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Sample records for adult cloned pig

  1. Proteomic analysis of pancreas derived from adult cloned pig

    SciTech Connect

    Chae, Jung-Il; Cho, Young Keun; Cho, Seong-Keun; Kim, Jin-Hoi; Han, Yong-Mahn; Koo, Deog-Bon Lee, Kyung-Kwang

    2008-02-08

    The potential medical applications of animal cloning include xenotransplantation, but the complex molecular cascades that control porcine organ development are not fully understood. Still, it has become apparent that organs derived from cloned pigs may be suitable for transplantation into humans. In this study, we examined the pancreas of an adult cloned pig developed through somatic cell nuclear transfer (SCNT) using two-dimensional electrophoresis (2-DE) and Western blotting. Proteomic analysis revealed 69 differentially regulated proteins, including such apoptosis-related species as annexins, lamins, and heat shock proteins, which were unanimously upregulated in the SCNT sample. Among the downregulated proteins in SCNT pancreas were peroxiredoxins and catalase. Western blot results indicate that several antioxidant enzymes and the anti-apoptotic protein were downregulated in SCNT pancreas, whereas several caspases were upregulated. Together, these data suggest that the accumulation of reactive oxygen species (ROS) in the pancreas of an adult cloned pig leads to apoptosis.

  2. Cloning. Pigs is pigs.

    PubMed

    Prather, R S

    2000-09-15

    Since the first report of a cloned animal (Dolly the sheep) 3 years ago, cloning mammals has become something of a cottage industry. As Prather discusses in his Perspective, pigs can now be added to the august list of cloned animals, which includes cows, goats, and mice. This is a particularly spectacular achievement because pig cloning has turned out to be notoriously difficult. The pig is also a valuable domestic animal to have cloned because, being physiologically close to humans, its organs can be used in xenotransplantation.

  3. Cloning. Pigs is pigs.

    PubMed

    Prather, R S

    2000-09-15

    Since the first report of a cloned animal (Dolly the sheep) 3 years ago, cloning mammals has become something of a cottage industry. As Prather discusses in his Perspective, pigs can now be added to the august list of cloned animals, which includes cows, goats, and mice. This is a particularly spectacular achievement because pig cloning has turned out to be notoriously difficult. The pig is also a valuable domestic animal to have cloned because, being physiologically close to humans, its organs can be used in xenotransplantation. PMID:11012362

  4. Phosphorylation of PrxII promotes JNK-dependent apoptosis in adult cloned pig kidney.

    PubMed

    Jeon, Young-Joo; Kim, Jumi; Lee, Dong-Seok; Shim, Jung-Hyun; Seo, Kang Seok; Chae, Jung-Il

    2014-08-01

    Organ transplantation is the most effective medical therapy for end-stage renal disease patients; however, there is a critical shortage of human donor organs. Therefore, xenotransplantation using genetically modified cloned porcine kidney is considered as a viable solution, but its fundamental therapeutic mechanism and difference from non-cloned porcine or human kidney for its clinical application is not well known. Here, we performed proteomic analysis to investigate the differentially expressed molecules in kidney tissue obtained from cloned porcine by SCNT, when compared with normal porcine kidney in same age as a control. A total of 80 protein spots were differentially expressed between cloned porcine kidney and control kidney, including apoptotic proteins, structural and anti-oxidant related proteins. Furthermore, very interestingly, the differential expression pattern of PrxII in the cloned porcine kidney was distinguishable from that in the control kidney in terms of the pI and molecular weight. Along with this, apoptotic marker proteins were up-regulated in the cloned porcine kidney. We suggested that these alterations were induced by post-translational modification such as phosphorylation in PrxII and could be mediated by JNK. With this result, we also observed that the down-regulation of JNK activity was caused by blockage of phosphorylation in PrxII T89A region. Taken together, cloned porcine kidney is more susceptible in JNK-induced apoptosis caused by PrxII phosphorylation, in oxidative stress condition. These results will be helpful in the application of cloned porcine xeno-transplants for treating end-stage renal disease patients in a clinical setting. PMID:24909612

  5. [Telomere lengthening by trichostatin A treatment in cloned pigs].

    PubMed

    Xie, Bing-Teng; Ji, Guang-Zhen; Kong, Qing-Ran; Mao, Jian; Shi, Yong-Qian; Liu, Shi-Chao; Wu, Mei-Ling; Wang, Juan; Liu, Lin; Liu, Zhong-Hua

    2012-12-01

    Telomeres are repeated GC rich sequences at the end of chromosomes, and shorten with each cell division due to DNA end replication problem. Previously, reprogrammed somatic cells of cloned animals display variable telomere elongation. However, it was reported that the cloned animals including Dolly do not reset telomeres and show premature aging. In this study, we investigated telomere function in cloned or transgenic cloned pigs, including the cloned Northeast Min pigs, eGFP, Mx, and PGC1α transgenic cloned pigs, and found that the telomere lengths of cloned pigs were significantly shorter than the nuclear donor adult fibroblasts and age-matched noncloned pigs (P<0.05), indicating that nuclear reprogramming did not restore cellular age of donor cells after somatic cell nuclear transfer (SCNT). Trichostatin A (TSA), an inhibitor of histone deacetylase, has proven to enhance the efficiency of nuclear reprogramming in several species. In order to test whether TSA also can effectively enhance reprogramming of telomeres, TSA (40 nmol/L) was used to treat porcine cloned embryos at 1-cell stage for 24 h. Consistent with previous reports, the developmental rate of SCNT embryos to the blastocyst stage was significantly increased compared with those of the control group (16.35% vs. 27.09%, 21.60% vs. 34.90%, P<0.05). Notably, the telomere length of cloned porcine blastocysts was also significantly elongated (P<0.05). Although TSA did not improve the cloning efficiency (1.3% vs. 1.7%, TSA vs. control), the telomere lengths of cloned pig-lets were significantly longer compared with those of the control group and the donor fibroblasts (P<0.05). In conclusion, telomeres have not been effectively restored by SCNT in pigs but TSA can effectively lengthen the telomere lengths of cloned pigs.

  6. [Telomere lengthening by trichostatin A treatment in cloned pigs].

    PubMed

    Xie, Bing-Teng; Ji, Guang-Zhen; Kong, Qing-Ran; Mao, Jian; Shi, Yong-Qian; Liu, Shi-Chao; Wu, Mei-Ling; Wang, Juan; Liu, Lin; Liu, Zhong-Hua

    2012-12-01

    Telomeres are repeated GC rich sequences at the end of chromosomes, and shorten with each cell division due to DNA end replication problem. Previously, reprogrammed somatic cells of cloned animals display variable telomere elongation. However, it was reported that the cloned animals including Dolly do not reset telomeres and show premature aging. In this study, we investigated telomere function in cloned or transgenic cloned pigs, including the cloned Northeast Min pigs, eGFP, Mx, and PGC1α transgenic cloned pigs, and found that the telomere lengths of cloned pigs were significantly shorter than the nuclear donor adult fibroblasts and age-matched noncloned pigs (P<0.05), indicating that nuclear reprogramming did not restore cellular age of donor cells after somatic cell nuclear transfer (SCNT). Trichostatin A (TSA), an inhibitor of histone deacetylase, has proven to enhance the efficiency of nuclear reprogramming in several species. In order to test whether TSA also can effectively enhance reprogramming of telomeres, TSA (40 nmol/L) was used to treat porcine cloned embryos at 1-cell stage for 24 h. Consistent with previous reports, the developmental rate of SCNT embryos to the blastocyst stage was significantly increased compared with those of the control group (16.35% vs. 27.09%, 21.60% vs. 34.90%, P<0.05). Notably, the telomere length of cloned porcine blastocysts was also significantly elongated (P<0.05). Although TSA did not improve the cloning efficiency (1.3% vs. 1.7%, TSA vs. control), the telomere lengths of cloned pig-lets were significantly longer compared with those of the control group and the donor fibroblasts (P<0.05). In conclusion, telomeres have not been effectively restored by SCNT in pigs but TSA can effectively lengthen the telomere lengths of cloned pigs. PMID:23262106

  7. Cloning pigs: advances and applications.

    PubMed

    Polejaeva, I A

    2001-01-01

    Although mouse embryonic stem cells have been used widely for over a decade as an important tool for introducing precise genetic modification into the genome, demonstrating the great value of this technology in a range of biomedical applications, similar technology does not exist for domestic animals. However, the development of somatic cell nuclear transfer has bypassed the need for embryonic stem cells from livestock. The production of offspring from differentiated cell nuclei provides information and opportunities in a number of areas including cellular differentiation, early development and ageing. However, the primary significance of cloning is probably in the opportunities that this technology brings to genetic manipulation. Potential applications of gene targeting in livestock species are described with particular emphasis on the generation of pigs that can be used for xenotransplantation, and the production of improved models for human physiology and disease. The development of techniques for somatic cell nuclear transfer in pigs and the challenges associated with this technology are also reviewed.

  8. Effects of donor fibroblast cell type and transferred cloned embryo number on the efficiency of pig cloning.

    PubMed

    Li, Zicong; Shi, Junsong; Liu, Dewu; Zhou, Rong; Zeng, Haiyu; Zhou, Xiu; Mai, Ranbiao; Zeng, Shaofen; Luo, Lvhua; Yu, Wanxian; Zhang, Shouquan; Wu, Zhenfang

    2013-02-01

    Currently, cloning efficiency in pigs is very low. Donor cell type and number of cloned embryos transferred to an individual surrogate are two major factors that affect the successful rate of somatic cell nuclear transfer (SCNT) in pigs. This study aimed to compare the influence of different donor fibroblast cell types and different transferred embryo numbers on recipients' pregnancy rate and delivery rate, the average number of total clones born, clones born alive and clones born healthy per litter, and the birth rate of healthy clones (=total number of healthy cloned piglets born /total number of transferred cloned embryos). Three types of donor fibroblasts were tested in large-scale production of cloned pigs, including fetal fibroblasts (FFBs) from four genetically similar Western swine breeds of Pietrain (P), Duroc (D), Landrace (L), and Yorkshire (Y), which are referred to as P,D,LY-FFBs, adult fibroblasts (AFBs) from the same four breeds, which are designated P,D,L,Y-AFBs, and AFBs from a Chinese pig breed of Laiwu (LW), which is referred to as LW-AFBs. Within each donor fibroblast cell type group, five transferred cloned embryo number groups were tested. In each embryo number group, 150-199, 200-249, 250-299, 300-349, or 350-450 cloned embryos were transferred to each individual recipient sow. For the entire experiment, 92,005 cloned embryos were generated from nearly 115,000 matured oocytes and transferred to 328 recipients; in total, 488 cloned piglets were produced. The results showed that the mean clones born healthy per litter resulted from transfer of embryos cloned from LW-AFBs (2.53 ± 0.34) was similar with that associated with P,D,L,Y-FFBs (2.72 ± 0.29), but was significantly higher than that resulted from P,D,L,Y-AFBs (1.47 ± 0.18). Use of LW-AFBs as donor cells for SCNT resulted in a significantly higher pregnancy rate (72.00% vs. 59.30% and 48.11%) and delivery rate (60.00% vs. 45.93% and 35.85%) for cloned embryo recipients, and a

  9. Application of genetically modified and cloned pigs in translational research.

    PubMed

    Matsunari, Hitomi; Nagashima, Hiroshi

    2009-06-01

    Pigs are increasingly being recognized as good large-animal models for translational research, linking basic science to clinical applications in order to establish novel therapeutics. This article reviews the current status and future prospects of genetically modified and cloned pigs in translational studies. It also highlights pigs specially designed as disease models, for xenotransplantation or to carry cell marker genes. Finally, use of porcine somatic stem and progenitor cells in preclinical studies of cell transplantation therapy is also discussed.

  10. Cloning and expression characteristics of the pig Stra8 gene.

    PubMed

    Wang, Xiaoyan; Chen, Tingfeng; Song, Chengyi; Gao, Bo; Zhang, Yani

    2014-07-15

    Stra8 (Stimulated by Retinoic Acid 8) is considered a meiotic gatekeeper gene. Using reverse transcriptase PCR and rapid amplification of cDNA ends (RACE), the complete sequence of the pig Stra8 gene was cloned. Bioinformatics analyses of this sequence were performed. Using semi-quantitative methods, the expression characteristics of Stra8 in Testis, cauda epididymis, body epididymis, caput epididymis, seminal vesicles, prostate gland, Cowper's gland, heart, liver, spleen, lung, kidney, stomach, hypothalamus, pituitary gland, cerebrum, cerebellum, and hippocampus of adult Meishan boar and sow tissues were examined. The expression pattern in the testis of 2-, 30-, 60-, 90-, and 150-day old Meishan boars were analyzed using real-time PCR. We constructed a eukaryotic expression vector for the Stra8 gene and used it to transfect NIH-3T3 cells and third generation pig spermatogonial stem cells (SSCs) cultured in vitro. Testes weight and sperm count in the cauda epididymis were evaluated at various time points. The results showed that the length of the pig Stra8 gene cDNA was 1444 bp encoding 366 amino acids with one typical helix-loop-helix (HLH) domain. It is testes-specific expression. Expression was first detected in boar testis starting at day 2, and its expression significantly (p<0.05) increased with age and body weight. When NIH-3T3 cells and pig SSCs were transfected with the eukaryotic expression vector EGFP (enhanced green fluorescent protein)-N1-pStra8, it was expressed in the cytoplasm of NIH-3T3 cells. However, in SSCs, Stra8 was expressed predominantly in cytoplasm and few in nucleus. Our data suggest that perhaps Stra8 acts as a transcription factor to initiate meiosis in young boar.

  11. Cloning and Expression Characteristics of the Pig Stra8 Gene

    PubMed Central

    Wang, Xiaoyan; Chen, Tingfeng; Song, Chengyi; Gao, Bo; Zhang, Yani

    2014-01-01

    Stra8 (Stimulated by Retinoic Acid 8) is considered a meiotic gatekeeper gene. Using reverse transcriptase PCR and rapid amplification of cDNA ends (RACE), the complete sequence of the pig Stra8 gene was cloned. Bioinformatics analyses of this sequence were performed. Using semi-quantitative methods, the expression characteristics of Stra8 in Testis, cauda epididymis, body epididymis, caput epididymis, seminal vesicles, prostate gland, Cowper’s gland, heart, liver, spleen, lung, kidney, stomach, hypothalamus, pituitary gland, cerebrum, cerebellum, and hippocampus of adult Meishan boar and sow tissues were examined. The expression pattern in the testis of 2-, 30-, 60-, 90-, and 150-day old Meishan boars were analyzed using real-time PCR. We constructed a eukaryotic expression vector for the Stra8 gene and used it to transfect NIH-3T3 cells and third generation pig spermatogonial stem cells (SSCs) cultured in vitro. Testes weight and sperm count in the cauda epididymis were evaluated at various time points. The results showed that the length of the pig Stra8 gene cDNA was 1444 bp encoding 366 amino acids with one typical helix-loop-helix (HLH) domain. It is testes-specific expression. Expression was first detected in boar testis starting at day 2, and its expression significantly (p < 0.05) increased with age and body weight. When NIH-3T3 cells and pig SSCs were transfected with the eukaryotic expression vector EGFP (enhanced green fluorescent protein)-N1-pStra8, it was expressed in the cytoplasm of NIH-3T3 cells. However, in SSCs, Stra8 was expressed predominantly in cytoplasm and few in nucleus. Our data suggest that perhaps Stra8 acts as a transcription factor to initiate meiosis in young boar. PMID:25029539

  12. Transplantation and differentiation of donor cells in the cloned pigs

    SciTech Connect

    Shimada, Arata; Tomii, Ryo; Kano, Koichiro; Nagashima, Hiroshi . E-mail: hnagas@isc.meiji.ac.jp

    2006-06-02

    The application of nuclear transfer technology is an interesting approach to investigate stem and progenitor cell transplantation therapy. If stem cells are used as a nuclear donor, donor cells can engraft into cloned animals without histocompatible problems. However, it is still uncertain whether donor cells can engraft to cloned animal and differentiate in vivo. To address this problem, we transplanted donor cells to dermal tissues of cloned pigs developed by using preadipocytes as donor cells. Preadipocytes are adipocytic progenitor which can differentiate to mature adipocytes in vitro. We showed that the donor preadipocytes were successfully transplanted into the cloned pigs without immune rejection and they differentiated into mature adipocytes in vivo 3 weeks after transplantation. In contrast, allogenic control preadipocytes, which can differentiate in vitro, did not differentiate in vivo. These results indicate that donor progenitor cells can differentiate in cloned animal.

  13. Pre-weaning performance and health of pigs born to cloned (fetal cell derived) swine versus non-cloned swine.

    PubMed

    Martin, M; Adams, C; Wiseman, B

    2004-07-01

    The objective of this study was to compare the pre-weaning performance of pigs derived from cloned versus non-cloned parents. Five cloned gilts and one cloned boar were used to produce five litters of pigs. One of five cloned females and the cloned boar were derived from two genetically unmanipulated fetal fibroblast cell lines. The remaining female clones were derived from a fetal fibroblast cell line in which random insertion of a alpha-1,3-galactosyltransferase gene targeting construct had occurred. Fetal cell lines had similar genetic backgrounds and were derived from three different fetuses in three different litters. Five litters of pigs were also generated from matings between two non-cloned boars and five non-cloned gilts. The mean gestation length, mean litter size, mean birth and weaning weights for male and female pigs were similar for litters derived from cloned parents versus non-cloned parents. The proportions of pigs born live and pigs that survived to weaning were also similar for pigs born to cloned as compared to non-cloned parents. In summary, matings between cloned swine derived from fetal fibroblast cell lines yielded litters of pigs that were similar in the number born, piglet birth weight and perinatal and pre-weaning mortality to litters produced by non-cloned swine.

  14. Advancing pig cloning technologies towards application in regenerative medicine.

    PubMed

    Nagashima, H; Matsunari, H; Nakano, K; Watanabe, M; Umeyama, K; Nagaya, M

    2012-08-01

    Regenerative medicine is expected to make a significant contribution by development of novel therapeutic treatments for intractable diseases and for improving the quality of life of patients. Many advances in regenerative medicine, including basic and translational research, have been developed and tested in experimental animals; pigs have played an important role in various aspects of this work. The value of pigs as a model species is being enhanced by the generation of specially designed animals through cloning and genetic modifications, enabling more sophisticated research to be performed and thus accelerating the clinical application of regenerative medicine. This article reviews the significant aspects of the creation and application of cloned and genetically modified pigs in regenerative medicine research and considers the possible future directions of the technology. We also discuss the importance of reproductive biology as an interface between basic science and clinical medicine.

  15. Molecular cloning and expression analysis of pig CD138.

    PubMed

    Bae, Joonbeom; Jeong, Seonah; Lee, Ju Yeon; Lee, Hyun-Jeong; Choi, Bong-Hwan; Kim, Ji-Eun; Choi, Inho; Chun, Taehoon

    2013-12-01

    CD138 (syndecan-1) interacts with various components of the extracellular matrix and associates with the actin cytoskeleton. In this study, we cloned pig CD138 cDNA and determined its complete cDNA sequence. Pig CD138 cDNA contained an open reading frame (930 bp) encoding 309 amino acids with five well conserved putative glycosaminoglycan attachment sites, a putative cleavage site for matrix metalloproteinases, and conserved motifs involved in signal transduction among mammalian species. Pig CD138 mRNA was detected in various tissues, including lymphoid and non-lymphoid organs, indicating the multicellular functions of CD138 in pigs. Western blot and flow cytometry analyses detected an approximate 35 kDa pig CD138 protein expressed on the cell surface. Further immunohistochemistry analysis revealed that CD138 expression was mainly observed in submucosa and lamina propria of the pig small intestine. Further study will be necessary to define the functional importance of CD138 during specific infectious diseases in pigs. PMID:24128845

  16. 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.

