence of animals produced by SCNT is indistinguishable from those generated by fertilization. With the current banning of all SCNT animal products from entering the food supply in some countries, the lack of a diagnostic test to identify SCNT animals may jeopardize market access for producers. The aim of this research was to exploit differences in epigenetic reprogramming that occur during SCNT and fertilization in the early embryo. The resulting differences in epigenetic signatures that persist to adulthood are proposed as the basis for a diagnostic test to identify animals generated by SCNT. Here we describe differences in DNA methylation at eight CpG sites in the retrotransposon-like 1 (Rtl1) promoter region in cattle blood and test whether these differences could be used as a diagnostic tool. For a definitive diagnosis, it is critical that no overlap in DNA methylation levels is observed between individuals produced by SCNT and fertilization. This was the case for the cohort of SCNT animals studied, their female half-siblings generated by AI, and a collection of unrelated cows also generated by AI. Further rigorous testing is required to determine what effects donor cell type, age, sex, genetic background, SCNT methods, and the environment have on the DNA methylation across this region, but the Rtl1 promoter is currently a promising candidate for the identification of SCNT generated cattle.

  17. Using a nano-flare probe to detect RNA in live donor cells prior to somatic cell nuclear transfer.

    PubMed

    Fu, Bo; Ren, Liang; Liu, Di; Ma, Jian-Zhang; An, Tie-Zhu; Yang, Xiu-Qin; Ma, Hong; Guo, Zhen-Hua; Zhu, Meng; Bai, Jing

    2016-01-01

    Many transgenes are silenced in mammalian cells (donor cells used for somatic cell nuclear transfer [SCNT]). Silencing correlated with a repressed chromatin structure or suppressed promoter, and it impeded the production of transgenic animals. Gene transcription studies in live cells are challenging because of the drawbacks of reverse-transcription polymerase chain reaction and fluorescence in situ hybridization. Nano-flare probes provide an effective approach to detect RNA in living cells. We used 18S RNA, a housekeeping gene, as a reference gene. This study aimed to establish a platform to detect RNA in single living donor cells using a Nano-flare probe prior to SCNT and to verify the safety and validity of the Nano-flare probe in order to provide a technical foundation for rescuing silenced transgenes in transgenic cloned embryos. We investigated cytotoxic effect of the 18S RNA-Nano-flare probe on porcine fetal fibroblasts, characterized the distribution of the 18S RNA-Nano-flare probe in living cells and investigated the effect of the 18S RNA-Nano-flare probe on the development of cloned embryos after SCNT. The cytotoxic effect of the 18S RNA-Nano-flare probe on porcine fetal fibroblasts was dose-dependent, and 18S RNA was detected using the 18S RNA-Nano-flare probe. In addition, treating donor cells with 500 pM 18S RNA-Nano-flare probe did not have adverse effects on the development of SCNT embryos at the pre-implantation stage. In conclusion, we established a preliminary platform to detect RNA in live donor cells using a Nano-flare probe prior to SCNT.

  18. Ultrastructural comparison of porcine putative embryonic stem cells derived by in vitro fertilization and somatic cell nuclear transfer

    PubMed Central

    YOO, Hyunju; KIM, Eunhye; HWANG, Seon-Ung; YOON, Junchul David; JEON, Yubyeol; PARK, Kyu-Mi; KIM, Kyu-Jun; JIN, Minghui; LEE, Chang-Kyu; LEE, Eunsong; KIM, Hyunggee; KIM, Gonhyung; HYUN, Sang-Hwan

    2016-01-01

    The ultrastructure of porcine putative embryonic stem cells and porcine fetal fibroblasts (PFFs) was analyzed by transmission electron microscopy. The aim of this study was to compare the features of organelles in in vitro fertilization (IVF) derived porcine embryonic stem cells (IVF-pESCs) and somatic cell nuclear transfer (SCNT) derived pESCs (SCNT-pESCs). Also, the features of organelles in high-passage IVF-pESCs were compared with those in low-passage cells. The ultrastructure of PFFs showed rare microvilli on the cell surfaces, polygonal or irregular nuclei with one to two reticular-shaped nucleoli and euchromatin, low cytoplasm-to-nucleus ratios, rare ribosomes, rare rough endoplasmic reticulum, elongated mitochondria, rich lysosomes and rich phagocytic vacuoles. IVF-pESCs showed rare microvilli on the cell surfaces, round or irregular nuclei with one to two reticular-shaped nucleoli and euchromatin, low cytoplasm-to-nucleus ratios, rich ribosomes, long stacks of rough endoplasmic reticulum, elongated mitochondria, rare lysosomes and rare autophagic vacuoles. By contrast, SCNT-pESCs showed rich microvilli with various lengths and frequencies on the cell surfaces, polygonal nuclei with one reticular shaped nucleoli and heterochromatin, high cytoplasm-to-nucleus ratios, rare ribosomes, rare rough endoplasmic reticulum, round mitochondria, rich lysosomes and rich phagocytic vacuoles with clear intercellular junctions. Furthermore, high-passage IVF-pESCs showed irregularly shaped colonies, pyknosis and numerous lysosomes associated with autophagic vacuoles showing signs of apoptosis. In conclusion, this study confirms that the ultrastructural characteristics of pESCs differ depending on their origin. These ultrastructural characteristics might be useful in biomedical research using pESCs, leading to new insights regarding regenerative medicine and tissue repair. PMID:26821870

  19. Aberrant gene expression patterns in placentomes are associated with phenotypically normal and abnormal cattle cloned by somatic cell nuclear transfer.