    PubMed

    Li, Zicong; He, Xiaoyan; Chen, Liwen; Shi, Junsong; Zhou, Rong; Xu, Weihua; Liu, Dewu; Wu, Zhenfang

    2013-10-01

    The somatic cell nuclear transfer (SCNT) technique has been widely applied to clone pigs or to produce genetically modified pigs. Currently, this technique relies mainly on using terminally differentiated fibroblasts as donor cells. To improve cloning efficiency, only partially differentiated multipotent mesenchymal stem cells (MSCs), thought to be more easily reprogrammed to a pluripotent state, have been used as nuclear donors in pig SCNT. Although in vitro-cultured embryos cloned from porcine MSCs (MSCs-embryos) were shown to have higher preimplantation developmental ability than cloned embryos reconstructed from fibroblasts (Fs-embryos), the difference in in vivo full-term developmental rate between porcine MSCs-embryos and Fs-embryos has not been investigated so far. In this study, we demonstrated that blastocyst total cell number and full-term survival abilities of MSCs-embryos were significantly higher than those of Fs-embryos cloned from the same donor pig. The enhanced developmental potential of MSCs-embryos may be associated with their nuclear donors' DNA methylation profile, because we found that the methylation level of imprinting genes and repeat sequences differed between MSCs and fibroblasts. In addition, we showed that use of transgenic porcine MSCs generated from transgene plasmid transfection as donor cells for SCNT can produce live transgenic cloned pigs. These results strongly suggest that porcine bone marrow MSCs are a desirable donor cell type for production of cloned pigs and genetically modified cloned pigs via SCNT.

  17. 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

  18. Embryo Aggregation in Pig Improves Cloning Efficiency and Embryo Quality.

    PubMed

    Buemo, Carla Paola; Gambini, Andrés; Moro, Lucia Natalia; Hiriart, María Inés; Fernández-Martín, Rafael; Collas, Philippe; Salamone, Daniel Felipe

    2016-01-01

    In this study, we analyzed the effects of the cloned embryo aggregation on in vitro embryo development and embryo quality by measuring blastocyst diameter and cell number, DNA fragmentation levels and the expression of genes associated with pluripotency, apoptosis, trophoblast and DNA methylation in the porcine. Zona-free reconstructed cloned embryos were cultured in the well of the well system, placing one (1x non aggregated group) or three (3x group) embryos per microwell. Our results showed that aggregation of three embryos increased blastocyst formation rate and blastocyst diameter of cloned pig embryos. DNA fragmentation levels in 3x aggregated cloned blastocysts were significantly decreased compared to 1x blastocysts. Levels of Oct4, Klf4, Igf2, Bax and Dnmt 1 transcripts were significantly higher in aggregated embryos, whereas Nanog levels were not affected. Transcripts of Cdx2 and Bcl-xl were essentially non-detectable. Our study suggests that embryo aggregation in the porcine may be beneficial for cloned embryo development and embryo quality, through a reduction in apoptotic levels and an improvement in cell reprogramming.

  19. Embryo Aggregation in Pig Improves Cloning Efficiency and Embryo Quality

    PubMed Central

    Buemo, Carla Paola; Gambini, Andrés; Moro, Lucia Natalia; Hiriart, María Inés; Fernández-Martín, Rafael; Collas, Philippe; Salamone, Daniel Felipe

    2016-01-01

    In this study, we analyzed the effects of the cloned embryo aggregation on in vitro embryo development and embryo quality by measuring blastocyst diameter and cell number, DNA fragmentation levels and the expression of genes associated with pluripotency, apoptosis, trophoblast and DNA methylation in the porcine. Zona-free reconstructed cloned embryos were cultured in the well of the well system, placing one (1x non aggregated group) or three (3x group) embryos per microwell. Our results showed that aggregation of three embryos increased blastocyst formation rate and blastocyst diameter of cloned pig embryos. DNA fragmentation levels in 3x aggregated cloned blastocysts were significantly decreased compared to 1x blastocysts. Levels of Oct4, Klf4, Igf2, Bax and Dnmt 1 transcripts were significantly higher in aggregated embryos, whereas Nanog levels were not affected. Transcripts of Cdx2 and Bcl-xl were essentially non-detectable. Our study suggests that embryo aggregation in the porcine may be beneficial for cloned embryo development and embryo quality, through a reduction in apoptotic levels and an improvement in cell reprogramming. PMID:26894831

  20. Embryo Aggregation in Pig Improves Cloning Efficiency and Embryo Quality.

    PubMed

    Buemo, Carla Paola; Gambini, Andrés; Moro, Lucia Natalia; Hiriart, María Inés; Fernández-Martín, Rafael; Collas, Philippe; Salamone, Daniel Felipe

    2016-01-01

    In this study, we analyzed the effects of the cloned embryo aggregation on in vitro embryo development and embryo quality by measuring blastocyst diameter and cell number, DNA fragmentation levels and the expression of genes associated with pluripotency, apoptosis, trophoblast and DNA methylation in the porcine. Zona-free reconstructed cloned embryos were cultured in the well of the well system, placing one (1x non aggregated group) or three (3x group) embryos per microwell. Our results showed that aggregation of three embryos increased blastocyst formation rate and blastocyst diameter of cloned pig embryos. DNA fragmentation levels in 3x aggregated cloned blastocysts were significantly decreased compared to 1x blastocysts. Levels of Oct4, Klf4, Igf2, Bax and Dnmt 1 transcripts were significantly higher in aggregated embryos, whereas Nanog levels were not affected. Transcripts of Cdx2 and Bcl-xl were essentially non-detectable. Our study suggests that embryo aggregation in the porcine may be beneficial for cloned embryo development and embryo quality, through a reduction in apoptotic levels and an improvement in cell reprogramming. PMID:26894831

  1. Reproductive and growth performance in Jin Hua pigs cloned from somatic cell nuclei and the meat quality of their offspring.

    PubMed

    Shibata, Masatoshi; Otake, Masayoshi; Tsuchiya, Seiko; Chikyu, Mikio; Horiuchi, Atsushi; Kawarasaki, Tatsuo

    2006-10-01

    Somatic cell cloning is expected to be a valuable method for conserving genetic resources in pigs. In this study, we compared the reproductive and growth performance of Jin Hua cloned pigs with that of naturally bred Jin Hua pigs. In addition, we generated offspring from the cloned sows and examined the productivity and quality of meat in the progeny. The birth weights and growth rates of somatic cell-cloned pigs were similar to those of Jin Hua pigs. The cloned pigs reached puberty very early, and this is typical of the Jin Hua breed. Furthermore, reproductive performance, in terms of traits such as gestation period, litter size, and raising rate in the cloned pigs were similar to Jin Hua pigs. Although the offspring of the cloned (OC) pigs had lower birth weights than the Jin Hua breed, the daily weight gain of the OC pigs was significantly higher, especially at the finishing stage. The carcass quality of the OC pigs had similar characteristics to the Jin Hua breed, namely thick back fat and a small loin area. Furthermore, the meat qualities of the OC pigs were similar to those of Jin Hua pigs in terms of intramuscular fat content and tenderness. These results demonstrate that cloned pigs and their offspring were similar to the Jin Hua breed in most of the growth, reproductive, and meat productive performances. This strongly suggests that pigs cloned from somatic cell nuclei have the potential to be a valuable genetic resource for breeding.

  2. A transgenic-cloned pig model expressing non-fluorescent modified Plum

    PubMed Central

    NAGAYA, Masaki; WATANABE, Masahito; KOBAYASHI, Mirina; NAKANO, Kazuaki; ARAI, Yoshikazu; ASANO, Yoshinori; TAKEISHI, Toki; UMEKI, Ikuma; FUKUDA, Tooru; YASHIMA, Sayaka; TAKAYANAGI, Shuko; WATANABE, Nobuyuki; ONODERA, Masafumi; MATSUNARI, Hitomi; UMEYAMA, Kazuhiro; NAGASHIMA, Hiroshi

    2016-01-01

    Genetically modified pigs that express fluorescent proteins such as green and red fluorescent proteins have become indispensable biomedical research tools in recent years. Cell or tissue transplantation studies using fluorescent markers should be conducted, wherein the xeno-antigenicity of the fluorescent proteins does not affect engraftment or graft survival. Thus, we aimed to create a transgenic (Tg)-cloned pig that was immunologically tolerant to fluorescent protein antigens. In the present study, we generated a Tg-cloned pig harboring a derivative of Plum modified by a single amino acid substitution in the chromophore. The cells and tissues of this Tg-cloned pig expressing the modified Plum (mPlum) did not fluoresce. However, western blot and immunohistochemistry analyses clearly showed that the mPlum had the same antigenicity as Plum. Thus, we have obtained primary proof of principle for creating a cloned pig that is immunologically tolerant to fluorescent protein antigens. PMID:27396383

  3. Mitochondrial DNA heteroplasmy in cloned cattle produced by fetal and adult cell cloning.

    PubMed

    Steinborn, R; Schinogl, P; Zakhartchenko, V; Achmann, R; Schernthaner, W; Stojkovic, M; Wolf, E; Müller, M; Brem, G

    2000-07-01

    Mammals have been cloned from adult donor cells. Here we report the first cases of mitochondrial DNA (mtDNA) heteroplasmy in adult mammalian clones generated from fetal and adult donor cells. The heteroplasmic clones included a healthy cattle equivalent of the sheep Dolly, for which a lack of heteroplasmy was reported.

  4. Mitochondrial DNA heteroplasmy in cloned cattle produced by fetal and adult cell cloning.

    PubMed

    Steinborn, R; Schinogl, P; Zakhartchenko, V; Achmann, R; Schernthaner, W; Stojkovic, M; Wolf, E; Müller, M; Brem, G

    2000-07-01

    Mammals have been cloned from adult donor cells. Here we report the first cases of mitochondrial DNA (mtDNA) heteroplasmy in adult mammalian clones generated from fetal and adult donor cells. The heteroplasmic clones included a healthy cattle equivalent of the sheep Dolly, for which a lack of heteroplasmy was reported. PMID:10888867

  5. Ovulation Statuses of Surrogate Gilts Are Associated with the Efficiency of Excellent Pig Cloning.

    PubMed

    Huan, Yanjun; Hu, Kui; Xie, Bingteng; Shi, Yongqian; Wang, Feng; Zhou, Yang; Liu, Shichao; Huang, Bo; Zhu, Jiang; Liu, Zhongfeng; He, Yilong; Li, Jingyu; Kong, Qingran; Liu, Zhonghua

    2015-01-01

    Somatic cell nuclear transfer (SCNT) is an assisted reproductive technique that can produce multiple copies of excellent livestock. However, low cloning efficiency limits the application of SCNT. In this study, we systematically investigated the major influencing factors related to the overall cloning efficiency in pigs. Here, 13620 cloned embryos derived from excellent pigs were transferred into 79 surrogate gilts, and 119 live cloned piglets were eventually generated. During cloning, group of cloned embryos derived from excellent Landrace or Large white pigs presented no significant differences of cleavage and blastocyst rates, blastocyst cell numbers, surrogate pregnancy and delivery rates, average numbers of piglets born and alive and cloning efficiencies, and group of 101-150, 151-200 or 201-250 cloned embryos transferred per surrogate also displayed a similar developmental efficiency. When estrus stage of surrogate gilts was compared, group of embryo transfer on Day 2 of estrus showed significantly higher pregnancy rate, delivery rate, average number of piglets born, average alive piglet number or cloning efficiency than group on Day 1, Day 3, Day 4 or Day 5, respectively (P<0.05). And, in comparison with the preovulation and postovulation groups, group of surrogate gilts during periovulation displayed a significantly higher overall cloning efficiency (P<0.05). Further investigation of surrogate estrus stage and ovulation status displayed that ovulation status was the real factor underlying estrus stage to determine the overall cloning efficiency. And more, follicle puncture for preovulation, not transfer position shallowed for preovulation or deepened for postovulation, significantly improved the average number of piglets alive and cloning efficiency (P<0.05). In conclusion, our results demonstrated that ovulation status of surrogate gilts was the fundamental factor determining the overall cloning efficiency of excellent pigs, and follicle puncture, not

  6. Ovulation Statuses of Surrogate Gilts Are Associated with the Efficiency of Excellent Pig Cloning

    PubMed Central

    Huan, Yanjun; Hu, Kui; Xie, Bingteng; Shi, Yongqian; Wang, Feng; Zhou, Yang; Liu, Shichao; Huang, Bo; Zhu, Jiang; Liu, Zhongfeng; He, Yilong; Li, Jingyu; Kong, Qingran; Liu, Zhonghua

    2015-01-01

    Somatic cell nuclear transfer (SCNT) is an assisted reproductive technique that can produce multiple copies of excellent livestock. However, low cloning efficiency limits the application of SCNT. In this study, we systematically investigated the major influencing factors related to the overall cloning efficiency in pigs. Here, 13620 cloned embryos derived from excellent pigs were transferred into 79 surrogate gilts, and 119 live cloned piglets were eventually generated. During cloning, group of cloned embryos derived from excellent Landrace or Large white pigs presented no significant differences of cleavage and blastocyst rates, blastocyst cell numbers, surrogate pregnancy and delivery rates, average numbers of piglets born and alive and cloning efficiencies, and group of 101–150, 151–200 or 201–250 cloned embryos transferred per surrogate also displayed a similar developmental efficiency. When estrus stage of surrogate gilts was compared, group of embryo transfer on Day 2 of estrus showed significantly higher pregnancy rate, delivery rate, average number of piglets born, average alive piglet number or cloning efficiency than group on Day 1, Day 3, Day 4 or Day 5, respectively (P<0.05). And, in comparison with the preovulation and postovulation groups, group of surrogate gilts during periovulation displayed a significantly higher overall cloning efficiency (P<0.05). Further investigation of surrogate estrus stage and ovulation status displayed that ovulation status was the real factor underlying estrus stage to determine the overall cloning efficiency. And more, follicle puncture for preovulation, not transfer position shallowed for preovulation or deepened for postovulation, significantly improved the average number of piglets alive and cloning efficiency (P<0.05). In conclusion, our results demonstrated that ovulation status of surrogate gilts was the fundamental factor determining the overall cloning efficiency of excellent pigs, and follicle puncture

  7. 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

  8. Cloning changes the response to obesity of innate immune factors in blood, liver, and adipose tissues in domestic pigs.

    PubMed

    Rødgaard, Tina; Skovgaard, Kerstin; Stagsted, Jan; Heegaard, Peter M H

    2013-06-01

    The objective of this study was to evaluate the usefulness of cloned pigs as porcine obesity models reflecting obesity-associated changes in innate immune factor gene expression profiles. Liver and adipose tissue expression of 43 innate immune genes as well as serum concentrations of six immune factors were analyzed in lean and diet-induced obese cloned domestic pigs and compared to normal domestic pigs (obese and lean). The number of genes affected by obesity was lower in cloned animals than in control animals. All genes affected by obesity in adipose tissues of clones were downregulated; both upregulation and downregulation were observed in the controls. Cloning resulted in a less differentiated adipose tissue expression pattern. Finally, the serum concentrations of two acute-phase proteins (APPs), haptoglobin (HP) and orosomucoid (ORM), were increased in obese clones as compared to obese controls as well as lean clones and controls. Generally, the variation in phenotype between individual pigs was not reduced in cloned siblings as compared to normal siblings. Therefore, we conclude that cloning limits both the number of genes responding to obesity as well as the degree of tissue-differentiated gene expression, concomitantly with an increase in APP serum concentrations only seen in cloned, obese pigs. This may suggest that the APP response seen in obese, cloned pigs is a consequence of the characteristic skewed gene response to obesity in cloned pigs, as described in this work. This should be taken into consideration when using cloned animals as models for innate responses to obesity.

  9. 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

  10. Generation of cloned transgenic pigs rich in omega-3 fatty acids.

    PubMed

    Lai, Liangxue; Kang, Jing X; Li, Rongfeng; Wang, Jingdong; Witt, William T; Yong, Hwan Yul; Hao, Yanhong; Wax, David M; Murphy, Clifton N; Rieke, August; Samuel, Melissa; Linville, Michael L; Korte, Scott W; Evans, Rhobert W; Starzl, Thomas E; Prather, Randall S; Dai, Yifan

    2006-04-01

    Meat products are generally low in omega-3 (n-3) fatty acids, which are beneficial to human health. We describe the generation of cloned pigs that express a humanized Caenorhabditis elegans gene, fat-1, encoding an n-3 fatty acid desaturase. The hfat-1 transgenic pigs produce high levels of n-3 fatty acids from n-6 analogs, and their tissues have a significantly reduced ratio of n-6/n-3 fatty acids (P < 0.001).

  11. Influence of embryo handling and transfer method on pig cloning efficiency.

    PubMed

    Shi, Junsong; Zhou, Rong; Luo, Lvhua; Mai, Ranbiao; Zeng, Haiyu; He, Xiaoyan; Liu, Dewu; Zeng, Fang; Cai, Gengyuan; Ji, Hongmei; Tang, Fei; Wang, Qinglai; Wu, Zhenfang; Li, Zicong

    2015-03-01

    The somatic cell nuclear transfer (SCNT) technique could be used to produce genetically superior or genetically engineered cloned pigs that have wide application in agriculture and bioscience research. However, the efficiency of porcine SCNT currently is very low. Embryo transfer (ET) is a key step for the success of SCNT. In this study, the effects of several ET-related factors, including cloned embryo culture time, recipient's ovulation status, co-transferred helper embryos and ET position, on the success rate of pig cloning were investigated. The results indicated that transfer of cloned embryos cultured for a longer time (22-24h vs. 4-6h) into pre-ovulatory sows decreased recipient's pregnancy rate and farrowing rate, and use of pre-ovulatory and post-ovulatory sows as recipients for SCNT embryos cultured for 22-24h resulted in a similar porcine SCNT efficiency. Use of insemination-produced in vivo fertilized, parthenogenetically activated and in vitro fertilized embryos as helper embryos to establish and/or maintain pregnancy of SCNT embryos recipients could not improve the success rate of porcine SCNT. Transfer of cloned embryos into double oviducts of surrogates significantly increased pregnancy rate as well as farrowing rate of recipients, and the developmental rate of transferred cloned embryos, as compared to unilateral oviduct transfer. This study provided useful information for optimization of the embryo handling and transfer protocol, which will help to improve the ability to generate cloned pigs.

  12. Valproic acid improved in vitro development of pig cloning embryos but did not improve survival of cloned pigs to adulthood.

    PubMed

    Kang, Jin-Dan; Li, Suo; Lu, Yue; Wang, Wei; Liang, Shuang; Liu, Xi; Jin, Jun-Xue; Hong, Yu; Yan, Chang-Guo; Yin, Xi-Jun

    2013-01-15

    The objective was to examine the effects of valproic acid (VPA), a histone deacetylase inhibitor, on in vitro and in vivo development of Wuzhishan miniature pig somatic cell nuclear transfer (SCNT) embryos. Experiment 1 compared in vitro developmental competence of nuclear transfer embryos treated with various concentrations of VPA for 24 h. Embryos treated with 2 mM VPA for 24 h had a greater rate of blastocyst formation compared with control or embryos treated with 4 or 8 mM VPA (21.5% vs. 10.5%, 12.6%, and 17.2%, P < 0.05). Experiment 2 examined the in vitro developmental competence of nuclear transfer embryos treated with 2 mM VPA for various intervals after chemical activation. Embryos treated for 24 h had higher rates of blastocyst formation than the control or those treated for 4 or 48 h (20.7% vs. 9.2%, 12.1%, and 9.1%, P < 0.05). In Experiment 3, an average of 207 (range, 192-216) nuclear transfer embryos from the VPA-treated group were transferred to surrogate mothers, resulting in three pregnancies. Two of the surrogates delivered a total of 11 live piglets. However, for unknown reasons, nine of 11 piglets in the VPA-treated group died within 1 to 5 d after birth. Untreated control embryos (average, 205; range, 179-225) transferred to four surrogate mothers resulted in three pregnancies, two of which delivered a total of 12 live offspring, although four of 12 piglets in the VPA-untreated group died (cause unknown) within 1 to 3 d, whereas eight of the 12 piglets in the VPA-untreated group survived more than 3 or 4 mo. The average birth weight of the two litters from the VPA-treated group tended (P < 0.05) to be lower than that from the control groups (551.6 g vs. 675.2 g). In conclusion, VPA treatment increased the blastocyst formation rate of SCNT porcine embryos; both VPA-treated and the untreated clones developed to term, but offspring from VPA-treated embryos had a lower survival to adulthood than those from control embryos (18.2% vs. 67.0%; P < 0.05).

  13. 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.

  14. 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

  15. Comparison of Gene Expression and Genome-Wide DNA Methylation Profiling between Phenotypically Normal Cloned Pigs and Conventionally Bred Controls

    PubMed Central

    Li, Shengting; Li, Jian; Lin, Lin; Nielsen, Anders Lade; Sørensen, Charlotte Brandt; Vajta, Gábor; Wang, Jun; Zhang, Xiuqing; Du, Yutao; Yang, Huanming; Bolund, Lars

    2011-01-01

    Animal breeding via Somatic Cell Nuclear Transfer (SCNT) has enormous potential in agriculture and biomedicine. However, concerns about whether SCNT animals are as healthy or epigenetically normal as conventionally bred ones are raised as the efficiency of cloning by SCNT is much lower than natural breeding or In-vitro fertilization (IVF). Thus, we have conducted a genome-wide gene expression and DNA methylation profiling between phenotypically normal cloned pigs and control pigs in two tissues (muscle and liver), using Affymetrix Porcine expression array as well as modified methylation-specific digital karyotyping (MMSDK) and Solexa sequencing technology. Typical tissue-specific differences with respect to both gene expression and DNA methylation were observed in muscle and liver from cloned as well as control pigs. Gene expression profiles were highly similar between cloned pigs and controls, though a small set of genes showed altered expression. Cloned pigs presented a more different pattern of DNA methylation in unique sequences in both tissues. Especially a small set of genomic sites had different DNA methylation status with a trend towards slightly increased methylation levels in cloned pigs. Molecular network analysis of the genes that contained such differential methylation loci revealed a significant network related to tissue development. In conclusion, our study showed that phenotypically normal cloned pigs were highly similar with normal breeding pigs in their gene expression, but moderate alteration in DNA methylation aspects still exists, especially in certain unique genomic regions. PMID:22022462

  16. [Cloning and bioinformatics analysis of SLA-DR genes in Hunan Shaziling pigs].

    PubMed

    Tang, Yi-Ya; Xing, Xiao-Wei; Xue, Li-Qun; Huang, Sheng-Qiang; Wang, Wei

    2007-12-01

    In order to clone class II DRA and DRB genes of swine leukocyte antigen (SLA) in Hunan Shaziling pigs, to analyze their characteristics and polymorphism and to provide immunological basic parameters for xenotransplantation from pigs to humans. SLA-DRA and SLA-DRB genes in two Shaziling pigs with the absence of porcine endogenous retrovirus (PERV) env-c were amplified by RT-PCR, cloned into PUCm-T vectors, sequenced and analyzed through BLAST in NCBI and related software in ExPASY. The obtained SLA-DRA and SLA-DRB genes of Shaziling pigs were 1,177 and 909 nucleotides in length with their accession numbers in Genbank as EF143987 and EF143988. Bioinformatics analyses have shown that they both contain opening reading frame (ORF) and encode 252 and 266 amino acids respectively. Comparing the ORF and protein sequences of the Shaziling SLA-DRA and SLA-DRB genes with their counterpart sequences of human, the homologies of nucleotide sequences were 83% and 83%, and the homologies of amino acid sequences 83 % and 79% respectively. Further comparison with SLA sequences published in GenBank indicated that SLA-DRB gene found in Shaziling pigs has polymorphism while the homology of SLA-DRA gene is up to 100 % .

  17. Production of transgenic cloned pigs expressing the far-red fluorescent protein monomeric Plum.

    PubMed

    Watanabe, Masahito; Kobayashi, Mirina; Nagaya, Masaki; Matsunari, Hitomi; Nakano, Kazuaki; Maehara, Miki; Hayashida, Gota; Takayanagi, Shuko; Sakai, Rieko; Umeyama, Kazuhiro; Watanabe, Nobuyuki; Onodera, Masafumi; Nagashima, Hiroshi

    2015-01-01

    Monomeric Plum (Plum), a far-red fluorescent protein with photostability and photopermeability, is potentially suitable for in vivo imaging and detection of fluorescence in body tissues. The aim of this study was to generate transgenic cloned pigs exhibiting systemic expression of Plum using somatic cell nuclear transfer (SCNT) technology. Nuclear donor cells for SCNT were obtained by introducing a Plum-expression vector driven by a combination of the cytomegalovirus early enhancer and chicken beta-actin promoter into porcine fetal fibroblasts (PFFs). The cleavage and blastocyst formation rates of reconstructed SCNT embryos were 81.0% (34/42) and 78.6% (33/42), respectively. At 36-37 days of gestation, three fetuses systemically expressing Plum were obtained from one recipient to which 103 SCNT embryos were transferred (3/103, 2.9%). For generation of offspring expressing Plum, rejuvenated PFFs were established from one cloned fetus and used as nuclear donor cells. Four cloned offspring and one stillborn cloned offspring were produced from one recipient to which 117 SCNT embryos were transferred (5/117, 4.3%). All offspring exhibited high levels of Plum fluorescence in blood cells, such as lymphocytes, monocytes and granulocytes. In addition, the skin, heart, kidney, pancreas, liver and spleen also exhibited Plum expression. These observations demonstrated that transfer of the Plum gene did not interfere with the development of porcine SCNT embryos and resulted in the successful generation of transgenic cloned pigs that systemically expressed Plum. This is the first report of the generation and characterization of transgenic cloned pigs expressing the far-red fluorescent protein Plum.

  18. Production of transgenic cloned pigs expressing the far-red fluorescent protein monomeric Plum

    PubMed Central

    WATANABE, Masahito; KOBAYASHI, Mirina; NAGAYA, Masaki; MATSUNARI, Hitomi; NAKANO, Kazuaki; MAEHARA, Miki; HAYASHIDA, Gota; TAKAYANAGI, Shuko; SAKAI, Rieko; UMEYAMA, Kazuhiro; WATANABE, Nobuyuki; ONODERA, Masafumi; NAGASHIMA, Hiroshi

    2015-01-01

    Monomeric Plum (Plum), a far-red fluorescent protein with photostability and photopermeability, is potentially suitable for in vivo imaging and detection of fluorescence in body tissues. The aim of this study was to generate transgenic cloned pigs exhibiting systemic expression of Plum using somatic cell nuclear transfer (SCNT) technology. Nuclear donor cells for SCNT were obtained by introducing a Plum-expression vector driven by a combination of the cytomegalovirus early enhancer and chicken beta-actin promoter into porcine fetal fibroblasts (PFFs). The cleavage and blastocyst formation rates of reconstructed SCNT embryos were 81.0% (34/42) and 78.6% (33/42), respectively. At 36–37 days of gestation, three fetuses systemically expressing Plum were obtained from one recipient to which 103 SCNT embryos were transferred (3/103, 2.9%). For generation of offspring expressing Plum, rejuvenated PFFs were established from one cloned fetus and used as nuclear donor cells. Four cloned offspring and one stillborn cloned offspring were produced from one recipient to which 117 SCNT embryos were transferred (5/117, 4.3%). All offspring exhibited high levels of Plum fluorescence in blood cells, such as lymphocytes, monocytes and granulocytes. In addition, the skin, heart, kidney, pancreas, liver and spleen also exhibited Plum expression. These observations demonstrated that transfer of the Plum gene did not interfere with the development of porcine SCNT embryos and resulted in the successful generation of transgenic cloned pigs that systemically expressed Plum. This is the first report of the generation and characterization of transgenic cloned pigs expressing the far-red fluorescent protein Plum. PMID:25739316

  19. Production of Pigs by Hand-Made Cloning Using Mesenchymal Stem Cells and Fibroblasts.

    PubMed

    Yang, Zhenzhen; Vajta, Gábor; Xu, Ying; Luan, Jing; Lin, Mufei; Liu, Cong; Tian, Jianing; Dou, Hongwei; Li, Yong; Liu, Tianbin; Zhang, Yijie; Li, Lin; Yang, Wenxian; Bolund, Lars; Yang, Huanming; Du, Yutao

    2016-08-01

    Mesenchymal stem cells (MSCs) exhibited self-renewal and less differentiation, making the MSCs promising candidates for adult somatic cell nuclear transfer (SCNT). In this article, we tried to produce genome identical pigs through hand-made cloning (HMC), with MSCs and adult skin fibroblasts as donor cells. MSCs were derived from either adipose tissue or peripheral blood (aMSCs and bMSCs, respectively). MSCs usually showed the expression pattern of CD29, CD73, CD90, and CD105 together with lack of expression of the hematopoietic markers CD34and CD45. Flow cytometry results demonstrated high expression of CD29 and CD90 in both MSC lines, while CD73, CD34, and CD45 expression were not detected. In contrary, in reverse transcription-polymerase chain reaction (RT-PCR) analysis, CD73 and CD34 were detected indicating that human antibodies CD73 and CD34 were not suitable to identify porcine cell surface markers and porcine MSC cellular surface markers of CD34 might be different from other species. MSCs also had potential to differentiate successfully into chondrocytes, osteoblasts, and adipocytes. After HMC, embryos reconstructed with aMSCs had higher blastocyst rate on day 5 and 6 than those reconstructed with bMSCs and fibroblasts (29.6% ± 1.3% and 41.1% ± 1.4% for aMSCs vs. 23.9% ± 1.2% and 35.5% ± 1.6% for bMSCs and 22.1% ± 0.9% and 33.3% ± 1.1% for fibroblasts, respectively). Live birth rate per transferred blastocyst achieved with bMSCs (1.59%) was the highest among the three groups. This article was the first report to compare the efficiency among bMSCs, aMSCs, and fibroblasts for boar cloning, which offered a realistic perspective to use the HMC technology for commercial breeding. PMID:27459584

  20. Molecular cloning and functional expression of a sodium bicarbonate cotransporter from guinea-pig parotid glands.

    PubMed

    Koo, Na-Youn; Li, Jingchao; Hwang, Sung Min; Choi, Se-Young; Lee, Sung Joong; Oh, Seog-Bae; Kim, Joong-Soo; Lee, Jong Heun; Park, Kyungpyo

    2006-04-21

    We recently found that the concentration of HCO3- in guinea-pig saliva is very similar to that of human saliva; however, the entity that regulates HCO3- transport has not yet been fully characterized. In order to investigate the mechanism of HCO3- transport, we identified, cloned, and characterized a sodium bicarbonate (Na(+)/HCO3- cotransporter found in guinea-pig parotid glands (gpNBC1). The gpNBC1 gene encodes a 1079-amino acid protein that has 95% and 96% homology with human and mouse parotid NBC1, respectively. Oocytes expressing gpNBC1 were exposed to HCO3- or Na(+)-free solutions, which resulted in a marked change in membrane potentials (V(m)), suggesting that gpNBC1 is electrogenic. Likewise, a gpNBC1-mediated pH recovery was observed in gpNBC1 transfected human hepatoma cells; however, in the presence of 4, 4-diisothiocyanostilbene-2,2-disulfonic acid, a specific NBC1 inhibitor, such changes in V(m) and pH(i) were not observed. Together, the data show that the cloned guinea-pig gene is a functional, as well as sequence homologue of human NBC1. PMID:16513089

  1. Growth, reproductive performance, carcass characteristics and meat quality in F1 and F2 progenies of somatic cell-cloned pigs.

    PubMed

    Adachi, Noritaka; Yamaguchi, Daisuke; Watanabe, Akiyuki; Miura, Narumi; Sunaga, Seiji; Oishi, Hitoshi; Hashimoto, Michiko; Oishi, Takatsugu; Iwamoto, Masaki; Hanada, Hirofumi; Kubo, Masanori; Onishi, Akira

    2014-04-24

    The objective of this study was to examine the health and meat production of cloned sows and their progenies in order to demonstrate the application of somatic cell cloning to the pig industry. This study compared the growth, reproductive performance, carcass characteristics and meat quality of Landrace cloned sows, F1 progenies and F2 progenies. We measured their body weight, growth rate and feed conversion and performed a pathological analysis of their anatomy to detect abnormalities. Three of the five cloned pigs were used for a growth test. Cloned pigs grew normally and had characteristics similar to those of the control purebred Landrace pigs. Two cloned gilts were bred with a Landrace boar and used for a progeny test. F1 progenies had characteristics similar to those of the controls. Two of the F1 progeny gilts were bred with a Duroc or Large White boar and used for the progeny test. F2 progenies grew normally. There were no biological differences in growth, carcass characteristics and amino acid composition among cloned sows, F1 progenies, F2 progenies and conventional pigs. The cloned sows and F1 progenies showed normal reproductive performance. No specific abnormalities were observed by pathological analysis, with the exception of periarteritis in the F1 progenies. All pigs had a normal karyotype. These results demonstrate that cloned female pigs and their progenies have similar growth, reproductive performance and carcass quality characteristics and that somatic cell cloning could be a useful technique for conserving superior pig breeds in conventional meat production.

  2. Generation of heterozygous fibrillin-1 mutant cloned pigs from genome-edited foetal fibroblasts.

    PubMed

    Umeyama, Kazuhiro; Watanabe, Kota; Watanabe, Masahito; Horiuchi, Keisuke; Nakano, Kazuaki; Kitashiro, Masateru; Matsunari, Hitomi; Kimura, Tokuhiro; Arima, Yoshimi; Sampetrean, Oltea; Nagaya, Masaki; Saito, Masahiro; Saya, Hideyuki; Kosaki, Kenjiro; Nagashima, Hiroshi; Matsumoto, Morio

    2016-01-01

    Marfan syndrome (MFS) is an autosomal dominant genetic disease caused by abnormal formation of the extracellular matrix with an incidence of 1 in 3, 000 to 5, 000. Patients with Marfan syndrome experience poor quality of life caused by skeletal disorders such as scoliosis, and they are at high risk of sudden death from cardiovascular impairment. Suitable animal models of MFS are essential for conquering this intractable disease. In particular, studies employing pig models will likely provide valuable information that can be extrapolated to humans because of the physiological and anatomical similarities between the two species. Here we describe the generation of heterozygous fibrillin-1 (FBN1) mutant cloned pigs (+/Glu433AsnfsX98) using genome editing and somatic cell nuclear transfer technologies. The FBN1 mutant pigs exhibited phenotypes resembling those of humans with MFS, such as scoliosis, pectus excavatum, delayed mineralization of the epiphysis and disrupted structure of elastic fibres of the aortic medial tissue. These findings indicate the value of FBN1 mutant pigs as a model for understanding the pathogenesis of MFS and for developing treatments. PMID:27074716

  3. Generation of heterozygous fibrillin-1 mutant cloned pigs from genome-edited foetal fibroblasts

    PubMed Central

    Umeyama, Kazuhiro; Watanabe, Kota; Watanabe, Masahito; Horiuchi, Keisuke; Nakano, Kazuaki; Kitashiro, Masateru; Matsunari, Hitomi; Kimura, Tokuhiro; Arima, Yoshimi; Sampetrean, Oltea; Nagaya, Masaki; Saito, Masahiro; Saya, Hideyuki; Kosaki, Kenjiro; Nagashima, Hiroshi; Matsumoto, Morio

    2016-01-01

    Marfan syndrome (MFS) is an autosomal dominant genetic disease caused by abnormal formation of the extracellular matrix with an incidence of 1 in 3, 000 to 5, 000. Patients with Marfan syndrome experience poor quality of life caused by skeletal disorders such as scoliosis, and they are at high risk of sudden death from cardiovascular impairment. Suitable animal models of MFS are essential for conquering this intractable disease. In particular, studies employing pig models will likely provide valuable information that can be extrapolated to humans because of the physiological and anatomical similarities between the two species. Here we describe the generation of heterozygous fibrillin-1 (FBN1) mutant cloned pigs (+/Glu433AsnfsX98) using genome editing and somatic cell nuclear transfer technologies. The FBN1 mutant pigs exhibited phenotypes resembling those of humans with MFS, such as scoliosis, pectus excavatum, delayed mineralization of the epiphysis and disrupted structure of elastic fibres of the aortic medial tissue. These findings indicate the value of FBN1 mutant pigs as a model for understanding the pathogenesis of MFS and for developing treatments. PMID:27074716

  4. First cloned swamp buffalo produced from adult ear fibroblast cell.

    PubMed

    Tasripoo, K; Suthikrai, W; Sophon, S; Jintana, R; Nualchuen, W; Usawang, S; Bintvihok, A; Techakumphu, M; Srisakwattana, K

    2014-07-01

    The world's first cloned swamp buffalo (Bubalus bubalis) derived from adult ear skin fibroblast has been reported. Donor fibroblast cells were produced from biopsies taken from adult male ear skin and in vitro matured oocytes obtained from a slaughterhouse were used as cytoplasts. A total of 39 blastocysts and 19 morulae fresh embryos were transferred into 12 recipient buffaloes. Progesterone assays indicated establishment of pregnancy in 10 of the 12 buffaloes (83.3%) after 45 days, with six animals still pregnant at 3 months. One recipient maintained pregnancy to term and naturally delivered a 40 kg male calf after 326 days of gestation. DNA analysis showed that the cloned calf was genetically identical to the donor cells. Genotype analyses, using 12 buffalo microsatellite markers, confirmed that the cloned calf was derived from the donor cell lines. In conclusion, the present study reports, for the first time, the establishment of pregnancy and birth of the first cloned Thai swamp buffalo derived from adult ear skin fibroblast cells.

  5. [Cloning and regulation of pig estrogen related receptor β gene (ESRRB) promoter].

    PubMed

    Yang, Yang; Wang, Yaxian; Du, Lixia; Wang, Huayan

    2015-04-01

    The estrogen related receptor family member Esrrb (Estrogen related receptor β) is a gene that expresses in the early stage of embryo and plays an important role in the core pluripotent network. Its function has been analyzed in human and mouse, although no report so far related to pig. Therefore, to explore its mechanism of transcriptional regulation and expression pattern, we cloned a 3.3 kb pig ESRRB promoter by PCR and constructed the green fluorescence protein (GFP) reporter vector pE3.3. We used these vectors to study the ESRRB expression pattern in 293T, Hela and C2C12. Sequence was analyzed for regulatory elements that share homology to known transcription factor binding sites by TFSEARCH and JASPER program. Some pluripotency related genes such as SMAD, STAT3, MYC, KLF4 and ESRRB have been found within the 3.3 kb sequence by co-transfected pig ESRRB promoter and these potential regulators. We found that ESRRB only expressed in 293T and SMAD could activate ESRRB expression obviously. To determine the core promoter region, a series of ESRRB promoter fragments with gradually truncated 5'-end were produced by PCR and inserted into pGL3-Basic vector. After transient transfection into 293T, dual luciferase assay was used to measure these promoter activities. The result suggested that the core promoter of pig ESRRB located within -25 bp to -269 bp region. These results suggest that these transcription factor binding sites and the core promoter region may be essential for transcriptional regulation of pig ESRRB gene. PMID:26380406

  6. [Molecular cloning and characteristics of cDNA encoding pig beta6 subunit for FMDV receptor].