    PubMed

    Everts, Robin E; Chavatte-Palmer, Pascale; Razzak, Anthony; Hue, Isabelle; Green, Cheryl A; Oliveira, Rosane; Vignon, Xavier; Rodriguez-Zas, Sandra L; Tian, X Cindy; Yang, Xiangzhong; Renard, Jean-Paul; Lewin, Harris A

    2008-03-14

    Transcription profiling of placentomes derived from somatic cell nuclear transfer (SCNT, n = 20), in vitro fertilization (IVF, n = 9), and artificial insemination (AI, n = 9) at or near term development was performed to better understand why SCNT and IVF often result in placental defects, hydrops, and large offspring syndrome (LOS). Multivariate analysis of variance was used to distinguish the effects of SCNT, IVF, and AI on gene expression, taking into account the effects of parturition (term or preterm), sex of fetus, breed of dam, breed of fetus, and pathological finding in the offspring (hydrops, normal, or other abnormalities). Differential expression of 20 physiologically important genes was confirmed with quantitative PCR. The largest effect on placentome gene expression was attributable to whether placentas were collected at term or preterm (i.e., whether the collection was because of disease or to obtain stage-matched controls) followed by placentome source (AI, IVF, or SCNT). Gene expression in SCNT placentomes was dramatically different from AI (n = 336 genes; 276 >2-fold) and from IVF (n = 733 genes; 162 >2-fold) placentomes. Functional analysis of differentially expressed genes (DEG) showed that IVF has significant effects on genes associated with cellular metabolism. In contrast, DEG associated with SCNT are involved in multiple pathways, including cell cycle, cell death, and gene expression. Many DEG were shared between the gene lists for IVF and SCNT comparisons, suggesting that common pathways are affected by the embryo culture methods used for IVF and SCNT. However, the many unique gene functions and pathways affected by SCNT suggest that cloned fetuses may be starved and accumulating toxic wastes due to placental insufficiency caused by reprogramming errors. Many of these genes are candidates for hydrops and LOS.

  20. Revival of extinct species using nuclear transfer: hope for the mammoth, true for the Pyrenean ibex, but is it time for "conservation cloning"?

    PubMed

    Piña-Aguilar, Raul E; Lopez-Saucedo, Janet; Sheffield, Richard; Ruiz-Galaz, Lilia I; Barroso-Padilla, Jose de J; Gutiérrez-Gutiérrez, Antonio

    2009-09-01

    Recent accomplishments in the fields of nuclear transfer and genomics, such as the cloned offspring production from frozen mouse cells, cryopreserved at not too low temperatures without cryoprotectors; or the sequencing of wooly mammoth genome, have opened the opportunity for the revival of extinct species. As expected, they are receiving a lot of publicity in the media and also scientific attention. Furthermore, it was recently published the "revival" of the first extinct subspecie: the Pyrenean ibex (Capra pyrenaica pyrenaica), a wild goat extinct in 2000. This strengthens the field of cloning as it had been tarnished by induced pluripotent stem cells (iPS) and other methods of reprogramming. However, for biological conservation purposes, cloning is not generally accepted as an alternative for animal conservation, and there is an ongoing debate between reproductive scientists and conservation specialists. Although we believe that nuclear transfer technologies have an opportunity in conservation efforts for some species that are on the brink of extinction and that population status, geographical isolation, reproductive characteristics, and human pressure create a situation that is almost unsustainable. In this article we discuss the barriers in cloning mammoths and cloning controversies in conservation from a zoological perspective, citing the species that might benefit from nuclear transfer techniques in the arduous journey so as not to disappear forever from this, our world.

  1. Revival of extinct species using nuclear transfer: hope for the mammoth, true for the Pyrenean ibex, but is it time for "conservation cloning"?

    PubMed

    Piña-Aguilar, Raul E; Lopez-Saucedo, Janet; Sheffield, Richard; Ruiz-Galaz, Lilia I; Barroso-Padilla, Jose de J; Gutiérrez-Gutiérrez, Antonio

    2009-09-01

    Recent accomplishments in the fields of nuclear transfer and genomics, such as the cloned offspring production from frozen mouse cells, cryopreserved at not too low temperatures without cryoprotectors; or the sequencing of wooly mammoth genome, have opened the opportunity for the revival of extinct species. As expected, they are receiving a lot of publicity in the media and also scientific attention. Furthermore, it was recently published the "revival" of the first extinct subspecie: the Pyrenean ibex (Capra pyrenaica pyrenaica), a wild goat extinct in 2000. This strengthens the field of cloning as it had been tarnished by induced pluripotent stem cells (iPS) and other methods of reprogramming. However, for biological conservation purposes, cloning is not generally accepted as an alternative for animal conservation, and there is an ongoing debate between reproductive scientists and conservation specialists. Although we believe that nuclear transfer technologies have an opportunity in conservation efforts for some species that are on the brink of extinction and that population status, geographical isolation, reproductive characteristics, and human pressure create a situation that is almost unsustainable. In this article we discuss the barriers in cloning mammoths and cloning controversies in conservation from a zoological perspective, citing the species that might benefit from nuclear transfer techniques in the arduous journey so as not to disappear forever from this, our world. PMID:19594389

  2. Plutonium from Above-Ground Nuclear Tests in Milk Teeth: Investigation of Placental Transfer in Children Born between 1951 and 1995 in Switzerland

    PubMed Central

    Froidevaux, Pascal; Haldimann, Max

    2008-01-01

    Background Occupational risks, the present nuclear threat, and the potential danger associated with nuclear power have raised concerns regarding the metabolism of plutonium in pregnant women. Objective We measured plutonium levels in the milk teeth of children born between 1951 and 1995 to assess the potential risk that plutonium incorporated by pregnant women might pose to the radiosensitive tissues of the fetus through placenta transfer. Methods We used milk teeth, whose enamel is formed during pregnancy, to investigate the transfer of plutonium from the mother’s blood plasma to the fetus. We measured plutonium using sensitive sector field inductively coupled plasma mass spectrometry techniques. We compared our results with those of a previous study on strontium-90 (90Sr) released into the atmosphere after nuclear bomb tests. Results Results show that plutonium activity peaks in the milk teeth of children born about 10 years before the highest recorded levels of plutonium fallout. By contrast, 90Sr, which is known to cross the placenta barrier, manifests differently in milk teeth, in accordance with 90Sr fallout deposition as a function of time. Conclusions These findings demonstrate that plutonium found in milk teeth is caused by fallout that was inhaled around the time the milk teeth were shed and not from any accumulation during pregnancy through placenta transfer. Thus, plutonium may not represent a radiologic risk for the radiosensitive tissues of the fetus. PMID:19079728

  3. An Overview of strategic measures to assess workforce needs and ensure technology transfer to meet current and future nuclear power operations

    SciTech Connect

    Vincenti, J.R.; Stigers, R.A.

    2007-07-01

    Between 1956 and 1989, the number of operating commercial nuclear power plants in the United States increased from none to 109. With the exception of a few plants that were still in final construction, no new nuclear power plants were ordered in the United States as the new millennium began. In 2005, the federal government pronounced the need for new electric power generating systems during the first quarter of the 21. century. The need comes from a desire to curb our reliance on fossil fuels, as well as to provide for a cleaner environment. One of those fuel systems noted was nuclear energy. Given the time between the last active period of nuclear power plant development and construction, there is a need to supply a talented and well-prepared workforce to operate the new plants. It will also be necessary to assess the needs of our current fleet of operating nuclear power plants, of which many are in the process of re-licensing, yet also facing an aging plant workforce. This paper will review and discuss measures to assess diverse workforce needs and technology transfer to meet current licensing requirements as that of future nuclear power plant development in the United States. (authors)

  4. Ovum pick up, intracytoplasmic sperm injection and somatic cell nuclear transfer in cattle, buffalo and horses: from the research laboratory to clinical practice.

    PubMed

    Galli, Cesare; Duchi, Roberto; Colleoni, Silvia; Lagutina, Irina; Lazzari, Giovanna

    2014-01-01

    Assisted reproductive techniques developed for cattle in the last 25 years, like ovum pick up (OPU), intracytoplasmic sperm injection (ICSI), and somatic cell nuclear transfer, have been transferred and adapted to buffalo and horses. The successful clinical applications of these techniques require both the clinical skills specific to each animal species and an experienced laboratory team to support the in vitro phase of the work. In cattle, OPU can be considered a consolidated technology that is rapidly outpacing conventional superovulation for embryo transfer. In buffalo, OPU represents the only possibility for embryo production to advance the implementation of embryo-based biotechnologies in that industry, although it is still mainly in the developmental phase. In the horse, OPU is now an established procedure for breeding from infertile and sporting mares throughout the year. It requires ICSI that in the horse, contrary to what happens in cattle and buffalo, is very efficient and the only option because conventional IVF does not work. Somatic cell nuclear transfer is destined to fill a very small niche for generating animals of extremely high commercial value. The efficiency is low, but because normal animals can be generated it is likely that advancing our knowledge in that field might improve the technology and reduce its cost.

  5. Generation of dwarf goat (Capra hircus) clones following nuclear transfer with transfected and nontransfected fetal fibroblasts and in vitro-matured oocytes.

    PubMed

    Keefer, C L; Baldassarre, H; Keyston, R; Wang, B; Bhatia, B; Bilodeau, A S; Zhou, J F; Leduc, M; Downey, B R; Lazaris, A; Karatzas, C N

    2001-03-01

    The developmental potential of caprine fetal fibroblast nuclei after in vitro transfection and nuclear transfer (NT) into enucleated, in vitro-matured oocytes was evaluated. Fetal fibroblasts were isolated from Day 27 to Day 30 fetuses from a dwarf breed of goat (BELE: breed early lactate early). Cells were transfected with constructs containing the enhanced green fluorescent protein (eGFP) and neomycin resistance genes and were selected with G418. Three eGFP lines and one nontransfected line were used as donor cells in NT. Donor cells were cultured in Dulbecco minimum Eagle medium plus 0.5% fetal calf serum for 4-8 days prior to use in NT. Immature oocytes were recovered by laparoscopic ovum pick-up and matured for 24 h prior to enucleation and NT. Reconstructed embryos were transferred as cleaved embryos into synchronized recipients. A total of 27 embryos derived from transgenic cells and 70 embryos derived from nontransgenic cells were transferred into 13 recipients. Five recipients (38%) were confirmed pregnant at Day 35 by ultrasound. Of these, four recipients delivered five male kids (7.1% of embryos transferred) derived from the nontransfected line. One recipient delivered a female kid derived from an eGFP line (7.7% of embryos transferred for that cell line). Presence of the eGFP transgene was confirmed by polymerase chain reaction, Southern blotting, and fluorescent in situ hybridization analyses. Nuclear transfer derivation from the donor cells was confirmed by single-strand confirmation polymorphism analysis. These results demonstrate that both in vitro-transfected and nontransfected caprine fetal fibroblasts can direct full-term development following NT.

  6. Quantifying inter-species differences in contractile function through biophysical modelling

    PubMed Central

    Tøndel, Kristin; Land, Sander; Niederer, Steven A; Smith, Nicolas P

    2015-01-01

    Abstract Animal models and measurements are frequently used to guide and evaluate clinical interventions. In this context, knowledge of inter-species differences in physiology is crucial for understanding the limitations and relevance of animal experimental assays for informing clinical applications. Extensive effort has been put into studying the structure and function of cardiac contractile proteins and how differences in these translate into the functional properties of muscles. However, integrating this knowledge into a quantitative description, formalising and highlighting inter-species differences both in the kinetics and in the regulation of physiological mechanisms, remains challenging. In this study we propose and apply a novel approach for the quantification of inter-species differences between mouse, rat and human. Assuming conservation of the fundamental physiological mechanisms underpinning contraction, biophysically based computational models are fitted to simulate experimentally recorded phenotypes from multiple species. The phenotypic differences between species are then succinctly quantified as differences in the biophysical model parameter values. This provides the potential of quantitatively establishing the human relevance of both animal-based experimental and computational models for application in a clinical context. Our results indicate that the parameters related to the sensitivity and cooperativity of calcium binding to troponin C and the activation and relaxation rates of tropomyosin/crossbridge binding kinetics differ most significantly between mouse, rat and human, while for example the reference tension, as expected, shows only minor differences between the species. Hence, while confirming expected inter-species differences in calcium sensitivity due to large differences in the observed calcium transients, our results also indicate more unexpected differences in the cooperativity mechanism. Specifically, the decrease in the unbinding