    PubMed

    Gao, Shan-Dian; Du, Jun-Zheng; Chang, Hui-Yun; Cong, Guo-Zheng; Shao, Jun-Jun; Shan, Yi Hua; Zhou, Jian-Hua; Xie, Qing-Ge

    2007-09-01

    In order to study the roles of integrin beta6 in Foot-and-Mouth Disease Virus infection, pig integrin beta6 was firstly molecularly cloned from RNA of the tongue and lung of recovered pig infected experimentally with foot-and-mouth-disease virus (FMDV), and was compared with the beta6 gene of other animals available in GenBank at nucleotide and amino acid leves. GeneBank association number of the beta6 gene is EF432729. Pig integrin beta6 gene (2367bp) encodes a polypeptide of 788 amino acids consisting of 9 potential N-linked glycosylation sites, 3 Glycosaminoglycan attachment sites, a cGMP-dependent protein kinase phosphorylation site, 10 Protein kinase C phosphorylation sites, 2 EGF-like domains and 2 cysteine-rich regions. Pig integrin beta6 subunit has a 26-residue putative signal peptide, a 681-residue ectodomain, a 29-residue transmembrane domain, and a 52-residue cytoplasmic domain. 11 mutant nucleotides were found in beta6 gene coding region and 9 amino acids were changed. The nucleotide sequence similarity of integrin beta6 gene between rheses monkey, mouse, Norway rat, dog, guinea pig, human, bovine, sheep is 79.5%, 84.9%, 85.4%, 85.2%, 88.7%, 90.1%, 91.9% and 91.9%, and the amino acid sequence similarity is 93.5%, 88.2%, 88.5%, 88.3%, 91.0%, 92.8%, 93.3% and 93.4% respectively. This study will lay a foundation for understanding the interactions of FMDV with receptors. PMID:18064756

  7. 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

  8. Production of Transgenic-Cloned Pigs Expressing Large Quantities of Recombinant Human Lysozyme in Milk

    PubMed Central

    Shang, Shengzhe; Wu, Fangfang; Wen, Xiao; Li, Zhiyuan; Li, Yan; Hu, Xiaoxiang; Zhao, Yaofeng; Li, Qiuyan; Li, Ning

    2015-01-01

    Human lysozyme is a natural non-specific immune factor in human milk that plays an important role in the defense of breastfed infants against pathogen infection. Although lysozyme is abundant in human milk, there is only trace quantities in pig milk. Here, we successfully generated transgenic cloned pigs with the expression vector pBAC-hLF-hLZ-Neo and their first generation hybrids (F1). The highest concentration of recombinant human lysozyme (rhLZ) with in vitro bioactivity was 2759.6 ± 265.0 mg/L in the milk of F0 sows. Compared with wild-type milk, rhLZ milk inhibited growth of Escherichia coli K88 during the exponential growth phase. Moreover, rhLZ in milk from transgenic sows was directly absorbed by the intestine of piglets with no observable anaphylactic reaction. Our strategy may provide a powerful tool for large-scale production of this important human protein in pigs to improve resistance to pathogen infection. PMID:25955256

  9. Production of Cloned Miniature Pigs Expressing High Levels of Human Apolipoprotein(a) in Plasma.

    PubMed

    Ozawa, Masayuki; Himaki, Takehiro; Ookutsu, Shoji; Mizobe, Yamato; Ogawa, Junki; Miyoshi, Kazuchika; Yabuki, Akira; Fan, Jianglin; Yoshida, Mitsutoshi

    2015-01-01

    High lipoprotein(a) [Lp(a)] levels are a major risk factor for the development of atherosclerosis. However, because apolipoprotein(a) [apo(a)], the unique component of Lp(a), is found only in primates and humans, the study of human Lp(a) has been hampered due to the lack of appropriate animal models. Using somatic cell nuclear transfer (SCNT) techniques, we produced transgenic miniature pigs expressing human apo(a) in the plasma. First, we placed the hemagglutinin (HA)-tagged cDNA of human apo(a) under the control of the β-actin promoter and cytomegalovirus enhancer, and then introduced this construct into kidney epithelial cells. Immunostaining of cells with anti-HA antibody allowed identification of cells stably expressing apo(a); one of the positive clones was used to provide donor cells for SCNT, yielding blastocysts that expressed apo(a). Immunohistochemical analysis of tissue sections and RT-PCR analysis of total RNA from organs of cloned piglet revealed that apo(a) is expressed in various tissues/organs including heart, liver, kidney, and intestine. More importantly, a transgenic line exhibited a high level (>400 mg/dL) of Lp(a) in plasma, and the transgenic apo(a) gene was transmitted to the offspring. Thus, we generated a human apo(a)-transgenic miniature pig that can be used as a model system to study advanced atherosclerosis related to human disease. The anatomical and physiological similarities between the swine and human cardiovascular systems will make this pig model a valuable source of information on the role of apo(a) in the formation of atherosclerosis, as well as the mechanisms underlying vascular health and disease. PMID:26147378

  10. Production of Cloned Miniature Pigs Expressing High Levels of Human Apolipoprotein(a) in Plasma

    PubMed Central

    Ozawa, Masayuki; Himaki, Takehiro; Ookutsu, Shoji; Mizobe, Yamato; Ogawa, Junki; Miyoshi, Kazuchika; Yabuki, Akira; Fan, Jianglin; Yoshida, Mitsutoshi

    2015-01-01

    High lipoprotein(a) [Lp(a)] levels are a major risk factor for the development of atherosclerosis. However, because apolipoprotein(a) [apo(a)], the unique component of Lp(a), is found only in primates and humans, the study of human Lp(a) has been hampered due to the lack of appropriate animal models. Using somatic cell nuclear transfer (SCNT) techniques, we produced transgenic miniature pigs expressing human apo(a) in the plasma. First, we placed the hemagglutinin (HA)-tagged cDNA of human apo(a) under the control of the β-actin promoter and cytomegalovirus enhancer, and then introduced this construct into kidney epithelial cells. Immunostaining of cells with anti-HA antibody allowed identification of cells stably expressing apo(a); one of the positive clones was used to provide donor cells for SCNT, yielding blastocysts that expressed apo(a). Immunohistochemical analysis of tissue sections and RT-PCR analysis of total RNA from organs of cloned piglet revealed that apo(a) is expressed in various tissues/organs including heart, liver, kidney, and intestine. More importantly, a transgenic line exhibited a high level (>400 mg/dL) of Lp(a) in plasma, and the transgenic apo(a) gene was transmitted to the offspring. Thus, we generated a human apo(a)–transgenic miniature pig that can be used as a model system to study advanced atherosclerosis related to human disease. The anatomical and physiological similarities between the swine and human cardiovascular systems will make this pig model a valuable source of information on the role of apo(a) in the formation of atherosclerosis, as well as the mechanisms underlying vascular health and disease. PMID:26147378

  11. Cloning and expression of pig kidney dopa decarboxylase: comparison of the naturally occurring and recombinant enzymes.

    PubMed Central

    Moore, P S; Dominici, P; Borri Voltattorni, C

    1996-01-01

    L-Aromatic amino acid decarboxylase (dopa decarboxylase; DDC) is a pyridoxal 5'-phosphate (PLP)-dependent homodimeric enzyme that catalyses the decarboxylation of L-dopa and other L-aromatic amino acids. To advance structure-function studies with the enzyme, a cDNA that codes for the protein from pig kidney has been cloned by joining a partial cDNA obtained by library screening with a synthetic portion constructed by the annealing and extension of long oligonucleotides. The hybrid cDNA was then expressed in Escherichia coli to produce recombinant protein. During characterization of the recombinant enzyme it was unexpectedly observed that it possesses certain differences from the enzyme purified from pig kidney. Whereas the later protein binds 1 molecule of PLP per dimer, the recombinant enzyme was found to bind two molecules of coenzyme per dimer. Moreover, the Vmax was twice that of the protein purified from tissue. On addition of substrate, the absorbance changes accompanying transaldimination were likewise 2-fold greater in the recombinant enzyme. Examination of the respective apoenzymes by absorbance, CD and fluorescence spectroscopy revealed distinct differences. The recombinant apoprotein has no significant absorbance at 335 nm, unlike the pig kidney apoenzyme; in the latter case this residual absorbance is associated with a positive dichroic signal. When excited at 335 nm the pig kidney apoenzyme has a pronounced emission maximum at 385 nm, in contrast with its recombinant counterpart, which shows a weak broad emission at about 400 nm. However, the holoenzyme-apoenzyme transition did not markedly alter the respective fluorescence properties of either recombinant or pig kidney DDC when excited at 335 nm. Taken together, these findings indicate that recombinant pig kidney DDC has two active-site PLP molecules and therefore displays structural characteristics typical of PLP-dependent homodimeric enzymes. The natural enzyme contains one active-site PLP molecule

  12. 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

  13. 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.

  14. The transgenic cloned pig population with integrated and controllable GH expression that has higher feed efficiency and meat production.

    PubMed

    Ju, Huiming; Zhang, Jiaqing; Bai, Lijing; Mu, Yulian; Du, Yutao; Yang, Wenxian; Li, Yong; Sheng, Anzhi; Li, Kui

    2015-01-01

    Sustained expression of the GH gene has been shown to have detrimental effects on the health of animals. In the current study, transgenic founder pigs, with controllable pig growth hormone (pGH) expression, were cloned via the handmade cloning method (HMC), and pGH expression levels were examined at the cellular and organismal levels. The serum pGH levels in 3 founder male pigs were found to be significantly higher after induction with intramuscular injection of doxycycline (DOX) compared to baseline. A daily dose of DOX was administered via feed to these animals for a period of 65 to 155 days. The growth rate, feed efficiency and pGH serum concentration increased in the DOX-induced transgenic group compared with the other groups. 8 numbers of animals were euthanized and the dressing percentage, loin muscle and lean meat percentage were significantly higher in the DOX-induced F1 transgenic group compared with the other groups. In this study a large population of transgenic pigs, with integrated controllable expression of a transgene, was obtained. The transgenic pigs were healthy and normal in terms of reproductive capability. At the same time, feed efficiency was improved, production processes were accelerated and meat yield was increased.

  15. SLC26 anion exchangers of guinea pig pancreatic duct: molecular cloning and functional characterization

    PubMed Central

    Stewart, Andrew K.; Shmukler, Boris E.; Vandorpe, David H.; Reimold, Fabian; Heneghan, John F.; Nakakuki, M.; Akhavein, Arash; Ko, Shigeru; Ishiguro, Hiroshi

    2011-01-01

    The secretin-stimulated human pancreatic duct secretes HCO3−-rich fluid essential for normal digestion. Optimal stimulation of pancreatic HCO3− secretion likely requires coupled activities of the cystic fibrosis transmembrane regulator (CFTR) anion channel and apical SLC26 Cl−/HCO3− exchangers. However, whereas stimulated human and guinea pig pancreatic ducts secrete ∼140 mM HCO3− or more, mouse and rat ducts secrete ∼40–70 mM HCO3−. Moreover, the axial distribution and physiological roles of SLC26 anion exchangers in pancreatic duct secretory processes remain controversial and may vary among mammalian species. Thus the property of high HCO3− secretion shared by human and guinea pig pancreatic ducts prompted us to clone from guinea pig pancreatic duct cDNAs encoding Slc26a3, Slc26a6, and Slc26a11 polypeptides. We then functionally characterized these anion transporters in Xenopus oocytes and human embryonic kidney (HEK) 293 cells. In Xenopus oocytes, gpSlc26a3 mediated only Cl−/Cl− exchange and electroneutral Cl−/HCO3− exchange. gpSlc26a6 in Xenopus oocytes mediated Cl−/Cl− exchange and bidirectional exchange of Cl− for oxalate and sulfate, but Cl−/HCO3− exchange was detected only in HEK 293 cells. gpSlc26a11 in Xenopus oocytes exhibited pH-dependent Cl−, oxalate, and sulfate transport but no detectable Cl−/HCO3− exchange. The three gpSlc26 anion transporters exhibited distinct pharmacological profiles of 36Cl− influx, including partial sensitivity to CFTR inhibitors Inh-172 and GlyH101, but only Slc26a11 was inhibited by PPQ-102. This first molecular and functional assessment of recombinant SLC26 anion transporters from guinea pig pancreatic duct enhances our understanding of pancreatic HCO3− secretion in species that share a high HCO3− secretory output. PMID:21593449

  16. 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.

  17. Cloning, sequencing, and polymorphism analysis of novel classical MHC class I alleles in northern pig-tailed macaques (Macaca leonina).

    PubMed

    Lian, Xiao-Dong; Zhang, Xi-He; Dai, Zheng-Xi; Zheng, Yong-Tang

    2016-04-01

    The northern pig-tailed macaque (Macaca leonina) has been confirmed to be an independent species from the pig-tailed macaque group of Old World monkey. We have previously reported that the northern pig-tailed macaques were also susceptible to HIV-1. Here, to make this animal a potential HIV/AIDS model and to discover the mechanism of virus control, we attempted to assess the role of major histocompatibility complex (MHC) class I-restricted immune responses to HIV-1 infection, which was associated with viral replication and disease progression. As an initial step, we first cloned and characterized the classical MHC class I gene of northern pig-tailed macaques. In this study, we identified 39 MHC class I alleles including 17 MHC-A and 22 MHC-B alleles. Out of these identified alleles, 30 were novel and 9 were identical to alleles previously reported from other macaque species. The MHC-A and MHC-B loci were both duplicates as rhesus macaques and southern pig-tailed macaques. In addition, we also detected the patterns of positive selection in northern pig-tailed macaques and revealed the existence of balance selection with 20 positive selection sites in the peptide binding region. The analysis of B and F peptide binding pockets in northern and southern pig-tailed macaques and rhesus macaques suggested that they were likely to share a few common peptides to present. Thus, this study provides important MHC immunogenetics information and adds values to northern pig-tailed macaques as a promising HIV/AIDS model.

  18. Cloning, sequencing, and polymorphism analysis of novel classical MHC class I alleles in northern pig-tailed macaques (Macaca leonina).

    PubMed

    Lian, Xiao-Dong; Zhang, Xi-He; Dai, Zheng-Xi; Zheng, Yong-Tang

    2016-04-01

    The northern pig-tailed macaque (Macaca leonina) has been confirmed to be an independent species from the pig-tailed macaque group of Old World monkey. We have previously reported that the northern pig-tailed macaques were also susceptible to HIV-1. Here, to make this animal a potential HIV/AIDS model and to discover the mechanism of virus control, we attempted to assess the role of major histocompatibility complex (MHC) class I-restricted immune responses to HIV-1 infection, which was associated with viral replication and disease progression. As an initial step, we first cloned and characterized the classical MHC class I gene of northern pig-tailed macaques. In this study, we identified 39 MHC class I alleles including 17 MHC-A and 22 MHC-B alleles. Out of these identified alleles, 30 were novel and 9 were identical to alleles previously reported from other macaque species. The MHC-A and MHC-B loci were both duplicates as rhesus macaques and southern pig-tailed macaques. In addition, we also detected the patterns of positive selection in northern pig-tailed macaques and revealed the existence of balance selection with 20 positive selection sites in the peptide binding region. The analysis of B and F peptide binding pockets in northern and southern pig-tailed macaques and rhesus macaques suggested that they were likely to share a few common peptides to present. Thus, this study provides important MHC immunogenetics information and adds values to northern pig-tailed macaques as a promising HIV/AIDS model. PMID:26782049

  19. T-cell-activating monoclonal antibodies, reacting with both leukocytes and erythrocytes, recognize the guinea pig Thy-1 differentiation antigen: characterization and cloning of guinea pig CD90.

    PubMed

    Schäfer, H; Bartels, T; Hahn, G; Otto, A; Burger, R

    1999-11-01

    A glycophosphatidylinositol (GPI)-linked differentiation antigen expressed on guinea pig T and B lymphocytes was identified by several monoclonal antibodies; it has been shown previously that this membrane protein induced strong polyclonal T cell proliferation upon antibody binding and costimulation by PMA. Purification by immunoadsorption and microsequencing revealed that this T-cell-activating protein is the homologue of Thy-1 or CD90. In contrast to the Thy-1 antigen of most other species, guinea pig Thy-1 has a much higher molecular weight, which is due to a more extensive N-linked glycosylation, bringing the molecular weight of the total antigen up to 36 kDa. Molecular cloning of guinea pig Thy-1 indicated that the deduced molecular weight of the protein backbone is 12,777 after removal of an N-terminal 19-amino-acid leader peptide and cleavage of the 31 amino acids for GPI anchoring the C-terminal end. Sequence comparison showed that guinea pig Thy-1 has an 82% homology to human and a 72% homology to mouse Thy-1 on the amino acid level. Immunohistological staining of cryostat sections revealed intensive staining with the monoclonal antibody H154 on fibroblasts, fibrocytes, Kupffer cells, alveolar macrophages, and mesangial cells. As observed in the human, mouse, and rat, Thy-1 is abundant in the guinea pig brain. Unlike Thy-1 expression in other species, guinea pig Thy-1 is strongly expressed on most resting, nonactivated B cells and, to a lesser extent, on erythrocytes. While treatment of erythrocytes and lymphocytes with GPI-specific phospholipase C largely decreased reactivity with mAb H154, T cells retained the proliferative response to antibody and phorbol esters.

  20. Analysis of nuclear reprogramming in cloned miniature pig embryos by expression of Oct-4 and Oct-4 related genes

    SciTech Connect

    Lee, Eugine; Lee, So Hyun; Kim, Sue

    2006-10-06

    Xenotransplantation is a rapidly expanding field of research and cloned miniature pigs have been considered as a model animal for it. However, the efficiency of somatic cell nuclear transfer (SCNT) is extremely low, with most clones resulting in early lethality and several kinds of aberrant development. A possible explanation for the developmental failure of SCNT embryos is insufficient reprogramming of the somatic cell nucleus by the oocyte. In order to test this, we analyzed the reprogramming capacity of differentiated fibroblast cell nuclei and embryonic germ cell nuclei with Oct-4 and Oct-4 related genes (Ndp5211, Dppa2, Dppa3, and Dppa5), which are important for embryonic development, Hand1 and GATA-4, which are important for placental development, as molecular markers using RT-PCR. The Oct-4 expression level was significantly lower (P < 0.05) in cloned hatched blastocysts derived from fibroblasts and many of fibroblast-derived clones failed to reactivate at least one of the tested genes, while most of the germ cell clones and control embryos correctly expressed these genes. In conclusion, our results suggest that the reprogramming of fibroblast-derived cloned embryos is highly aberrant and this improper reprogramming could be one reason of the early lethality and post-implantation anomalies of somatic cell-derived clones.

  1. Molecular cloning of the flavin-containing monooxygenase (form II) cDNA from adult human liver.

    PubMed Central

    Lomri, N; Gu, Q; Cashman, J R

    1992-01-01

    Complementary DNA (cDNA) clones encoding the adult human liver flavin-containing monooxygenase (FMO; dimethylaniline N-oxidase, EC 1.14.13.8) were isolated from lambda gt10 and lambda gt11 libraries. The cDNA libraries were screened with three synthetic 36-mer oligonucleotide probes derived from the nucleic acid sequence of the pig liver FMO cDNA. The deduced amino acid sequence for the adult human liver FMO was quite distinct from the pig liver FMO, and adult human liver FMO was designated form II (HLFMO II). The full-length cDNA sequence of HLFMO II [2119 base pairs (bp)] had an open reading frame of 1599 nucleotides, which encoded a 533-amino acid protein of Mr 59,179, a 5'-noncoding region of 136 nucleotides and a 3'-noncoding region of 369 nucleotides excluding the poly(A) tail. The deduced amino acid sequence of HLFMO II had 80% similarity with the rabbit liver FMO II but only a 52%, 55%, and 53% amino acid similarity with the rabbit liver (form I), the pig liver (form I), and fetal human liver (form I) FMOs, respectively. RNA analysis of adult human liver RNA showed that there was one HLFMO II mRNA species. Analysis of genomic DNA indicated that HLFMO II was the product of a single gene. These results indicated that the deduced amino acid sequence for HLFMO II contained highly conserved residues and suggested that FMO enzymes were closely related and, undoubtedly, derived from the same ancestral gene. Images PMID:1542660

  2. 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.