  7. Construction of murine coronavirus mutants containing interspecies chimeric nucleocapsid proteins.

    PubMed Central

    Peng, D; Koetzner, C A; McMahon, T; Zhu, Y; Masters, P S

    1995-01-01

    Targeted RNA recombination was used to construct mouse hepatitis virus (MHV) mutants containing chimeric nucleocapsid (N) protein genes in which segments of the bovine coronavirus N gene were substituted in place of their corresponding MHV sequences. This defined portions of the two N proteins that, despite evolutionary divergence, have remained functionally equivalent. These regions included most of the centrally located RNA-binding domain and two putative spacers that link the three domains of the N protein. By contrast, the amino terminus of N, the acidic carboxy-terminal domain, and a serine- and arginine-rich segment of the central domain could not be transferred from bovine coronavirus to MHV, presumably because these parts of the molecule participate in protein-protein interactions that are specific for each virus (or, possibly, each host). Our results demonstrate that targeted recombination can be used to make extensive substitutions in the coronavirus genome and can generate recombinants that could not otherwise be made between two viruses separated by a species barrier. The implications of these findings for N protein structure and function as well as for coronavirus RNA recombination are discussed. PMID:7636993

  8. Thermal-Hydraulic Analyses of Heat Transfer Fluid Requirements and Characteristics for Coupling A Hydrogen Production Plant to a High-Temperature Nuclear Reactor

    SciTech Connect

    C. B. Davis; C. H. Oh; R. B. Barner; D. F. Wilson

    2005-06-01

    The Department of Energy is investigating the use of high-temperature nuclear reactors to produce hydrogen using either thermochemical cycles or high-temperature electrolysis. Although the hydrogen production processes are in an early stage of development, coupling either of these processes to the hightemperature reactor requires both efficient heat transfer and adequate separation of the facilities to assure that off-normal events in the production facility do not impact the nuclear power plant. An intermediate heat transport loop will be required to separate the operations and safety functions of the nuclear and hydrogen plants. A next generation high-temperature reactor could be envisioned as a single-purpose facility that produces hydrogen or a dual-purpose facility that produces hydrogen and electricity. Early plants, such as the proposed Next Generation Nuclear Plant, may be dual-purpose facilities that demonstrate both hydrogen and efficient electrical generation. Later plants could be single-purpose facilities. At this stage of development, both single- and dual-purpose facilities need to be understood. Seven possible configurations for a system that transfers heat between the nuclear reactor and the hydrogen and/or electrical generation plants were identified. These configurations included both direct and indirect cycles for the production of electricity. Both helium and liquid salts were considered as the working fluid in the intermediate heat transport loop. Methods were developed to perform thermalhydraulic and cycle-efficiency evaluations of the different configurations and coolants. The thermalhydraulic evaluations estimated the sizes of various components in the intermediate heat transport loop for the different configurations. The relative sizes of components provide a relative indication of the capital cost associated with the various configurations. Estimates of the overall cycle efficiency of the various configurations were also determined. The

  9. Effects of long-term in vitro culturing of transgenic bovine donor fibroblasts on cell viability and in vitro developmental potential after nuclear transfer.

    PubMed

    Bressan, F F; Miranda, M S; Bajgelman, M C; Perecin, F; Mesquita, L G; Fantinato-Neto, P; Merighe, G F K; Strauss, B E; Meirelles, F V

    2013-04-01

    Genetically modified animals have numerous applications, ranging from basic research to livestock production and agriculture. Recent progress in animal cloning by nuclear transfer has made possible the production of transgenic animals using previously genetically modified cell lineages. However, to produce such lineages, an additional time for in vitro culturing and great manipulation is needed. Herein, we aimed to characterize different aspects of genetically modified cells compared to control cells, and we also analyzed the development rate of embryos produced by nuclear transfer by using them as nuclei donors after short or long periods of in vitro culturing (early versus late passages). We hypothesized that the genetic material inserted in the genome of these cells, associated with the prolonged time in culture, ultimately alters cell growth physiology and cell viability, which leads to impaired nuclei reprogramming potential and consequent reduction in the production of cloned blastocysts. Fetal fibroblasts expressing the enhanced Green Fluorescent Protein gene (eGFP) cultured for different periods in vitro were analyzed with respect to chromosomal numeric abnormalities, nuclear DNA fragmentation, the ratio of BAX and BCL2 gene transcripts, and the intensity of mitochondrial membrane potential, and they were then used as nuclei donors for somatic cell nuclear transfer (SCNT). Early passages were defined as fewer than 11 passages, and late passages were 18th passage (18(th)p) to 21(st)p. No differences were observed in the percentage of cells with chromosomal abnormalities or in the mitochondrial membrane potential analysis. eGFP cells in late passages and control cells in early passages were not different regarding DNA fragmentation; however, control cells in late passages presented higher fragmentation (P < 0.05). The Bax and Bcl2 gene expression ratio in control and transgenic cells presented different patterns regarding cell conditions during culture. For

  10. Technical aspects of the piezo, laser-assisted, and conventional methods for nuclear transfer of mouse oocytes and their efficiency and efficacy: Piezo minimizes damage of the ooplasmic membrane at injection.

    PubMed

    Chen, Shee-Uan; Chao, Kuang-Han; Chang, Chia-Yi; Hsieh, Fon-Jou; Ho, Hong-Nerng; Yang, Yu-Shih

    2004-04-01

    Assessment of the advantages and disadvantages of the piezo, laser, and conventional methods for nuclear transfer has remained elusive. Furthermore, although the piezo method had been used by some investigators for research of sperm injection and nuclear transfer for several years, many researchers have failed to operate the technique smoothly and achieve reproducible results. The procedures of nuclear transfer using piezo were ascertained and described in detail. Mouse oocytes were enucleated, and injected with cumulus cells using the piezo, laser, or conventional methods. We investigated the time needed and survival of nuclear transfer. Development was compared among the three methods and parthenogenetic control specimens. The average time of nuclear transfer for each oocyte was significantly shorter using the piezo (118 +/- 9 s) and laser methods (120 +/- 11 s) than using the conventional method (170 +/- 11 s). The damage rate was smaller for the piezo group (10%) than the laser (37%) and conventional (40%) groups. The percentages of blastocyst formation (14%, 12%, and 11%) and the number of nuclei of blastocysts (54 +/- 13, 51 +/- 11, and 52 +/- 12) were similar among the piezo, laser, and conventional groups, but significantly lower than for the control group (83%, 105 +/- 14). The piezo technique is more efficient than the conventional method for nuclear transfer. The laser method is easy to operate, but the equipment is expensive. In addition, piezo induced fewer traumas while breaking the membrane than the aspiration techniques used in the laser and conventional methods. PMID:15039993