  3. Developmental kinetics of pig embryos by parthenogenetic activation or by handmade cloning.

    PubMed

    Li, J; Li, R; Liu, Y; Villemoes, K; Purup, S; Callesen, H

    2013-10-01

    The developmental kinetics of pig embryos produced by parthenogenetic activation without (PAZF) or with (PAZI) zona pellucida or by handmade cloning (HMC) was compared by time-lapse videography. After cumulus cell removal, the matured oocytes were either left zona intact (PAZI) or were made zona free by pronase digestion (PAZF) before they were activated (PA). Other matured oocytes were used for HMC based on foetal fibroblast cells. On Day 0 (day of PA or reconstruction), the embryos were cultured for 7 days in vitro in our time-lapse system. Pictures were taken every 30 min, and afterwards, each cell cycle was identified for each embryo to be analysed. Results showed that the PA embryos (both PAZF and PAZI) had shorter first cell cycle compared with HMC (17.4. 17.8 vs 23.6 h), but had a longer time length from four cell to morula stages (57.9, 53.8 vs 44.9 h). However, at the second cell cycle, PAZF embryos needed shorter time, while PAZI embryos had similar time length as HMC embryos, and both were longer than PAZF (23.4, 24.8 vs 14.6 h). Both PAZF and PAZI embryos used similar time to reach the blastocyst stage, and this was later than HMC embryos. In addition, when all of these embryos were grouped into viable (developed to blastocysts) and non-viable (not developed to blastocysts), the only difference in the time length was observed on the first cell cycle (18.6 vs 24.5 h), but not on the later cell cycles. In conclusion, our results not only give detailed information regarding the time schedule of in vitro-handled pig embryos, but also indicate that the first cell cycle could be used as a selecting marker for embryo viability. However, to evaluate the effect of the produced techniques, the whole time schedule of the pre-implantation developmental kinetics should be observed.

  4. 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.

  5. Abcb1 in Pigs: Molecular cloning, tissues distribution, functional analysis, and its effect on pharmacokinetics of enrofloxacin.

    PubMed

    Guo, Tingting; Huang, Jinhu; Zhang, Hongyu; Dong, Lingling; Guo, Dawei; Guo, Li; He, Fang; Bhutto, Zohaib Ahmed; Wang, Liping

    2016-01-01

    P-glycoprotein (P-gp) is one of the best-known ATP-dependent efflux transporters, contributing to differences in pharmacokinetics and drug-drug interactions. Until now, studies on pig P-gp have been scarce. In our studies, the full-length porcine P-gp cDNA was cloned and expressed in a Madin-Darby Canine Kidney (MDCK) cell line. P-gp expression was then determined in tissues and its role in the pharmacokinetics of oral enrofloxacin in pigs was studied. The coding region of pig Abcb1 gene was 3,861 bp, encoding 1,286 amino acid residues (Mw = 141,966). Phylogenetic analysis indicated a close evolutionary relationship between porcine P-gp and those of cow and sheep. Pig P-gp was successfully stably overexpressed in MDCK cells and had efflux activity for rhodamine 123, a substrate of P-gp. Tissue distribution analysis indicated that P-gp was highly expressed in brain capillaries, small intestine, and liver. In MDCK-pAbcb1 cells, enrofloxacin was transported by P-gp with net efflux ratio of 2.48 and the efflux function was blocked by P-gp inhibitor verapamil. High expression of P-gp in the small intestine could modify the pharmacokinetics of orally administrated enrofloxacin by increasing the Cmax, AUC and Ka, which was demonstrated using verapamil, an inhibitor of P-gp. PMID:27572343

  6. Abcb1 in Pigs: Molecular cloning, tissues distribution, functional analysis, and its effect on pharmacokinetics of enrofloxacin

    PubMed Central

    Guo, Tingting; Huang, Jinhu; Zhang, Hongyu; Dong, Lingling; Guo, Dawei; Guo, Li; He, Fang; Bhutto, Zohaib Ahmed; Wang, Liping

    2016-01-01

    P-glycoprotein (P-gp) is one of the best-known ATP-dependent efflux transporters, contributing to differences in pharmacokinetics and drug-drug interactions. Until now, studies on pig P-gp have been scarce. In our studies, the full-length porcine P-gp cDNA was cloned and expressed in a Madin-Darby Canine Kidney (MDCK) cell line. P-gp expression was then determined in tissues and its role in the pharmacokinetics of oral enrofloxacin in pigs was studied. The coding region of pig Abcb1 gene was 3,861 bp, encoding 1,286 amino acid residues (Mw = 141,966). Phylogenetic analysis indicated a close evolutionary relationship between porcine P-gp and those of cow and sheep. Pig P-gp was successfully stably overexpressed in MDCK cells and had efflux activity for rhodamine 123, a substrate of P-gp. Tissue distribution analysis indicated that P-gp was highly expressed in brain capillaries, small intestine, and liver. In MDCK-pAbcb1 cells, enrofloxacin was transported by P-gp with net efflux ratio of 2.48 and the efflux function was blocked by P-gp inhibitor verapamil. High expression of P-gp in the small intestine could modify the pharmacokinetics of orally administrated enrofloxacin by increasing the Cmax, AUC and Ka, which was demonstrated using verapamil, an inhibitor of P-gp. PMID:27572343

  7. Cloning adult animals - what is the genetic age of the clones?

    PubMed

    Yang, X; Tian, X C

    2000-01-01

    The rapid progress in cloning research along with its many ramifications will soon have a significant beneficial impact on basic research, agriculture, and biomedicine. However, for the nuclear transfer technology to reach its fullest potential, it is important to understand whether the cloning procedure can reverse cellular aging and generate clones with normal genetic and physiological age, similar to those produced from natural reproduction. Telomere shortening is believed to correlate with cellular aging both in vitro and in vivo. Telomere lengths in cells of cloned individuals thus may reflect their genetic age. However, controversies have developed over whether the eroded telomere in somatic cells used for nuclear transfer can be restored during the cloning process.

  8. Increasing histone acetylation of cloned embryos, but not donor cells, by sodium butyrate improves their in vitro development in pigs.

    PubMed

    Das, Ziban Chandra; Gupta, Mukesh Kumar; Uhm, Sang Jun; Lee, Hoon Taek

    2010-02-01

    Previous studies have demonstrated that increased histone acetylation in donor cells or cloned embryos, by applying a histone deacetylase inhibitor (HDACi) such as trichostatin A (TSA), significantly enhances their developmental competence. However, its effect may vary with the type of HDACi and the target species, with some research showing nonsignificant or detrimental effects of TSA on in vitro and in vivo development of embryos. In this study, we show that sodium salt of butyric acid, a short-chain fatty acid produced naturally in the body by bacterial degradation of dietary fibers in the colon and rectum, increases histone acetylation in pig fibroblast and embryos at a concentration of 1.0 and 5.0 mM, respectively. However, treatment of donor cells with NaBu did not affect the rate of blastocyst formation or embryo quality in terms of histone acetylation and total nuclei per blastocyst (p > 0.05). On the contrary, treatment of cloned pig embryos with NaBu for 4 h significantly enhanced (p < 0.01) the rate of blastocyst formation (18.3 +/- 2.1 vs. 11.2 +/- 3.0%), although the total nuclei number per blastocyst did not differ. More importantly, blastocysts generated from NaBu-treated cloned embryos had increased levels of histone acetylation that was comparable to those of in vitro fertilized (IVF) embryos (36.7 +/- 3.6 vs. 45.9 +/- 2.5). In conclusion, our data suggest that histone hyperacetylation by NaBu treatment of cloned embryos, but not donor cell, enhances their in vitro development up to blastocyst stage.

  9. 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.

  10. Cloning

    MedlinePlus

    ... copies of whole animals Therapeutic cloning, which creates embryonic stem cells. Researchers hope to use these cells to grow healthy tissue to replace injured or diseased tissues in the human body. NIH: National Human Genome Research Institute

  11. cAMP sensitivity conferred to the epithelial Na+ channel by alpha-subunit cloned from guinea-pig colon.

    PubMed

    Schnizler, M; Mastroberardino, L; Reifarth, F; Weber, W M; Verrey, F; Clauss, W

    2000-03-01

    The rate of Na+ (re)absorption across tight epithelia such as in distal kidney nephron and colon is to a large extent controlled at the level of the epithelial Na+ channel (ENaC). In kidney, antidiuretic hormone (ADH, vasopressin) stimulates the expression/activity of this channel by a cAMP/protein-kinase-A- (PKA-) mediated pathway. However, a clear upregulation of ENaC function by cAMP could not be reproduced with cloned channel subunits in the Xenopus oocyte expression system, suggesting the hypothesis that an additional factor is missing. In contrast, we show here that membrane-permeant cAMP can activate ENaC expressed in Xenopus oocytes (3.8-fold) upon replacement of the rat alpha-subunit by a new alpha-subunit cloned from guinea-pig colon (gpalpha). This alpha-subunit is 76% identical with its rat orthologue originating from ADH-insensitive rat colon. The biophysical fingerprints of the hybrid ENaC formed by this guinea-pig alpha-subunit together with rat beta- and gamma-subunits are indistinguishable from those of rat ENaC (rENaC). Injection of the PKA inhibitor PKI-(6-22)-amide into the oocyte had no effect on the basal activity of rat ENaC but inhibited the activity of gpalpha-containing hybrid ENaC and greatly decreased its stimulation by cAMP. This suggests that, unlike for rat ENaC, tonic PKA activity is required for basal function of gpalpha-containing ENaC and that PKA mediates its cAMP-induced activation. This regulatory behaviour is not common to all ENaCs containing an alpha-subunit cloned from an ADH-responsive tissue since xENaC, which was cloned from the ADH-sensitive Xenopus laevis A6 epithelia, is, when expressed in oocytes, resistant to cAMP, similar to rat ENaC. This study demonstrates that the PKA sensitivity of ENaC can depend on the nature of the ENaC alpha-subunit and raises the possibility that cAMP can stimulate ENaCs by different mechanisms. PMID:10764218

  12. Expression of Huntington's disease protein results in apoptotic neurons in the brains of cloned transgenic pigs

    PubMed Central

    Yang, Dongshan; Wang, Chuan-En; Zhao, Bentian; Li, Wei; Ouyang, Zhen; Liu, Zhaoming; Yang, Huaqiang; Fan, Pei; O'Neill, Ashley; Gu, Weiwang; Yi, Hong; Li, Shihua; Lai, Liangxue; Li, Xiao-Jiang

    2010-01-01

    Neurodegeneration is a hallmark of many neurological diseases, including Alzheimer's, Parkinson's and the polyglutamine diseases, which are all caused by misfolded proteins that accumulate in neuronal cells of the brain. Although apoptosis is believed to contribute to neurodegeneration in these cases, genetic mouse models of these diseases often fail to replicate apoptosis and overt neurodegeneration in the brain. Using nuclear transfer, we generated transgenic Huntington's disease (HD) pigs that express N-terminal (208 amino acids) mutant huntingtin with an expanded polyglutamine tract (105Q). Postnatal death, dyskinesia and chorea-like movement were observed in some transgenic pigs that express mutant huntingtin. Importantly, the transgenic HD pigs, unlike mice expressing the same transgene, displayed typical apoptotic neurons with DNA fragmentation in their brains. Also, expression of mutant huntingtin resulted in more neurons with activated caspase-3 in transgenic pig brains than that in transgenic mouse brains. Our findings suggest that species differences determine neuropathology and underscore the importance of large mammalian animals for modeling neurological disorders. PMID:20660116

  13. Analysis of Gene Expression and Ultrastructure of Stifle Menisci from Juvenile and Adult Pigs

    PubMed Central

    Kreinest, Michael; Reisig, Gregor; Ströbel, Philipp; Fickert, Stefan; Brade, Joachim; Wennemuth, Gunther; Lipp, Peter; Schwarz, Markus L

    2016-01-01

    The origin of the age-associated degenerative processes in meniscal tissue is poorly understood and may be related to an imbalance of anabolic and catabolic metabolism. The aim of the current study was to compare medial menisci isolated from juvenile pigs and degenerated medial menisci from adult pigs in terms of gene expression profile and ultrastructure. Medial menisci were isolated from the knee joints of juvenile and adult pigs (n = 8 for each group). Degeneration was determined histologically according to a scoring system. In addition, the gene expression profiles of 14 genes encoding extracellular matrix proteins, catabolic matrix metalloproteinases and mediators of inflammation were analyzed. Changes in the ultrastructure of the collagen network of the meniscal tissue were analyzed by using transmission electron microscopy. The histologic analysis of menisci showed significantly higher grade of degeneration in tissue isolated from adult porcine knee joints compared with menisci isolated from juvenile knee joints. In particular, destruction of the collagen network was greater in adult menisci than in juvenile menisci. Degenerated menisci showed significantly decreased gene expression of COL1A1 and increased expression of MMP2, MMP13, and IL8. The menisci from adult porcine knee joints can serve as a model for meniscal degeneration. Degenerative changes were manifested as differences in histopathology, gene expression and ultrastructure of collagen network. PMID:26884408

  14. Molecular cloning and anti-HIV-1 activities of APOBEC3s from northern pig-tailed macaques (Macaca leonina).

    PubMed

    Zhang, Xiao-Liang; Song, Jia-Hao; Pang, Wei; Zheng, Yong-Tang

    2016-07-18

    Northern pig-tailed macaques (NPMs, Macaca leonina) are susceptible to HIV-1 infection largely due to the loss of HIV-1-restricting factor TRIM5α. However, great impediments still exist in the persistent replication of HIV-1 in vivo, suggesting some viral restriction factors are reserved in this host. The APOBEC3 proteins have demonstrated a capacity to restrict HIV-1 replication, but their inhibitory effects in NPMs remain elusive. In this study, we cloned the NPM A3A-A3H genes, and determined by BLAST searching that their coding sequences (CDSs) showed 99% identity to the corresponding counterparts from rhesus and southern pig-tailed macaques. We further analyzed the anti-HIV-1 activities of the A3A-A3H genes, and found that A3G and A3F had the greatest anti-HIV-1 activity compared with that of other members. The results of this study indicate that A3G and A3F might play critical roles in limiting HIV-1 replication in NPMs in vivo. Furthermore, this research provides valuable information for the optimization of monkey models of HIV-1 infection. PMID:27469256

  15. Molecular cloning and anti-HIV-1 activities of APOBEC3s from northern pig-tailed macaques (Macaca leonina)

    PubMed Central

    ZHANG, Xiao-Liang; SONG, Jia-Hao; PANG, Wei; ZHENG, Yong-Tang

    2016-01-01

    Northern pig-tailed macaques (NPMs, Macaca leonina) are susceptible to HIV-1 infection largely due to the loss of HIV-1-restricting factor TRIM5α. However, great impediments still exist in the persistent replication of HIV-1 in vivo, suggesting some viral restriction factors are reserved in this host. The APOBEC3 proteins have demonstrated a capacity to restrict HIV-1 replication, but their inhibitory effects in NPMs remain elusive. In this study, we cloned the NPM A3A-A3H genes, and determined by BLAST searching that their coding sequences (CDSs) showed 99% identity to the corresponding counterparts from rhesus and southern pig-tailed macaques. We further analyzed the anti-HIV-1 activities of the A3A-A3H genes, and found that A3G and A3F had the greatest anti-HIV-1 activity compared with that of other members. The results of this study indicate that A3G and A3F might play critical roles in limiting HIV-1 replication in NPMs in vivo. Furthermore, this research provides valuable information for the optimization of monkey models of HIV-1 infection. PMID:27469256

  16. Chronic prenatal ethanol exposure increases disinhibition and perseverative responding in the adult guinea pig.

    PubMed

    Olmstead, Mary C; Martin, Amanda; Brien, James F; Reynolds, James N

    2009-09-01

    Cognitive and behavioural deficits, including increased impulsivity and perseveration, are associated with chronic prenatal ethanol exposure (CPEE) in humans. We tested whether these same deficits occur in the guinea pig after CPEE treatment. Pregnant guinea pigs received oral administration of ethanol (4 g/kg maternal body weight/day), or isocaloric-sucrose/pair-feeding throughout gestation. Young adult offspring were trained in lever-pressing paradigms to work for a sucrose-pellet food reward. CPEE increased No-Go, but not Go, responses in the Go/No-Go paradigm, indicative of a disinhibition deficit in these animals. Perseverative responses in the Cued Alternation task were also increased in CPEE offspring. These data show that CPEE induces behavioural deficits in the guinea pig that are remarkably similar to the executive function deficits that follow prenatal ethanol exposure in humans.

  17. Cloning of the full length pig PIT1 (POU1F1) CDNA and a novel alternative PIT1 transcript, and functional studies of their encoded proteins.

    PubMed

    Yu, T P; Sun, H S; Wahls, S; Sanchez-Serrano, I; Rothschild, M F; Tuggle, C K

    2001-05-01

    PIT1 is an essential regulatory gene of growth hormone (GH), prolactin (PRL) and thyrotropin beta subunit (TSHbeta). Previously, a partial pig PIT1 cDNA and a genomic clone of the entire 3' end of the PIT1 gene was isolated, and polymorphisms at PIT1 were associated with several performance traits in the pig. In order to understand the biological function of the pig PIT1 gene and its possible application in swine genetics, reverse transcriptase-polymerase chain reaction (RT-PCR) was used to complete the cloning of the full length cDNA for pig PIT1. The pig PIT1 cDNA and its deduced protein sequence have approximately 90% and 95% identity, respectively, with the PIT1 cDNA and protein of other mammals (human, bovine, sheep and rodents). Surprisingly, sequence comparison to other pig PIT1 sequences indicated only approximately 93% identity. Additional sequencing confirmed our sequence, and identified a new polymorphism in exon 4. Phylogenetic analysis of several mammalian PIT1 sequences indicates sequencing errors may account for the discrepancies observed in the other pig sequences reported. Several PIT1 alternative spliced forms were also identified by RT-PCR. They were the delta3PIT1 (missing entire exon 3), delta4PIT1 (missing entire exon 4) and PIT1beta (additional 26 amino acids inserted in front of exon 2) transcripts. The delta4PIT1 and PIT1beta transcripts have been found to encode functionally different proteins in rodents. The delta3PIT1 transcript is a novel isoform of PIT1. Potentially different functions between pig delta3PIT1 and PIT1 were analyzed by expressing these proteins in bacteria. The E. coli-expressed PIT1 and delta3PIT1 proteins were used with rat growth hormone (rGH) and rat prolactin (rPRL) promoter DNA in DNA mobility shift assays. The results showed that pig PIT1 can specifically bind rGH and rPRL promoter regions, but that the pig delta3PIT1 cannot, even at very high protein concentrations. Possible protein-protein interactions between

  18. Isozygous and selectable marker-free MSTN knockout cloned pigs generated by the combined use of CRISPR/Cas9 and Cre/LoxP.

    PubMed

    Bi, Yanzhen; Hua, Zaidong; Liu, Ximei; Hua, Wenjun; Ren, Hongyan; Xiao, Hongwei; Zhang, Liping; Li, Li; Wang, Zhirui; Laible, Götz; Wang, Yan; Dong, Faming; Zheng, Xinmin

    2016-01-01

    Predictable, clean genetic modification (GM) in livestock is important for reliable phenotyping and biosafety. Here we reported the generation of isozygous, functional myostatin (MSTN) knockout cloned pigs free of selectable marker gene (SMG) by CRISPR/Cas9 and Cre/LoxP. CRISPR/Cas9-mediated homologous recombination (HR) was exploited to knock out (KO) one allele of MSTN in pig primary cells. Cre recombinase was then used to excise the SMG with an efficiency of 82.7%. The SMG-free non-EGFP cells were isolated by flow cytometery and immediately used as donor nuclei for nuclear transfer. A total of 685 reconstructed embryos were transferred into three surrogates with one delivering two male live piglets. Molecular testing verified the mono-allelic MSTN KO and SMG deletion in these cloned pigs. Western blots showed approximately 50% decrease in MSTN and concurrent increased expression of myogenic genes in muscle. Histological examination revealed the enhanced myofiber quantity but myofiber size remained unaltered. Ultrasonic detection showed the increased longissimus muscle size and decreased backfat thickness. Precision editing of pig MSTN gene has generated isozygous, SMG-free MSTN KO cloned founders, which guaranteed a reliable route for elite livestock production and a strategy to minimize potential biological risks. PMID:27530319

  19. Isozygous and selectable marker-free MSTN knockout cloned pigs generated by the combined use of CRISPR/Cas9 and Cre/LoxP

    PubMed Central

    Bi, Yanzhen; Hua, Zaidong; Liu, Ximei; Hua, Wenjun; Ren, Hongyan; Xiao, Hongwei; Zhang, Liping; Li, Li; Wang, Zhirui; Laible, Götz; Wang, Yan; Dong, Faming; Zheng, Xinmin

    2016-01-01

    Predictable, clean genetic modification (GM) in livestock is important for reliable phenotyping and biosafety. Here we reported the generation of isozygous, functional myostatin (MSTN) knockout cloned pigs free of selectable marker gene (SMG) by CRISPR/Cas9 and Cre/LoxP. CRISPR/Cas9-mediated homologous recombination (HR) was exploited to knock out (KO) one allele of MSTN in pig primary cells. Cre recombinase was then used to excise the SMG with an efficiency of 82.7%. The SMG-free non-EGFP cells were isolated by flow cytometery and immediately used as donor nuclei for nuclear transfer. A total of 685 reconstructed embryos were transferred into three surrogates with one delivering two male live piglets. Molecular testing verified the mono-allelic MSTN KO and SMG deletion in these cloned pigs. Western blots showed approximately 50% decrease in MSTN and concurrent increased expression of myogenic genes in muscle. Histological examination revealed the enhanced myofiber quantity but myofiber size remained unaltered. Ultrasonic detection showed the increased longissimus muscle size and decreased backfat thickness. Precision editing of pig MSTN gene has generated isozygous, SMG-free MSTN KO cloned founders, which guaranteed a reliable route for elite livestock production and a strategy to minimize potential biological risks. PMID:27530319

  20. Molecular cloning of the. cap alpha. subunit of human and guinea pig leukocyte adhesion glycoprotein Mo1: Chromosomal localization and homology to the. cap alpha. subunits of integrins

    SciTech Connect

    Arnaout, M.A.; Remold-O'Donnell, E.; Pierce, M.W.; Harris, P.; Tenen, D.G.

    1988-04-01

    The cell surface-glycoprotein Mo1 is a member of the family of leukocyte cell adhesion molecules (Leu-CAMs) that includes lymphocyte function-associated antigen 1 (LFA-1) and p150,95. Each Leu-CAM is a heterodimer with a distinct ..cap alpha.. subunit noncovalently associated with a common ..beta.. subunit. The authors describe the isolation and analysis of two partial cDNA clones encoding the ..cap alpha.. subunit of the Leu-CAM Mo1 in humans and guinea pigs. A monoclonal antibody directed against an epitope in the carboxyl-terminal portion of the guinea pig ..cap alpha.. chain was used for immunoscreening a lambdagt11 expression library. The sequence of a 378-base-pair insert from one immunoreactive clone revealed a single continuous open reading frame encoding 126 amino acids including a 26-amino acid tryptic peptide isolated from the purified guinea pig ..cap alpha.. subunit. A cDNA clone of identical size was isolated from a human monocyte/lymphocyte cDNA library by using the guinea pig clone as a probe. The human clone also encoded a 126-amino acid peptide including the sequence of an additional tryptic peptide present in purified human Mo1..cap alpha.. chain. Southern analysis of DNA from hamster-human hybrids localized the human Mo1..cap alpha.. chain to chromosome 16, which has been shown to contain the gene for the ..cap alpha.. chain of lymphocyte function-associated antigen 1. These data suggest that the ..cap alpha.. subunits of Leu-CAMs evolved by gene duplication from a common ancestral gene and strengthen the hypothesis that the ..cap alpha.. subunits of these heterodimeric cell adhesion molecules on myeloid and lymphoid cells, platelets, and fibroblasts are evolutionary related.

  1. BIX-01294 increases pig cloning efficiency by improving epigenetic reprogramming of somatic cell nuclei.

    PubMed

    Huang, Jiaojiao; Zhang, Hongyong; Yao, Jing; Qin, Guosong; Wang, Feng; Wang, Xianlong; Luo, Ailing; Zheng, Qiantao; Cao, Chunwei; Zhao, Jianguo

    2016-01-01

    Accumulating evidence suggests that faulty epigenetic reprogramming leads to the abnormal development of cloned embryos and results in the low success rates observed in all mammals produced through somatic cell nuclear transfer (SCNT). The aberrant methylation status of H3K9me and H3K9me2 has been reported in cloned mouse embryos. To explore the role of H3K9me2 and H3K9me in the porcine somatic cell nuclear reprogramming, BIX-01294, known as a specific inhibitor of G9A (histone-lysine methyltransferase of H3K9), was used to treat the nuclear-transferred (NT) oocytes for 14-16 h after activation. The results showed that the developmental competence of porcine SCNT embryos was significantly enhanced both in vitro (blastocyst rate 16.4% vs 23.2%, P<0.05) and in vivo (cloning rate 1.59% vs 2.96%) after 50 nm BIX-01294 treatment. BIX-01294 treatment significantly decreased the levels of H3K9me2 and H3K9me at the 2- and 4-cell stages, which are associated with embryo genetic activation, and increased the transcriptional expression of the pluripotency genes SOX2, NANOG and OCT4 in cloned blastocysts. Furthermore, the histone acetylation levels of H3K9, H4K8 and H4K12 in cloned embryos were decreased after BIX-01294 treatment. However, co-treatment of activated NT oocytes with BIX-01294 and Scriptaid rescued donor nuclear chromatin from decreased histone acetylation of H4K8 that resulted from exposure to BIX-01294 only and consequently improved the preimplantation development of SCNT embryos (blastocyst formation rates of 23.7% vs 21.5%). These results indicated that treatment with BIX-01294 enhanced the developmental competence of porcine SCNT embryos through improvements in epigenetic reprogramming and gene expression.

  2. Telomere-to-centromere ratio of bovine clones, embryos, gametes, fetal cells, and adult cells.

    PubMed

    Meerdo, Lora N; Reed, William A; White, Kenneth L

    2005-01-01

    In 1997, Dolly, the first animal cloned from an adult cell, was born. It was announced in 1999 that Dolly might be aging faster than normal because her telomeres were shorter than age-matched control sheep. Telomeres, a repeated DNA sequence located at the ends of linear chromosomes, allow for base pair loss during DNA replication. Telomere shortening acts as a "mitotic clock," leading to replicative senescence. By using whole cell lysate and slot-blot analysis, we determined the telomere-to-centromere ratio (T/C) for bovine gametes, embryos, fetal tissues (brain, heart, lung, kidney, uterus, ovary, and skin), adult donor cells, and cloned embryos. Our data indicates a consistency in T/C among the various fetal tissues. The T/C of sperm is significantly lower than in oocytes. The T/C decreases from the oocyte to the 2-8-cell stage embryo, increases dramatically at the morula stage, and decreases at the blastocyst stage. Our data shows no significant difference in T/C between cloned embryos and in vitro fertilized (IVF) embryos, but there is a significant difference between cloned embryos and adult donor cells. In conclusion, the enucleated bovine oocyte has the ability to reestablish the telomere length of adult somatic cell donor nuclei. PMID:15996118

  3. Layer I as a putative neurogenic niche in young adult guinea pig cerebrum.

    PubMed

    Xiong, Kun; Cai, Yan; Zhang, Xue-Mei; Huang, Ju-Fang; Liu, Zhong-Yu; Fu, Guang-Ming; Feng, Jia-Chun; Clough, Richard W; Patrylo, Peter R; Luo, Xue-Gang; Hu, Chun-Hong; Yan, Xiao-Xin

    2010-10-01

    A considerable number of cells expressing typical immature neuronal markers including doublecortin (DCX+) are present around layer II in the cerebral cortex of young and adult guinea pigs and other larger mammals, and their origin and biological implication await further characterization. We show here in young adult guinea pigs that these DCX+ cells are accompanied by in situ cell division around the superficial cortical layers mostly in layer I, but they co-express proliferating cell nuclear antigen (PCNA) and an early neuronal fate determining factor, PAX6. A small number of these DCX+ cells also colocalize with BrdU following administration of this mitotic indicator. Cranial X-ray irradiation causes a decline of DCX+ cells around layer II, and novel environmental exploration induces c-Fos expression among these cells in several neocortical areas. Together, these data are compatible with a notion that DCX+ cortical neurons around layer II might derive from proliferable neuronal precursors around layer I in young adult guinea pig cerebrum, and that these cells might be modulated by experience under physiological conditions.

  4. Pigs fed cholesterol neonatally have increased cerebrum cholesterol as young adults.

    PubMed

    Boleman, S L; Graf, T L; Mersmann, H J; Su, D R; Krook, L P; Savell, J W; Park, Y W; Pond, W G

    1998-12-01

    Sixty-eight female neonatal pigs selected for seven (Experiment 1) or eight (Experiment 2) generations for high (HG) or low (LG) plasma cholesterol were used to test the hypothesis that neonatal dietary cholesterol fed during the first 4 or 8 wk of postnatal life increases the cholesterol content of the cerebrum in young adulthood following free access to a high-fat (15%), high-cholesterol (0.5%) diet from 8 to 20 or 24 wk of age. Pigs were removed from their dams at 1 d of age and given free access to a sow-milk replacer diet containing 9.5% coconut fat and 0 or 0.5 % cholesterol. All pigs (except four HG and four LG pigs in Experiment 2, which were deprived of cholesterol throughout the study) were fed the high-fat, high-cholesterol diet from 8 wk to termination at 20 or 24 wk of age. Cerebrum weight and cholesterol concentration were higher in pigs fed cholesterol neonatally than in those deprived of cholesterol neonatally in both experiments, but weight and cholesterol concentration were unaffected by genetic line. Cholesterol concentrations in longissimus and semitendinosus muscles and in subcutaneous fat were unaffected by diet or genetic line. We conclude that dietary cholesterol deprivation during the first 4 to 8 wk of life in piglets is associated with lower cholesterol concentration and total content in the young adult cerebrum than in pigs supplemented with cholesterol in early life. These data support previous observations and suggest the possibility of a metabolic need for neonatal dietary cholesterol in normal brain development. PMID:9868199

  5. Production of cloned NIBS (Nippon Institute for Biological Science) and α-1, 3-galactosyltransferase knockout MGH miniature pigs by somatic cell nuclear transfer using the NIBS breed as surrogates

    PubMed Central

    Shimatsu, Yoshiki; Yamada, Kazuhiko; Horii, Wataru; Hirakata, Atsushi; Sakamoto, Yuji; Waki, Shiori; Sano, Junichi; Saitoh, Toshiki; Sahara, Hisashi; Shimizu, Akira; Yazawa, Hajime; Sachs, David H.; Nunoya, Tetsuo

    2013-01-01

    Background Nuclear transfer (NT) technologies offer a means for producing the genetically modified pigs necessary to develop swine models for mechanistic studies of disease processes as well as to serve as organ donors for xenotransplantation. Most previous studies have used commercial pigs as surrogates. Method and Results In this study, we established a cloning technique for miniature pigs by somatic cell nuclear transfer (SCNT) using Nippon Institute for Biological Science (NIBS) miniature pigs as surrogates. Moreover, utilizing this technique, we have successfully produced an α-1, 3-galactosyltransferase knockout (GalT-KO) miniature swine. Fibroblasts procured from a NIBS miniature pig fetus were injected into 1312 enucleated oocytes. The cloned embryos were transferred to 11 surrogates of which five successfully delivered 13 cloned offspring; the production efficiency was 1.0% (13/1312). In a second experiment, lung fibroblasts obtained from neonatal GalT-KO MGH miniature swine were used as donor cells and 1953 cloned embryos were transferred to 12 surrogates. Six cloned offspring were born from five surrogates, a production efficiency of 0.3% (6/1953). Conclusions These results demonstrate successful establishment of a miniature pig cloning technique by SCNT using NIBS miniature pigs as surrogates. To our knowledge, this is the first demonstration of successful production of GalT-KO miniature swine using miniature swine surrogates. This technique could help to ensure a stable supply of the cloned pigs through the use of miniature pig surrogates and could expand production in countries with limited space or in facilities with special regulations such as specific pathogen-free or good laboratory practice. PMID:23581451

  6. Neuromedin B and Its Receptor: Gene Cloning, Tissue Distribution and Expression Levels of the Reproductive Axis in Pigs.

    PubMed

    Ma, Zhiyu; Su, Juan; Guo, Tingting; Jin, Mengmeng; Li, Xiang; Lei, Zhihai; Hou, Yuanlong; Li, Xiaoliang; Jia, Cuicui; Zhang, Zheng; Ahmed, Ejlal

    2016-01-01

    Neuromedin B is one member of a family of bombesin-like peptides, which performs a variety of physiological functions via their receptor (NMBR) in most mammals. However, the genes encoding NMB and NMBR and their functions especially reproduction of the pigs are currently not fully understood. To research the physiological functions of NMB, we cloned and analyzed the NMB and NMBR genes, and systematically investigated the expression levels of NMB and NMBR mRNA using relative real-time PCR and the distribution of NMBR by immunohistochemistry (IHC). Experimental results show that the sequences of the amino acid and gene of NMB and NMBR were highly conservative and homology in many species, Significantly, the relative RT-PCR results revealed that NMB was mainly expressed in the central nervous system (CNS), whereas NMBR is highly expressed in peripheral tissues and organs, such as endocrine tissues, glands and reproductive organs. The IHC results show that NMBR positive cells were widely distributed in the body, such as respiratory and circulatory system, digestive system, urogenital system, in lymphatic organs and in the endocrine system. We also systematically investigated expression levels of NMB and NMBR in the reproductive axis using relative real-time PCR. In sow estrous cycle, the hypothalamic levels of both NMB and NMBR mRAN were similar, but the expression levels of the pituitary were negatively correlated. Expression levels in the ovarian system are lowest in metestrus phases and highest in proestrus and estrus phases. In boar post-natal development stages, the hypothalamic, pituitary and testicular levels of NMB and NMBR mRNAs showed developmental changes on postnatal day 30, 60, 90 and 120. Taken together, this study provided molecular and morphological data necessary for further research of physiological function of NMB/NMBR system in the pigs.

  7. Neuromedin B and Its Receptor: Gene Cloning, Tissue Distribution and Expression Levels of the Reproductive Axis in Pigs.

    PubMed

    Ma, Zhiyu; Su, Juan; Guo, Tingting; Jin, Mengmeng; Li, Xiang; Lei, Zhihai; Hou, Yuanlong; Li, Xiaoliang; Jia, Cuicui; Zhang, Zheng; Ahmed, Ejlal

    2016-01-01

    Neuromedin B is one member of a family of bombesin-like peptides, which performs a variety of physiological functions via their receptor (NMBR) in most mammals. However, the genes encoding NMB and NMBR and their functions especially reproduction of the pigs are currently not fully understood. To research the physiological functions of NMB, we cloned and analyzed the NMB and NMBR genes, and systematically investigated the expression levels of NMB and NMBR mRNA using relative real-time PCR and the distribution of NMBR by immunohistochemistry (IHC). Experimental results show that the sequences of the amino acid and gene of NMB and NMBR were highly conservative and homology in many species, Significantly, the relative RT-PCR results revealed that NMB was mainly expressed in the central nervous system (CNS), whereas NMBR is highly expressed in peripheral tissues and organs, such as endocrine tissues, glands and reproductive organs. The IHC results show that NMBR positive cells were widely distributed in the body, such as respiratory and circulatory system, digestive system, urogenital system, in lymphatic organs and in the endocrine system. We also systematically investigated expression levels of NMB and NMBR in the reproductive axis using relative real-time PCR. In sow estrous cycle, the hypothalamic levels of both NMB and NMBR mRAN were similar, but the expression levels of the pituitary were negatively correlated. Expression levels in the ovarian system are lowest in metestrus phases and highest in proestrus and estrus phases. In boar post-natal development stages, the hypothalamic, pituitary and testicular levels of NMB and NMBR mRNAs showed developmental changes on postnatal day 30, 60, 90 and 120. Taken together, this study provided molecular and morphological data necessary for further research of physiological function of NMB/NMBR system in the pigs. PMID:27010315

  8. Neuromedin B and Its Receptor: Gene Cloning, Tissue Distribution and Expression Levels of the Reproductive Axis in Pigs

    PubMed Central

    Ma, Zhiyu; Su, Juan; Guo, Tingting; Jin, Mengmeng; Li, Xiang; Lei, Zhihai; Hou, Yuanlong; Li, Xiaoliang; Jia, Cuicui; Zhang, Zheng; Ahmed, Ejlal

    2016-01-01

    Neuromedin B is one member of a family of bombesin-like peptides, which performs a variety of physiological functions via their receptor (NMBR) in most mammals. However, the genes encoding NMB and NMBR and their functions especially reproduction of the pigs are currently not fully understood. To research the physiological functions of NMB, we cloned and analyzed the NMB and NMBR genes, and systematically investigated the expression levels of NMB and NMBR mRNA using relative real-time PCR and the distribution of NMBR by immunohistochemistry (IHC). Experimental results show that the sequences of the amino acid and gene of NMB and NMBR were highly conservative and homology in many species, Significantly, the relative RT-PCR results revealed that NMB was mainly expressed in the central nervous system (CNS), whereas NMBR is highly expressed in peripheral tissues and organs, such as endocrine tissues, glands and reproductive organs. The IHC results show that NMBR positive cells were widely distributed in the body, such as respiratory and circulatory system, digestive system, urogenital system, in lymphatic organs and in the endocrine system. We also systematically investigated expression levels of NMB and NMBR in the reproductive axis using relative real-time PCR. In sow estrous cycle, the hypothalamic levels of both NMB and NMBR mRAN were similar, but the expression levels of the pituitary were negatively correlated. Expression levels in the ovarian system are lowest in metestrus phases and highest in proestrus and estrus phases. In boar post-natal development stages, the hypothalamic, pituitary and testicular levels of NMB and NMBR mRNAs showed developmental changes on postnatal day 30, 60, 90 and 120. Taken together, this study provided molecular and morphological data necessary for further research of physiological function of NMB/NMBR system in the pigs. PMID:27010315

  9. 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.

  10. Effects of dietary fibers with different fermentation characteristics on feeding motivation in adult female pigs.

    PubMed

    Souza da Silva, Carol; Bolhuis, J Elizabeth; Gerrits, Walter J J; Kemp, Bas; van den Borne, Joost J G C

    2013-02-17

    Dietary fibers can be fermented in the colon, resulting in production of short-chain fatty acids (SCFA) and secretion of satiety-related peptides. Fermentation characteristics (fermentation kinetics and SCFA-profile) differ between fibers and could impact their satiating potential. We investigated the effects of fibers with varying fermentation characteristics on feeding motivation in adult female pigs. Sixteen pair-housed pigs received four diets in four periods in a Latin square design. Starch from a control (C) diet was exchanged, based on gross energy, for inulin (INU), guar gum (GG), or retrograded tapioca starch (RS), each at a low (L) and a high (H) inclusion level. This resulted in a decreased metabolizable energy intake when feeding fiber diets as compared with the C diet. According to in vitro fermentation measurements, INU is rapidly fermentable and yields relatively high amounts of propionate, GG is moderately rapidly fermentable and yields relatively high amounts of acetate, and RS is slowly fermentable and yields relatively high amounts of butyrate. Feeding motivation was assessed using behavioral tests at 1h, 3h and 7h after the morning meal, and home pen behavioral observations throughout the day. The number of wheel turns paid for a food reward in an operant test was unaffected by diet. Pigs on H-diets ran 25% slower for a food reward in a runway test than pigs on L-diets, and showed less spontaneous physical activity and less stereotypic behavior in the hours before the afternoon meal, reflecting increased interprandial satiety. Reduced feeding motivation with increasing inclusion level was most pronounced for RS, as pigs decreased speed in the runway test and tended to have a lower voluntary food intake in an ad libitum food intake test when fed RS-H. In conclusion, increasing levels of fermentable fibers in the diet seemed to enhance satiety in adult pigs, despite a reduction in metabolizable energy supply. RS was the most satiating fiber

  11. The osmotic stability of lysosomes from adult and foetal guinea-pig liver tissue.

    PubMed

    Turnbull, J M; Neil, M W

    1969-02-01

    1. Lysosome-rich fractions were obtained from foetal liver tissues as early as 35 days uterine age. Foetal lysosomes showed the same ;structure-linked latency' and acid hydrolytic potentiality characteristic of their adult counterparts. 2. The osmotic stability of lysosome-rich fraction from foetal guinea-pig liver tissue was greater than that of the corresponding adult lysosome fractions, p-nitrophenyl-phosphatase being used as marker enzyme. 3. The observation was confirmed by using beta-glycerophosphatase and phenolphthalein beta-glucuronidase as alternative marker enzymes. p-Nitrophenyl phosphate and beta-glycerophosphate appear to act as substrates for the same enzyme. 4. By using p-nitrophenylphosphatase activity measurements it was shown that the osmotic stability of foetal lysosomal fractions decreased with increasing foetal age, but at no time achieved the degree of osmotic instability characteristic of adult lysosomal fractions. 5. The correlation of these findings with the intracellular environment of lysosomes is discussed.