  11. Generation and transfer of coherence in electron-nuclear spin systems by non-ideal microwave pulses

    NASA Astrophysics Data System (ADS)

    Jeschke, Gunnar

    A quantum mechanical treatment based on a suitably truncated Hamiltonian is developed for the evolution during microwave (MW) irradiation of an S = 1/2, I = 1/2 electron-nuclear spin system with anisotropic hyperfine coupling. It is shown that there is a close analogy between this problem and the well understood phenomena in heteronuclear spin systems with resolved J couplings during double irradiation (L. Muller and R. R. Ernst, 1979, Molec. Phys., 38, 963). A general expression is derived for the MW field strength necessary to establish the modified Hartmann-Hahn condition where electron and nuclear spin states mix completely and maximum nuclear coherence can be generated. The theory is then applied to the discussion of hyperfine decoupling and coherence generation from Boltzmann equilibrium by a MW pulse of arbitrary field strength and duration. It is demonstrated that the build-up of nuclear coherence during such a pulse is slow and oscillatory, and that maximum nuclear coherence is achieved at nominal flip angles considerably larger than 2 pi. The creation of both nuclear coherence and coherence on forbidden electron transitions is found to be optimum at the modified Hartmann-Hahn condition, and not to be limited by the modulation depth parameter known from electron spin echo envelope modulation (ESEEM) theory. The theoretical results are used to interpret a number of recent experimental observations that could not be understood by existing theories, and to propose an alternative explanation for nuclear modulations observed in experiments on bacterial photosystems. Effects in systems with a nuclear spin I = 1 are discussed taking into account nuclear quadrupole interactions. It is demonstrated both by numerical simulations and experiments that the well established two-pulse ESEEM experiment can be optimized by using pulses with matched microwave field strength and nominal flip angles larger than 2 pi.

  12. Energy metabolism during microsurgical transfer of human skeletal muscle assessed by high-pressure liquid chromatography and by 31P-nuclear magnetic resonance.

    PubMed

    Lundberg, Jonas; Elander, Anna; Rakotonirainy, Olivier; Zetterlund, Therese; Fogdestam, Ingemar; Soussi, Bassam

    2002-01-01

    The effect of ischaemia and reperfusion on human skeletal muscle was studied during free vascularised muscle transfer. Muscle biopsy specimens were taken from patients having microsurgical muscle transfer, 18 cases (17 patients; 12 men, 5 women). The biopsies were taken three times: before transfer of the muscle (control), at maximum ischaemic time, and one hour after revascularisation. The biopsy specimens were analysed for purine nucleotides, by high-pressure liquid chromatography (HPLC), and by nuclear magnetic resonance (NMR) at 500 MHz. Phosphocreatine (PCr) recovered only partially (79%) and adenosine triphosphate (ATP) did not differ significantly from normal control after revascularisation and a mean ischaemic time of 114 minutes. NMR measurements showed an accumulation of glucose-6-phosphate (G-6-P) during the ischaemic period, indicating anaerobic metabolism. After three hours of ischaemia and one hour of reperfusion the PCr recovery was less than 60% (r = 0.7). The results confirm those of previous animal studies, which set three hours normothermic ischaemia as a safe limit for tissue preservation when transferring skeletal muscle. Longer ischaemic times may cause serious postoperative healing problems and reduced muscle function.

  13. Momentum transfer dependence of nuclear transparency from the quasielastic [sup 12]C([ital e],[ital e][prime][ital p]) reaction

    SciTech Connect

    Makins, N.C.R.; Ent, R.; Chapman, M.S.; Hansen, J.; Lee, K.; Milner, R.G.; Nelson, J. ); Arnold, R.G.; Bosted, P.E.; Keppel, C.E.; Lung, A.; Rock, S.E.; Spengos, M.; Szalata, Z.M.; Tao, L.H.; White, J.L. ); Coulter, K.P.; Geesaman, D.F.; Holt, R.J.; Jackson, H.E.; Papavassiliou, V.; Potterveld, D.H.; Zeidman, B. ); Arrington, J.; Beise, E.J.; Belz, E.; Filippone, B.W.; Gao, H.; Lorenzon, W.; Mueller, B.; McKeown, R.D.; O'Neill, T.G. ); Epstein, M.; Margaziotis, D.J. ); Napolitano, J. (Rensselaer Polytechnic Institute, Troy, New York 12180 (United S

    1994-03-28

    The cross section for quasielastic [sup 12]C([ital e],[ital e][prime][ital p]) scattering has been measured at momentum transfer [ital Q][sup 2]=1, 3, 5, and 6.8 (GeV/[ital c])[sup 2]. The results are consistent with scattering from a single nucleon as the dominant process. The nuclear transparency is obtained and compared with theoretical calculations that incorporate color transparency effects. No significant rise of the transparency with [ital Q][sup 2] is observed.

  14. Space transfer concepts and analysis for exploration missions. Implementation plan and element description document (draft final). Volume 3: Nuclear thermal rocket vehicle

    NASA Technical Reports Server (NTRS)

    1991-01-01

    This document presents the nuclear thermal rocket (NTR) concept design developed in support of the Space Transfer Concepts and Analysis for Exploration Missions (STCAEM) study. The evolution of the NTR concept is described along with the requirements, guidelines and assumptions for the design. Operating modes and options are defined and a systems description of the vehicle is presented. Artificial gravity configuration options and space and ground support systems are discussed. Finally, an implementation plan is presented which addresses technology needs, schedules, facilities and costs.

  15. Chemically Assisted Enucleation Results in Higher G6PD Expression in Early Bovine Female Embryos Obtained by Somatic Cell Nuclear Transfer

    PubMed Central

    Oliveira, Clara Slade; Tetzner, Tatiane Almeida Drummond; de Lima, Marina Ragagnin; de Melo, Danilas Salinet; Niciura, Simone Cristina Méo; Garcia, Joaquim Mansano

    2012-01-01

    Abstract Despite extensive efforts, low efficiency is still an issue in bovine somatic cell nuclear transfer (SCNT). The hypothesis of our study was that the use of cytoplasts produced by chemically assisted enucleation (EN) would improve nuclear reprogramming in nuclear transfer (NT)–derived embryos because it results in lower damage and higher cytoplasm content than conventional EN. For that purpose, we investigated the expression of two X-linked genes: X inactive-specific transcript (XIST) and glucose 6-phosphate dehydrogenase (G6PD). In the first experiment, gene expression was assessed in day-7 female blastocysts from embryonic cell NT (ECNT) groups [conventional, ECNT conv; chemically assisted, ECNT deme (demecolcine)]. Whereas in the ECNT conv group, only one embryo (25%; n=4) expressed XIST transcripts, most embryos showed XIST expression (75%; n=4) in the ECNT deme group. However, no significant differences in transcript abundance of XIST and G6PD were found when comparing the embryos from all groups. In a second experiment using somatic cells as nuclear donors, we evaluated gene expression profiles in female SCNT-derived embryos. No significant differences in relative abundance (RA) of XIST transcripts were observed among the groups. Nonetheless, higher (p<0.05) levels of G6PD were observed in SCNT deme and in vitro–derived groups in comparison to SCNT conv. To know whether higher G6PD expression in embryos derived from SCNT chemically assisted EN indicates higher metabolism in embryos considered of superior quality or if the presence of higher reactive oxygen species (ROS) levels generated by the increased oxygen consumption triggers G6PD activation, the expression of genes related to stress response should be investigated in embryos produced by that technique. PMID:22908977

  16. uv excision-repair gene transfer in Chinese hamster ovary (CHO) cells

    SciTech Connect

    MacInnes, M.A.; Bingham, J.M.; Strniste, G.F.; Thompson, L.H.

    1983-01-01

    uvc-sensitive mutants of CHO cells provide a model system for molecular studies of DNA repair. We present our recent results which show that these mutants are competent recipients for plasmid marker gene transfer and incorporation of a putative CHO repair gene. The applicability and advantages of this system for interspecies human repair gene identification are discussed.

  17. Predicting passive and active tissue:plasma partition coefficients: interindividual and interspecies variability.

    PubMed

    Ruark, Christopher D; Hack, C Eric; Robinson, Peter J; Mahle, Deirdre A; Gearhart, Jeffery M

    2014-07-01

    A mechanistic tissue composition model incorporating passive and active transport for the prediction of steady-state tissue:plasma partition coefficients (K(t:pl)) of chemicals in multiple mammalian species was used to assess interindividual and interspecies variability. This approach predicts K(t:pl) using chemical lipophilicity, pKa, phospholipid membrane binding, and the unbound plasma fraction, together with tissue fractions of water, neutral lipids, neutral and acidic phospholipids, proteins, and pH. Active transport K(t:pl) is predicted using Michaelis-Menten transport parameters. Species-specific biological properties were identified from 126 peer reviewed journal articles, listed in the Supporting Information, for mouse, rat, guinea pig, rabbit, beagle dog, pig, monkey, and human species. Means and coefficients of variation for biological properties were used in a Monte Carlo analysis to assess variability. The results show K(t:pl) interspecies variability for the brain, fat, heart, kidney, liver, lung, muscle, red blood cell, skin, and spleen, but uncertainty in the estimates obscured some differences. Compounds undergoing active transport are shown to have concentration-dependent K(t:pl). This tissue composition-based mechanistic model can be used to predict K(t:pl) for organic chemicals across eight species and 10 tissues, and can be an important component in drug development when scaling K(t:pl) from animal models to humans.

  18. Morphological and molecular characterization of yellow oyster mushroom, Pleurotus citrinopileatus, hybrids obtained by interspecies mating.

    PubMed

    Rosnina, A G; Tan, Yee Shin; Abdullah, Noorlidah; Vikineswary, S

    2016-02-01

    Pleurotus citrinopileatus (yellow oyster mushroom) has an attractive shape and yellow colour but the fragile texture complicates packaging, and its strong aroma is unappealing to consumers. This study aimed to improve the characteristics and yield of P. citrinopileatus by interspecies mating between monokaryotic cultures of P. citrinopileatus and P. pulmonarius. Ten monokaryon cultures of the parental lines were crossed in all combinations to obtain hybrids. Eleven compatible mating pairs were obtained and cultivated to observe their sporophore morphology and yield. The selected hybrid, i.e. P1xC9, was beige in colour while hybrid P3xC8 was yellow in colour. Their sporophores had less offensive aroma, improved texture and higher yield. The DNA sequences of these hybrids were found to be in the same clade as the P. citrinopileatus parent with a bootstrap value of 99%. High bootstrap values indicate high genetic homology between hybrids and the P. citrinopileatus parent. The biological efficiencies of these hybrids P1xC9 (70.97%) and P3xC8 (52.14%) were also higher than the P. citrinopileatus parent (35.63%). Interspecies hybrids obtained by this mating technique can lead to better strains of mushrooms for genetic improvement of the Pleurotus species. PMID:26745978

  19. A RANKL mutant used as an inter-species vaccine for efficient immunotherapy of osteoporosis

    PubMed Central

    Liu, Changzhen; Zhao, Yunfeng; He, Wen; Wang, Wei; Chen, Yuan; Zhang, Shiqian; Ma, Yijing; Gohda, Jin; Ishida, Takaomi; Walter, Thomas S.; Owens, Raymond J.; Stuart, David I.; Ren, Jingshan; Gao, Bin