  12. Effects of Eszopiclone and Zolpidem on Sleep and Waking States in the Adult Guinea Pig

    PubMed Central

    Xi, Mingchu; Chase, Michael H.

    2008-01-01

    Study Objective: The present study was designed to compare and contrast the effects of eszopiclone and zolpidem on the states of sleep and wakefulness in chronically instrumented, unanesthetized adult guinea pigs. Design: Adult guinea pigs were implanted with electrodes to record sleep and waking states and to perform a frequency analysis of the EEG. Eszopiclone (1 and 3 mg/kg) and zolpidem (1 and 3 mg/kg) were administered intraperitoneally. Measurements and Results: The administration of eszopiclone (1 and 3 mg/kg) resulted in a significant dose-dependent increase in NREM sleep. Zolpidem produced a significant increase in NREM sleep, but only at a dose of 3 mg/kg. The following changes in NREM and REM sleep, as well as in the power spectra, were all significant when the effects of 1 and 3 mg/kg of eszopiclone were compared with responses induced with 1 and 3 mg/kg of zolpidem, respectively: The increase in NREM sleep produced by eszopiclone was greater than that following the administration of zolpidem. The mean latency to NREM sleep following the administration of eszopiclone was significantly shorter than zolpidem. Eszopiclone significantly increased the latency to REM sleep. The mean duration of episodes of NREM sleep was increased by eszopiclone, but not by zolpidem. The EEG power increased in the delta band and decreased in the theta band during NREM sleep following the administration of eszopiclone. No significant changes occurred in any of the frequency bands analyzed following zolpidem administration. Conclusions: The differences in the effects of eszopiclone and zolpidem on sleep and waking states and the power spectra of the EEG likely reflect the fact that eszopiclone and zolpidem bind to different subunits of the GABAA receptor complex. Citation: Xi M; Chase MH. Effects of eszopiclone and zolpidem on sleep and waking states in the adult guinea pig. SLEEP 2008;31(7):1043-1051. PMID:18652100

  13. Supplemental Analysis for N-linked Sugars in Adult Pig Islets.

    PubMed

    Eguchi, H; Kawamura, T; Kashiyama, N; Matsuura, R; Sakai, R; Nakahata, K; Lo, P-C; Asada, M; Maeda, A; Goto, M; Toyoda, M; Okuyama, H; Miyagawa, S

    2016-05-01

    The pig pancreas is considered to be one of the most suitable sources of islets for clinical xenotransplantation. However, after producing α1-3galactosyltransferase knockout pigs, most of the organs of these pigs showed less antigenicity to the human body. Wild-type adult pig islets (APIs) that originally produced negligible levels of α-Gal, different from neonatal porcine islet-like cell clusters, showed a clear antigenicity to human serum. Concerning the so-called non-Gal epitopes, many studies related to glycoproteins and glycolipids are ongoing in efforts to identify them. However, our knowledge of non-Gal glycoantigens remains incomplete. In our previous study, N-glycans were isolated from APIs, and the structures of 28 of the N-glycans were detected. In this study, to identify additional structures, further analyses were performed by liquid chromatography-mass spectrometry (LC-MS). N-glycans were isolated from APIs by the method described by O'Neil et al with minor modifications and LC-MS-based structural analyses were then performed. The detected N-glycan peaks in the LC-MS spectra were selected using the FLexAnalysis software program and the structures of the glycans were predicted using the GlyocoMod Tool. The API preparation contained 11 peaks and 16 structures were then nominated as containing N-linked sugars. Among them, 5 sulfated glycans were estimated, confirming the existence of sulfate structures in N-glycans in API. In addition, these data may supplement several N-glycan structures that contain two deoxyhexose units, such as fucose, to our previous report. The data herein will be helpful for future studies of antigenicity associated with API. PMID:27320609

  14. 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.

  15. Construction of a normalized directionally cloned cDNA library from adult heart and analysis of 3040 clones by partial sequencing.

    PubMed

    Tanaka, T; Ogiwara, A; Uchiyama, I; Takagi, T; Yazaki, Y; Nakamura, Y

    1996-07-01

    Large-scale sequencing of clones from cDNA libraries derived from specific tissues is a rapid and efficient way of discovering novel genes expressed in those tissues. However, because the heart is continually contracting and relaxing, it strongly expresses muscle-contractile genes and/or mitochondrial genes, a bias that reduces the efficiency of this method. To improve the efficiency of identifying novel genes expressed in the heart, we constructed a normalized directionally cloned cDNA library from adult heart and partially sequenced 3040 clones. Comparisons of these sequence data with known DNA sequences in the database revealed that 57.1% of the clones matched human genes already known, 23.4% were identical or almost identical to human expressed sequence tags (ESTs), 14.2% bore no significant homology to any sequences in the database, and 1.2% represented repetitive sequences. The remaining 4.1% showed some homology with known genes, and Northern blot analysis of several clones in this category revealed that most of them were expressed mainly in the heart and skeletal muscle. After redundancy was excluded, the 3040 clones accounted for 1395 distinctive ESTs, 446 of which exhibited no match to any known sequence. Our results suggest that our normalized library is less redundant than standard libraries and is a useful resource for cataloging genes expressed in the heart. PMID:8661126

  16. Expression of recombinant human α-lactalbumin in milk of transgenic cloned pigs is sufficient to enhance intestinal growth and weight gain of suckling piglets.

    PubMed

    Ma, Jin; Li, Qiuyan; Li, Yan; Wen, Xiao; Li, Zhiyuan; Zhang, Zaihu; Zhang, Jiuming; Yu, Zhengquan; Li, Ning

    2016-06-10

    Human α-lactalbumin (HLA) has very high nutritional value and important physiological functions during the neonatal period. The peptides derived from HLA provide diverse health benefits including antimicrobial, antiviral, immune-modulating, and antihypertensive effects. Thus, it is worth investigating the effects on offspring development of increasing HLA in milk. In this study, we found that recombinant human α-lactalbumin (rHLA) exhibits efficient inhibition of dipeptidyl peptidase-IV (DPP-IV) activity in an in vitro simulated gastrointestinal digestion system. Using a BAC clone containing the complete HLA gene as a candidate vector, we generated two lines of transgenic cloned sows via somatic cell nuclear transfer that over-expressed rHLA. The average concentrations of rHLA in milk from the two lines of transgenic cloned sows were 2.24 ± 0.71 mg/ml and 2.67 ± 1.29 mg/ml. The feeding experiments revealed that rHLA represses dipeptidyl peptidase-IV (DPP-IV) activity in vivo. Furthermore, the piglets reared by rHLA transgenic cloned sows exhibit better performance in gain of body weight and intestine growth than the control piglets reared by non-transgenic sows. Therefore, these findings indicate that rHLA could serve as a natural precursor for a DPP-IV inhibitor, and the transgenic technology that produced the over-expression of rHLA could be a useful method for pig breeders to improve lactation performance.

  17. Expression of recombinant human α-lactalbumin in milk of transgenic cloned pigs is sufficient to enhance intestinal growth and weight gain of suckling piglets.

    PubMed

    Ma, Jin; Li, Qiuyan; Li, Yan; Wen, Xiao; Li, Zhiyuan; Zhang, Zaihu; Zhang, Jiuming; Yu, Zhengquan; Li, Ning

    2016-06-10

    Human α-lactalbumin (HLA) has very high nutritional value and important physiological functions during the neonatal period. The peptides derived from HLA provide diverse health benefits including antimicrobial, antiviral, immune-modulating, and antihypertensive effects. Thus, it is worth investigating the effects on offspring development of increasing HLA in milk. In this study, we found that recombinant human α-lactalbumin (rHLA) exhibits efficient inhibition of dipeptidyl peptidase-IV (DPP-IV) activity in an in vitro simulated gastrointestinal digestion system. Using a BAC clone containing the complete HLA gene as a candidate vector, we generated two lines of transgenic cloned sows via somatic cell nuclear transfer that over-expressed rHLA. The average concentrations of rHLA in milk from the two lines of transgenic cloned sows were 2.24 ± 0.71 mg/ml and 2.67 ± 1.29 mg/ml. The feeding experiments revealed that rHLA represses dipeptidyl peptidase-IV (DPP-IV) activity in vivo. Furthermore, the piglets reared by rHLA transgenic cloned sows exhibit better performance in gain of body weight and intestine growth than the control piglets reared by non-transgenic sows. Therefore, these findings indicate that rHLA could serve as a natural precursor for a DPP-IV inhibitor, and the transgenic technology that produced the over-expression of rHLA could be a useful method for pig breeders to improve lactation performance. PMID:26899869

  18. The Complete Genome Sequences, Unique Mutational Spectra, and Developmental Potency of Adult Neurons Revealed by Cloning.

    PubMed

    Hazen, Jennifer L; Faust, Gregory G; Rodriguez, Alberto R; Ferguson, William C; Shumilina, Svetlana; Clark, Royden A; Boland, Michael J; Martin, Greg; Chubukov, Pavel; Tsunemoto, Rachel K; Torkamani, Ali; Kupriyanov, Sergey; Hall, Ira M; Baldwin, Kristin K

    2016-03-16

    Somatic mutation in neurons is linked to neurologic disease and implicated in cell-type diversification. However, the origin, extent, and patterns of genomic mutation in neurons remain unknown. We established a nuclear transfer method to clonally amplify the genomes of neurons from adult mice for whole-genome sequencing. Comprehensive mutation detection and independent validation revealed that individual neurons harbor ∼100 unique mutations from all classes but lack recurrent rearrangements. Most neurons contain at least one gene-disrupting mutation and rare (0-2) mobile element insertions. The frequency and gene bias of neuronal mutations differ from other lineages, potentially due to novel mechanisms governing postmitotic mutation. Fertile mice were cloned from several neurons, establishing the compatibility of mutated adult neuronal genomes with reprogramming to pluripotency and development. PMID:26948891

  19. The Complete Genome Sequences, Unique Mutational Spectra, and Developmental Potency of Adult Neurons Revealed by Cloning.

    PubMed

    Hazen, Jennifer L; Faust, Gregory G; Rodriguez, Alberto R; Ferguson, William C; Shumilina, Svetlana; Clark, Royden A; Boland, Michael J; Martin, Greg; Chubukov, Pavel; Tsunemoto, Rachel K; Torkamani, Ali; Kupriyanov, Sergey; Hall, Ira M; Baldwin, Kristin K

    2016-03-16

    Somatic mutation in neurons is linked to neurologic disease and implicated in cell-type diversification. However, the origin, extent, and patterns of genomic mutation in neurons remain unknown. We established a nuclear transfer method to clonally amplify the genomes of neurons from adult mice for whole-genome sequencing. Comprehensive mutation detection and independent validation revealed that individual neurons harbor ∼100 unique mutations from all classes but lack recurrent rearrangements. Most neurons contain at least one gene-disrupting mutation and rare (0-2) mobile element insertions. The frequency and gene bias of neuronal mutations differ from other lineages, potentially due to novel mechanisms governing postmitotic mutation. Fertile mice were cloned from several neurons, establishing the compatibility of mutated adult neuronal genomes with reprogramming to pluripotency and development.

  20. Pancreatic Islets: Methods for Isolation and Purification of Juvenile and Adult Pig Islets.

    PubMed

    Brandhorst, Heide; Johnson, Paul R V; Brandhorst, Daniel

    2016-01-01

    The current situation of organ transplantation is mainly determined by the disbalance between the number of available organs and the number of patients on the waiting list. This obvious dilemma might be solved by the transplantation of porcine organs into human patients. The metabolic similarities which exist between both species made pancreatic islets of Langerhans to that donor tissue which will be most likely transplanted in human recipients. Nevertheless, the successful isolation of significant yields of viable porcine islets is extremely difficult and requires extensive experiences in the field. This review is focussing on the technical challenges, pitfalls and particularities that are associated with the isolation of islets from juvenile and adult pigs considering donor variables that can affect porcine islet isolation outcome.

  1. Pancreatic Islets: Methods for Isolation and Purification of Juvenile and Adult Pig Islets.

    PubMed

    Brandhorst, Heide; Johnson, Paul R V; Brandhorst, Daniel

    2016-01-01

    The current situation of organ transplantation is mainly determined by the disbalance between the number of available organs and the number of patients on the waiting list. This obvious dilemma might be solved by the transplantation of porcine organs into human patients. The metabolic similarities which exist between both species made pancreatic islets of Langerhans to that donor tissue which will be most likely transplanted in human recipients. Nevertheless, the successful isolation of significant yields of viable porcine islets is extremely difficult and requires extensive experiences in the field. This review is focussing on the technical challenges, pitfalls and particularities that are associated with the isolation of islets from juvenile and adult pigs considering donor variables that can affect porcine islet isolation outcome. PMID:27586421

  2. Potent spinal parenchymal AAV9-mediated gene delivery by subpial injection in adult rats and pigs

    PubMed Central

    Miyanohara, Atsushi; Kamizato, Kota; Juhas, Stefan; Juhasova, Jana; Navarro, Michael; Marsala, Silvia; Lukacova, Nada; Hruska-Plochan, Marian; Curtis, Erik; Gabel, Brandon; Ciacci, Joseph; Ahrens, Eric T; Kaspar, Brian K; Cleveland, Don; Marsala, Martin

    2016-01-01

    Effective in vivo use of adeno-associated virus (AAV)-based vectors to achieve gene-specific silencing or upregulation in the central nervous system has been limited by the inability to provide more than limited deep parenchymal expression in adult animals using delivery routes with the most clinical relevance (intravenous or intrathecal). Here, we demonstrate that the spinal pia membrane represents the primary barrier limiting effective AAV9 penetration into the spinal parenchyma after intrathecal AAV9 delivery. We develop a novel subpial AAV9 delivery technique and AAV9-dextran formulation. We use these in adult rats and pigs to show (i) potent spinal parenchymal transgene expression in white and gray matter including neurons, glial and endothelial cells after single bolus subpial AAV9 delivery; (ii) delivery to almost all apparent descending motor axons throughout the length of the spinal cord after cervical or thoracic subpial AAV9 injection; (iii) potent retrograde transgene expression in brain motor centers (motor cortex and brain stem); and (iv) the relative safety of this approach by defining normal neurological function for up to 6 months after AAV9 delivery. Thus, subpial delivery of AAV9 enables gene-based therapies with a wide range of potential experimental and clinical utilizations in adult animals and human patients. PMID:27462649

  3. Mechanisms of the biphasic effects of peroxides on the retinal vasculature of newborn and adult pigs.

    PubMed

    Abran, D; Hardy, P; Varma, D R; Chemtob, S

    1995-09-01

    We tested whether the ontogenic differences in the constrictor effects of peroxides on the retinal vasculature were modulated by dilator cyclo-oxygenase products. Retinal arteriole (100-200 microns) vasomotor response to H2O2, t-butyl hydroperoxide, and cumene hydroperoxide were studied in isolated eyecup preparations using video camera monitoring of vessel diameter. A time- and dose-dependent biphasic retinal vasomotor response to all peroxides was observed on tissues of newborn and adult pigs. A rapid vasoconstriction (first 2 min) was followed by a relaxation which was greater in the adult than in the newborn tissues. The constrictor as well as the dilator response to peroxides and the observed increase in prostanoids were blocked by the cyclo-oxygenase inhibitor indomethacin. The peroxide-induced relaxation was inhibited or markedly attenuated by the prostaglandin I2 synthase blockers, trans-2-phenyl cyclopropylamine and minoxidil on tissues of newborn and adult animals. These agents also prevented the increase of the prostaglandin I2 receptor-coupled second messenger, cyclic 3',5'-adenosine monophosphate. Our data indicate that prostaglandin I2 plays a major role in counteracting the initial constrictor effects of peroxides in the retinal vasculature, and that the reversal of this constriction is greater in the adult than the newborn. These findings suggest that reduced reversal of vasoconstriction by the dilator prostaglandin I2 during an oxidative stress in the newborn may facilitate vasoconstriction by the dilator prostaglandin I2 during an oxidative stress in the newborn may facilitate neovascularization in retinopathy of prematurity.

  4. High-fat diet-dependent modulation of the delayed rectifier K(+) current in adult guinea pig atrial myocytes.

    PubMed

    Aromolaran, Ademuyiwa S; Colecraft, Henry M; Boutjdir, Mohamed

    2016-06-01

    Obesity is associated with hyperlipidemia, electrical remodeling of the heart, and increased risk of supraventricular arrhythmias in both male and female patients. The delayed rectifier K(+) current (IK), is an important regulator of atrial repolarization. There is a paucity of studies on the functional role of IK in response to obesity. Here, we assessed the obesity-mediated functional modulation of IK in low-fat diet (LFD), and high-fat diet (HFD) fed adult guinea pigs. Guinea pigs were randomly divided into control and obese groups fed, ad libitum, with a LFD (10 kcal% fat) or a HFD (45 kcal% fat) respectively. Action potential duration (APD), and IK were studied in atrial myocytes and IKr and IKs in HEK293 cells using whole-cell patch clamp electrophysiology. HFD guinea pigs displayed a significant increase in body weight, total cholesterol and total triglycerides within 50 days. Atrial APD at 30% (APD30) and 90% (APD90) repolarization were shorter, while atrial IK density was significantly increased in HFD guinea pigs. Exposure to palmitic acid (PA) increased heterologously expressed IKr and IKs densities, while oleic acid (OA), severely reduced IKr and had no effect on IKs. The data are first to show that in obese guinea pigs abbreviated APD is due to increased IK density likely through elevations of PA. Our findings may have crucial implications for targeted treatment options for obesity-related arrhythmias.