    2015-01-01

    Anti-cytokine therapeutic antibodies have been demonstrated to be effective in the treatment of several auto-immune disorders. However, The problems in antibody manufacture and the immunogenicity caused by multiple doses of antibodies inspire people to use auto-cytokine as immunogen to induce anti-cytokine antibodies. Nevertheless, the tolerance for inducing immune response against self-antigen has hindered the wide application of the strategy. To overcome the tolerance, here we proposed a strategy using the inter-species cytokine as immunogen for active immunization (TISCAI) to induce anti-cytokine antibody. As a proof of concept, an inter-species cytokine RANKL was successfully used as immunogen to induce anti-RANKL immune response. Furthermore, to prevent undesirable side-effects, the human RANKL was mutated based on the crystal structure of the complex of human RANKL and its rodent counterpart receptor RANK. We found, the antibodies produced blocked the osteoclast development in vitro and osteoporosis in OVX rat models. The results demonstrated this strategy adopted is very useful for general anti-cytokine immunotherapy for different diseases settings. PMID:26412210

  20. Interspecies competition triggers virulence and mutability in Candida albicans–Pseudomonas aeruginosa mixed biofilms

    PubMed Central

    Trejo-Hernández, Abigail; Andrade-Domínguez, Andrés; Hernández, Magdalena; Encarnación, Sergio

    2014-01-01

    Inter-kingdom and interspecies interactions are ubiquitous in nature and are important for the survival of species and ecological balance. The investigation of microbe-microbe interactions is essential for understanding the in vivo activities of commensal and pathogenic microorganisms. Candida albicans, a polymorphic fungus, and Pseudomonas aeruginosa, a Gram-negative bacterium, are two opportunistic pathogens that interact in various polymicrobial infections in humans. To determine how P. aeruginosa affects the physiology of C. albicans and vice versa, we compared the proteomes of each species in mixed biofilms versus single-species biofilms. In addition, extracellular proteins were analyzed. We observed that, in mixed biofilms, both species showed differential expression of virulence proteins, multidrug resistance-associated proteins, proteases and cell defense, stress and iron-regulated proteins. Furthermore, in mixed biofilms, both species displayed an increase in mutability compared with monospecific biofilms. This characteristic was correlated with the downregulation of enzymes conferring protection against DNA oxidation. In mixed biofilms, P. aeruginosa regulates its production of various molecules involved in quorum sensing and induces the production of virulence factors (pyoverdine, rhamnolipids and pyocyanin), which are major contributors to the ability of this bacterium to cause disease. Overall, our results indicate that interspecies competition between these opportunistic pathogens enhances the production of virulence factors and increases mutability and thus can alter the course of host-pathogen interactions in polymicrobial infections. PMID:24739628

  1. Automated aural classification used for inter-species discrimination of cetaceans.

    PubMed

    Binder, Carolyn M; Hines, Paul C

    2014-04-01

    Passive acoustic methods are in widespread use to detect and classify cetacean species; however, passive acoustic systems often suffer from large false detection rates resulting from numerous transient sources. To reduce the acoustic analyst workload, automatic recognition methods may be implemented in a two-stage process. First, a general automatic detector is implemented that produces many detections to ensure cetacean presence is noted. Then an automatic classifier is used to significantly reduce the number of false detections and classify the cetacean species. This process requires development of a robust classifier capable of performing inter-species classification. Because human analysts can aurally discriminate species, an automated aural classifier that uses perceptual signal features was tested on a cetacean data set. The classifier successfully discriminated between four species of cetaceans-bowhead, humpback, North Atlantic right, and sperm whales-with 85% accuracy. It also performed well (100% accuracy) for discriminating sperm whale clicks from right whale gunshots. An accuracy of 92% and area under the receiver operating characteristic curve of 0.97 were obtained for the relatively challenging bowhead and humpback recognition case. These results demonstrated that the perceptual features employed by the aural classifier provided powerful discrimination cues for inter-species classification of cetaceans.

  2. Development of quantitative interspecies toxicity relationship modeling of chemicals to fish.

    PubMed

    Fatemi, M H; Mousa Shahroudi, E; Amini, Z

    2015-09-01

    In this work, quantitative interspecies-toxicity relationship methodologies were used to improve the prediction power of interspecies toxicity model. The most relevant descriptors selected by stepwise multiple linear regressions and toxicity of chemical to Daphnia magna were used to predict the toxicities of chemicals to fish. Modeling methods that were used for developing linear and nonlinear models were multiple linear regression (MLR), random forest (RF), artificial neural network (ANN) and support vector machine (SVM). The obtained results indicate the superiority of SVM model over other models. Robustness and reliability of the constructed SVM model were evaluated by using the leave-one-out cross-validation method (Q(2)=0.69, SPRESS=0.822) and Y-randomization test (R(2)=0.268 for 30 trail). Furthermore, the chemical applicability domains of these models were determined via leverage approach. The developed SVM model was used for the prediction of toxicity of 46 compounds that their experimental toxicities to a fish were not being reported earlier from their toxicities to D. magna and relevant molecular descriptors.

  3. Automated aural classification used for inter-species discrimination of cetaceans.

    PubMed

    Binder, Carolyn M; Hines, Paul C

    2014-04-01

    Passive acoustic methods are in widespread use to detect and classify cetacean species; however, passive acoustic systems often suffer from large false detection rates resulting from numerous transient sources. To reduce the acoustic analyst workload, automatic recognition methods may be implemented in a two-stage process. First, a general automatic detector is implemented that produces many detections to ensure cetacean presence is noted. Then an automatic classifier is used to significantly reduce the number of false detections and classify the cetacean species. This process requires development of a robust classifier capable of performing inter-species classification. Because human analysts can aurally discriminate species, an automated aural classifier that uses perceptual signal features was tested on a cetacean data set. The classifier successfully discriminated between four species of cetaceans-bowhead, humpback, North Atlantic right, and sperm whales-with 85% accuracy. It also performed well (100% accuracy) for discriminating sperm whale clicks from right whale gunshots. An accuracy of 92% and area under the receiver operating characteristic curve of 0.97 were obtained for the relatively challenging bowhead and humpback recognition case. These results demonstrated that the perceptual features employed by the aural classifier provided powerful discrimination cues for inter-species classification of cetaceans. PMID:25235008

  4. Interspecies differences in DNA single strand breaks caused by benzo(a)pyrene and marine environment.

    PubMed

    Bihari, Nevenka; Fafandel, Maja

    2004-08-18

    The presence of DNA single strand breaks in untreated specimens of selected species, mosquito fish Gambusia affinis, painted comber Serranus scriba, blue mussel Mytilus galloprovincialis, spiny crab Maja crispata and sea cucumber Holothuria tubulosa as well as in 10 microg/g benzo(a)pyrene (BaP) treated mosquito fish, blue mussel and spiny crab was measured, using alkaline filter elution. Interspecies differences in alkaline elution profiles were observed and attributed to different lengths of DNA from different sources and to differences in the number of strand breaks present during normal cellular events in different phyla. Spiny crab hemocytes are more sensitive to action of BaP then blue mussel hemocytes and mosquito fish hepatocytes that could be explained by differences in the rates of distinct metabolic reactions and DNA repair among the investigated species. In field study, DNA single strand breaks were measured in hepatocytes of painted comber and in hemocytes of blue mussel and spiny crab from natural population specimens collected at eight sampling sites along Istrian coast, Croatia. Spatial variations in DNA integrity for each species were detected and revealed for the first time that spiny crab is responsive to different environmental conditions. Interspecies variations in the DNA integrity due to environmental conditions, confirmed species specific susceptibility to genotoxicity of certain environment that in long-term may modify the structure of marine communities. The multi-species approach in designing biomonitoring studies was suggested.

  5. Differential Transfer and Dissemination of Hypovirus and Nuclear and Mitochondrial Genomes of a Hypovirus-Infected Cryphonectria parasitica Strain after Introduction into a Natural Population

    PubMed Central

    Hoegger, Patrik J.; Heiniger, Ursula; Holdenrieder, Ottmar; Rigling, Daniel

    2003-01-01

    Biological control of plant diseases generally requires release of living organisms into the environment. Cryphonectria hypoviruses function as biological control agents for the chestnut blight fungus, Cryphonectria parasitica, and hypovirus-infected C. parasitica strains can be used to treat infected trees. We used naturally occurring molecular marker polymorphisms to examine the persistence and dissemination of the three genomes of a hypovirus-infected C. parasitica strain, namely, the double-stranded RNA genome of Cryphonectria hypovirus 1 (CHV1) and the nuclear and mitochondrial genomes of its fungal host. The hypovirus-infected strain was experimentally introduced into a blight-infested chestnut coppice forest by treating 73 of 246 chestnut blight cankers. Two years after introduction, the hypovirus had disseminated to 36% of the untreated cankers and to 35% of the newly established cankers. Spread of the hypovirus was more frequent within treated sprout clusters than between sprout clusters. Mitochondrial DNA of the introduced fungus also was transferred into the resident C. parasitica population. Concomitant transfer of both the introduced hypovirus and mitochondrial DNA was detected in almost one-half of the treated cankers analyzed. The introduced mitochondrial DNA haplotype also was found in three resident isolates from newly established cankers. The nuclear genome of the introduced strain persisted in the treated cankers but did not spread beyond them. PMID:12839742

  6. A Heat Transfer Model for a Stratified Corium-Metal Pool in the Lower Plenum of a Nuclear Reactor

    SciTech Connect

    M. S. Sohal; L. J. Siefken

    1999-08-01

    This preliminary design report describes a model for heat transfer in a corium-metal stratified pool. It was decided to make use of the existing COUPLE model. Currently available correlations for natural convection heat transfer in a pool with and without internal heat generation were obtained. The appropriate correlations will be incorporated in the existing COUPLE model. Heat conduction and solidification modeling will be done with existing algorithms in the COUPLE. Assessment of the new model will be done by simple energy conservation problems.

  7. A Heat Transfer Model for a Stratified Corium-metal Pool in the Lower Plenum of a Nuclear Reactor

    SciTech Connect

    Sohal, Manohar Singh; Siefken, Larry James

    1999-08-01

    This preliminary design report describes a model for heat transfer in a corium-metal stratified pool. It was decided to make use of the existing COUPLE model. Currently available correlations for natural convection heat transfer in a pool with and without internal heat generation were obtained. The appropriate correlations will be incorporated in the existing COUPLE model. Heat conduction and solidification modeling will be done with existing algorithms in the COUPLE. Assessment of the new model will be done by simple energy conservation problems.

  8. A protocol for adult somatic cell nuclear transfer in medaka fish (Oryzias latipes) with a high rate of viable clone formation.

    PubMed

    Bubenshchikova, Ekaterina; Kaftanovskaya, Elena; Adachi, Tomoko; Hashimoto, Hisashi; Kinoshita, Masato; Wakamatsu, Yuko

    2013-12-01

    Previously, we successfully generated fully grown, cloned medaka (the Japanese rice fish, Oryzias latipes) using donor nuclei from primary culture cells of adult caudal fin tissue and nonenucleated recipient eggs that were heat shock-treated to induce diploidization of the nuclei. However, the mechanism of clone formation using this method is unknown, and the rate of adult clone formation is not high enough for studies in basic and applied sciences. To gain insight into the mechanism and increase the success rate of this method of clone formation, we tested two distinct nuclear transfer protocols. In one protocol, the timing of transfer of donor nuclei was changed, and in the other, the size of the donor cells was changed; each protocol was based on our original methodology. Ultimately, we obtained an unexpectedly high rate of adult clone formation using the protocol that differed with respect to the timing of donor nuclei transfer. Specifically, 17% of the transplants that developed to the blastula stage ultimately developed into adult clones. The success rate with this method was 13 times higher than that obtained using the original method. Analyses focusing on the reasons for this high success rate of clone formation will help to elucidate the mechanism of clone formation that occurs with this method.

  9. Bone marrow mesenchymal stem cells are an attractive donor cell type for production of cloned pigs as well as genetically modified cloned pigs by somatic cell nuclear transfer.