  5. Third-Generation Sequencing and Analysis of Four Complete Pig Liver Esterase Gene Sequences in Clones Identified by Screening BAC Library

    PubMed Central

    Zhou, Qiongqiong; Sun, Wenjuan; Liu, Xiyan; Wang, Xiliang; Xiao, Yuncai; Bi, Dingren; Yin, Jingdong; Shi, Deshi

    2016-01-01

    Aim Pig liver carboxylesterase (PLE) gene sequences in GenBank are incomplete, which has led to difficulties in studying the genetic structure and regulation mechanisms of gene expression of PLE family genes. The aim of this study was to obtain and analysis of complete gene sequences of PLE family by screening from a Rongchang pig BAC library and third-generation PacBio gene sequencing. Methods After a number of existing incomplete PLE isoform gene sequences were analysed, primers were designed based on conserved regions in PLE exons, and the whole pig genome used as a template for Polymerase chain reaction (PCR) amplification. Specific primers were then selected based on the PCR amplification results. A three-step PCR screening method was used to identify PLE-positive clones by screening a Rongchang pig BAC library and PacBio third-generation sequencing was performed. BLAST comparisons and other bioinformatics methods were applied for sequence analysis. Results Five PLE-positive BAC clones, designated BAC-10, BAC-70, BAC-75, BAC-119 and BAC-206, were identified. Sequence analysis yielded the complete sequences of four PLE genes, PLE1, PLE-B9, PLE-C4, and PLE-G2. Complete PLE gene sequences were defined as those containing regulatory sequences, exons, and introns. It was found that, not only did the PLE exon sequences of the four genes show a high degree of homology, but also that the intron sequences were highly similar. Additionally, the regulatory region of the genes contained two 720bps reverse complement sequences that may have an important function in the regulation of PLE gene expression. Significance This is the first report to confirm the complete sequences of four PLE genes. In addition, the study demonstrates that each PLE isoform is encoded by a single gene and that the various genes exhibit a high degree of sequence homology, suggesting that the PLE family evolved from a single ancestral gene. Obtaining the complete sequences of these PLE genes

  6. Transition of adult T-cell leukemia/lymphoma clones during clinical progression.

    PubMed

    Aoki, Sakura; Firouzi, Sanaz; López, Yosvany; Yamochi, Tadanori; Nakano, Kazumi; Uchimaru, Kaoru; Utusnomiya, Atae; Iwanaga, Masako; Watanabe, Toshiki

    2016-09-01

    Adult T-cell leukemia/lymphoma (ATLL) is a peripheral T-cell neoplasm caused by the transformation of HTLV-1-infected T cells. ATLL, especially its aggressive form, is known for its poor prognosis, even with intensive chemotherapy. ATLL cells are considered to be monoclonal; however, multiclonal proliferation or emergence of a new clone over time has been reported based on Southern blot analysis, although direct molecular evidence remains elusive. Furthermore, it is thought that clonal change may be a cause of early drug resistance in ATLL. To directly analyze potential clonal changes in ATLL during its clinical course, we used inverse PCR to detect integration sites in combination with a newly developed method using next-generation sequencing, and compared ATLL cell clonality at different time points. The results of inverse PCR indicated that the major clone was altered in three of 19 patients. Together with results from five patients, using this new method, we found direct evidence of clonal change occurring during the clinical course or in response to chemotherapy in ATLL. These results also highlight the importance of clonality analysis for understanding the mechanisms of ATLL development and drug resistance. PMID:27383637

  7. Cloning of calves from various somatic cell types of male and female adult, newborn and fetal cows.

    PubMed

    Kato, Y; Tani, T; Tsunoda, Y

    2000-11-01

    Twenty-four calves were cloned from six somatic cell types of female and male adult, newborn and fetal cows. The clones were derived from female cumulus (n = 3), oviduct (n = 2) and uterine (n = 2) cells, female and male skin cells (n = 10), and male ear (n = 5) and liver (n = 2) cells. On the basis of the number of cloned embryos transferred (n = 172) to surrogate cows, the overall rate of success was 14%, but based on the number of surrogate mothers that became pregnant (n = 50), the success rate was 48%. Cell nuclei from uterus, ear and liver cells, which have not been tested previously, developed into newborn calves after nuclear transfer into enucleated oocytes. To date, seven female and six male calves have survived: six of the females were from adult cells (cumulus (n = 3), oviduct (n = 2) and skin (n = 1) cells) and one was from newborn skin cells, whereas the male calves were derived from adult ear cells (n = 3), newborn liver and skin cells (n = 2), and fetal cells (n = 1). Clones derived from adult cells frequently aborted in the later stages of pregnancy and calves developing to term showed a higher number of abnormalities than did those derived from newborn or fetal cells. The telomeric DNA lengths in the ear cells of three male calves cloned from the ear cells of a bull aged 10 years were similar to those of the original bull. However, the telomeric DNA lengths from the white blood cells of the clones, although similar to those in an age-matched control, were shorter than those of the original bull, which indicates that telomeric shortening varies among tissues.

  8. Adult-brain-derived neural stem cells grafting into a vein bridge increases postlesional recovery and regeneration in a peripheral nerve of adult pig.

    PubMed

    Liard, Olivier; Segura, Stéphanie; Sagui, Emmanuel; Nau, André; Pascual, Aurélie; Cambon, Melissa; Darlix, Jean-Luc; Fusai, Thierry; Moyse, Emmanuel

    2012-01-01

    We attempted transplantation of adult neural stem cells (ANSCs) inside an autologous venous graft following surgical transsection of nervis cruralis with 30 mm long gap in adult pig. The transplanted cell suspension was a primary culture of neurospheres from adult pig subventricular zone (SVZ) which had been labeled in vitro with BrdU or lentivirally transferred fluorescent protein. Lesion-induced loss of leg extension on the thigh became definitive in controls but was reversed by 45-90 days after neurosphere-filled vein grafting. Electromyography showed stimulodetection recovery in neurosphere-transplanted pigs but not in controls. Postmortem immunohistochemistry revealed neurosphere-derived cells that survived inside the venous graft from 10 to 240 post-lesion days and all displayed a neuronal phenotype. Newly formed neurons were distributed inside the venous graft along the severed nerve longitudinal axis. Moreover, ANSC transplantation increased CNPase expression, indicating activation of intrinsic Schwann cells. Thus ANSC transplantation inside an autologous venous graft provides an efficient repair strategy. PMID:22448170

  9. Adult-Brain-Derived Neural Stem Cells Grafting into a Vein Bridge Increases Postlesional Recovery and Regeneration in a Peripheral Nerve of Adult Pig

    PubMed Central

    Liard, Olivier; Segura, Stéphanie; Sagui, Emmanuel; Nau, André; Pascual, Aurélie; Cambon, Melissa; Darlix, Jean-Luc; Fusai, Thierry; Moyse, Emmanuel

    2012-01-01

    We attempted transplantation of adult neural stem cells (ANSCs) inside an autologous venous graft following surgical transsection of nervis cruralis with 30 mm long gap in adult pig. The transplanted cell suspension was a primary culture of neurospheres from adult pig subventricular zone (SVZ) which had been labeled in vitro with BrdU or lentivirally transferred fluorescent protein. Lesion-induced loss of leg extension on the thigh became definitive in controls but was reversed by 45–90 days after neurosphere-filled vein grafting. Electromyography showed stimulodetection recovery in neurosphere-transplanted pigs but not in controls. Postmortem immunohistochemistry revealed neurosphere-derived cells that survived inside the venous graft from 10 to 240 post-lesion days and all displayed a neuronal phenotype. Newly formed neurons were distributed inside the venous graft along the severed nerve longitudinal axis. Moreover, ANSC transplantation increased CNPase expression, indicating activation of intrinsic Schwann cells. Thus ANSC transplantation inside an autologous venous graft provides an efficient repair strategy. PMID:22448170

  10. Nano-crystalline diamond-coated titanium dental implants - a histomorphometric study in adult domestic pigs.

    PubMed

    Metzler, Philipp; von Wilmowsky, Cornelius; Stadlinger, Bernd; Zemann, Wolfgang; Schlegel, Karl Andreas; Rosiwal, Stephan; Rupprecht, Stephan

    2013-09-01

    Promising biomaterial characteristics of diamond-coatings in biomedicine have been described in the literature. However, there is a lack of knowledge about implant osseointegration of this surface modification compared to the currently used sandblasted acid-etched Ti-Al6-V4 implants. The aim of this study was to investigate the osseointegration of microwave plasma-chemical-vapour deposition (MWP-CVD) diamond-coated Ti-Al6-V4 dental implants after healing periods of 2 and 5 months. Twenty-four MWP-CVD diamond-coated and 24 un-coated dental titanium-alloy implants (Ankylos(®)) were placed in the frontal skull of eight adult domestic pigs. To evaluate the effects of the nano-structured surfaces on bone formation, a histomorphometric analysis was performed after 2 and 5 months of implant healing. Histomorphometry analysed the bone-to-implant contact (BIC). No significant difference in BIC for the diamond-coated implants in comparison to reference implants could be observed for both healing periods. Scanning electron microscopy revealed an adequate interface between the bone and the diamond surface. No delamination or particle-dissociation due to shearing forces could be detected. In this study, diamond-coated dental titanium-alloy implants and sandblasted acid-etched implants showed a comparable degree of osseointegration.

  11. A low α-linolenic intake during early life increases adiposity in the adult guinea pig

    PubMed Central

    2010-01-01

    Background The composition of dietary fatty acids (FA) during early life may impact adult adipose tissue (AT) development. We investigated the effects of α-linolenic acid (ALA) intake during the suckling/weaning period on AT development and metabolic markers in the guinea pig (GP). Methods Newborn GP were fed a 27%-fat diet (w/w %) with high (10%-ALA group), moderate (2.4%-ALA group) or low (0.8%-ALA group) ALA content (w/w % as total FA) until they were 21 days old (d21). Then all animals were switched to a 15%-fat diet containing 2% ALA (as total FA) until 136 days of age (d136). Results ALA and docosapentaenoic acid measured in plasma triglycerides (TG) at d21 decreased with decreasing ALA intake. Total body fat mass was not different between groups at d21. Adipose tissue TG synthesis rates and proliferation rate of total adipose cells, as assessed by 2H2O labelling, were unchanged between groups at d21, while hepatic de novo lipogenesis was significantly 2-fold increased in the 0.8%-ALA group. In older GP, the 0.8%-ALA group showed a significant 15-%-increased total fat mass (d79 and d107, p < 0.01) and epididymal AT weight (d136) and tended to show higher insulinemia compared to the 10%-ALA group. In addition, proliferation rate of cells in the subcutaneous AT was higher in the 0.8%-ALA (15.2 ± 1.3% new cells/5d) than in the 10%-ALA group (8.6 ± 1.7% new cells/5d, p = 0.021) at d136. AT eicosanoid profiles were not associated with the increase of AT cell proliferation. Conclusion A low ALA intake during early postnatal life promotes an increased adiposity in the adult GP. PMID:20205840

  12. A novel method for the production of transgenic cloned pigs: electroporation-mediated gene transfer to non-cultured cells and subsequent selection with puromycin.

    PubMed

    Watanabe, Satoshi; Iwamoto, Masaki; Suzuki, Shun-ichi; Fuchimoto, Daiichiro; Honma, Daisuke; Nagai, Takashi; Hashimoto, Michiko; Yazaki, Satoko; Sato, Masahiro; Onishi, Akira

    2005-02-01

    Puromycin N-acetyl transferase gene (pac), of which the gene product catalyzes antibiotic puromycin (an effective inhibitor of protein synthesis), has been widely used as a dominant selection marker in embryonic stem (ES) cell-mediated transgenesis. The present study is the first to report on the usefulness of puromycin for production of enhanced green fluorescent protein (EGFP) transgenic piglets after somatic cell cloning and embryo transfer. Somatic cells isolated from porcine fetuses at 73 days of gestation were immediately electroporated with a transgene (pCAG-EGFPac) carrying both EGFP cDNA and pac. This procedure aims to avoid aging effects thought to be generated during cell culture. The recombinant cells were selected with puromycin at a low concentration (2 microg/ml), cultured for 7 days, and then screened for EGFP expression before somatic cell cloning. The manipulated embryos were transplanted into the oviducts of 14 foster mother sows. Four of the foster sows became pregnant and nine piglets were delivered. Of the nine piglets, eight died shortly after birth and one grew healthy after weaning. Results indicate that puromycin can be used for the selection of recombinant cells from noncultured cells, and moreover, may confer the production of genetically engineered newborns via nuclear transfer techniques in pigs. PMID:15385422

  13. Immunolocalization of MAP-2 in Routinely Formalin-Fixed, Paraffin-Embedded Guinea Pig Brain Sections Using Microwave Irradiation: A Comparison of Different Combinations of Antibody Clones and Antigen Retrieval Buffer Solutions

    NASA Astrophysics Data System (ADS)

    Kan, Robert K.; Pleva, Christina M.; Hamilton, Tracey A.; Petrali, John P.

    2005-04-01

    The present study was designed to evaluate the efficacy of different microwave pretreatment methods to retrieve microtubule-associated protein 2 (MAP-2) immunoreactivity in formalin-fixed, paraffin-embedded guinea pig brain sections. Brain sections, microwave pretreated in boiling sodium citrate, citric acid, Tris hydrochloride, and EDTA buffers of pH 4, 6, and 8, were labeled with four different clones of MAP-2 monoclonal antibodies. No MAP-2 immunoreactivity was observed in control sections processed without microwave pretreatment. Optimal MAP-2 immunoreactivity was observed only when MAP-2 antibody clone AP18 was used in conjunction with citric acid buffer of pH 6.0. Using this combination, brain sections from nerve agent soman-exposed guinea pigs were found to exhibit marked reduction in MAP-2 immunostaining in the hippocampus. These observations suggest that the clone of the antibody in addition to the type and pH of antigen retrieval (AR) solution are important variables to be considered for establishing an optimal AR technique. When studying counterpart antigens of species other than that to which the antibodies were originally raised, different antibody clones must be tested in combination with different microwave-assisted AR (MAR) methods. This MAR method makes it possible to conduct retrospective studies on archival guinea pig brain paraffin blocks to evaluate changes in neuronal MAP-2 expression as a consequence of chemical warfare nerve agent toxicity.

  14. 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.

  15. Cloned cDNA to cholecystokinin mRNA predicts an identical preprocholecystokinin in pig brain and gut.

    PubMed Central

    Gubler, U; Chua, A O; Hoffman, B J; Collier, K J; Eng, J

    1984-01-01

    Molecular cloning has established the structure of preprocholecystokinin in porcine cerebral cortex and duodenal mucosa. This precursor is 114 amino acids long, is identical in brain and gut, contains all the cholecystokinin (CCK) peptides previously isolated, and has the characteristics of a prohormone. It contains a putative amino-terminal signal peptide, basic processing sites, and a carboxyl-terminal amidation signal. The CCK mRNAs from brain and gut are approximately 850 nucleotides long and differ by only a few single base changes. This analysis establishes by a strict criterion that CCK is synthesized in both brain and gut and that the different distributions of molecular forms of CCK in the two tissues are most probably a consequence of tissue-specific posttranslational events. Images PMID:6205394

  16. Volume-activated Na/H exchange activity in fetal and adult pig red cells: inhibition by cyclic AMP.

    PubMed

    Sergeant, S; Sohn, D H; Kim, H D

    1989-08-01

    Hyposmotic swelling of pig red cells leads to a selective increase in K permeability, whereas hyperosmotic cell shrinkage augments the Na permeability. In this regard, the ouabain-resistant (OR) Na flux of red cells of newborn and adult pigs is characterized in detail. A reduction in cell volume by approximately 18% leads to an increase in the OR Na efflux of fetal and adult cells by 15- and fourfold, respectively. The OR Na influx in both cell types is equally influenced by cell shrinkage. Depletion of cellular K does not influence the volume-activated OR Na efflux. Nor does OR Na influx require external K. Both OR Na efflux and influx activated by shrinkage are inhibited by the diuretics furosemide and amiloride. The rank order of decreasing anion sensitivity for diuretic-sensitive Na efflux was acetate greater than chloride greater than gluconate greater than nitrate. Cell shrinkage induced by the addition of hypertonic salts results in an acidification of the unbuffered and CO2-free media, provided that both Na and DIDS are present. The acidification process can be reversed by either of the diuretic agents. These findings suggest that the shrinkage-activated OR Na flux is primarily mediated by a Na/H exchanger rather than by a Na/K/Cl cotransporter. Once loaded with either cAMP or cGMP, cell swelling can no longer activate the Na/H exchanger. The Na/H exchanger activity is detectable in the fetal cells of normal volume but quiescent in adult cells, indicating that the exchanger undergoes a developmental change during the transition from the fetal to adult stage. PMID:2552123

  17. Chronic prenatal ethanol exposure alters expression of central and peripheral insulin signaling molecules in adult guinea pig offspring.

    PubMed

    Dobson, Christine C; Thevasundaram, Kersh; Mongillo, Daniel L; Winterborn, Andrew; Holloway, Alison C; Brien, James F; Reynolds, James N

    2014-11-01

    Maternal ethanol consumption during pregnancy can produce a range of teratogenic outcomes in offspring. The mechanism of ethanol teratogenicity is multi-faceted, but may involve alterations in insulin and insulin-like growth factor (IGF) signaling pathways. These pathways are not only important for metabolism, but are also critically involved in neuronal survival and plasticity, and they can be altered by chronic prenatal ethanol exposure (CPEE). The objective of this study was to test the hypothesis that CPEE alters expression of insulin and IGF signaling molecules in the prefrontal cortex and liver of adult guinea pig offspring. Pregnant Dunkin-Hartley-strain guinea pigs received ethanol (4 g/kg maternal body weight/day) or isocaloric-sucrose/pair-feeding (nutritional control) throughout gestation. Fasting blood glucose concentration was measured in male and female offspring at postnatal day 150-200, followed by euthanasia, collection of prefrontal cortex and liver, and RNA extraction. IGF-1, IGF-1 receptor (IGF-1R), IGF-2, IGF-2 receptor (IGF-2R), insulin receptor substrate (IRS)-1, IRS-2, and insulin receptor (INSR) mRNA expression levels were measured in tissues using quantitative real-time PCR. The mean maternal blood ethanol concentration was 281 ± 15 mg/dL at 1 h after the second divided dose of ethanol on GD 57. CPEE resulted in increased liver weight in adult offspring, but produced no difference in fasting blood glucose concentration compared with nutritional control. In the liver, CPEE decreased mRNA expression of IGF-1, IGF-1R, and IGF-2, and increased IRS-2 mRNA expression in male offspring only compared with nutritional control. Female CPEE offspring had decreased INSR hepatic mRNA expression compared with male CPEE offspring. In the prefrontal cortex, IRS-2 mRNA expression was increased in CPEE offspring compared with nutritional control. The data demonstrate that CPEE alters both central and peripheral expression of insulin and IGF signaling

  18. Chronic prenatal ethanol exposure alters expression of central and peripheral insulin signaling molecules in adult guinea pig offspring.

    PubMed

    Dobson, Christine C; Thevasundaram, Kersh; Mongillo, Daniel L; Winterborn, Andrew; Holloway, Alison C; Brien, James F; Reynolds, James N

    2014-11-01

    Maternal ethanol consumption during pregnancy can produce a range of teratogenic outcomes in offspring. The mechanism of ethanol teratogenicity is multi-faceted, but may involve alterations in insulin and insulin-like growth factor (IGF) signaling pathways. These pathways are not only important for metabolism, but are also critically involved in neuronal survival and plasticity, and they can be altered by chronic prenatal ethanol exposure (CPEE). The objective of this study was to test the hypothesis that CPEE alters expression of insulin and IGF signaling molecules in the prefrontal cortex and liver of adult guinea pig offspring. Pregnant Dunkin-Hartley-strain guinea pigs received ethanol (4 g/kg maternal body weight/day) or isocaloric-sucrose/pair-feeding (nutritional control) throughout gestation. Fasting blood glucose concentration was measured in male and female offspring at postnatal day 150-200, followed by euthanasia, collection of prefrontal cortex and liver, and RNA extraction. IGF-1, IGF-1 receptor (IGF-1R), IGF-2, IGF-2 receptor (IGF-2R), insulin receptor substrate (IRS)-1, IRS-2, and insulin receptor (INSR) mRNA expression levels were measured in tissues using quantitative real-time PCR. The mean maternal blood ethanol concentration was 281 ± 15 mg/dL at 1 h after the second divided dose of ethanol on GD 57. CPEE resulted in increased liver weight in adult offspring, but produced no difference in fasting blood glucose concentration compared with nutritional control. In the liver, CPEE decreased mRNA expression of IGF-1, IGF-1R, and IGF-2, and increased IRS-2 mRNA expression in male offspring only compared with nutritional control. Female CPEE offspring had decreased INSR hepatic mRNA expression compared with male CPEE offspring. In the prefrontal cortex, IRS-2 mRNA expression was increased in CPEE offspring compared with nutritional control. The data demonstrate that CPEE alters both central and peripheral expression of insulin and IGF signaling

  19. Using adult cloned trees grown under natural conditions to characterize BVOC emission variation

    NASA Astrophysics Data System (ADS)

    Persson, Ylva; Schurgers, Guy; Ekberg, Anna; Rinnan, Riikka; Holst, Thomas

    2015-04-01

    Biogenic Volatile Organic Compounds (BVOCs) are diverse chemical species produced and emitted from the vegetation as trace gases. BVOCs are commonly grouped into isoprene, monoterpenes and sesquiterpenes, where isoprene is mainly emitted by deciduous trees and monoterpenes and sesquiterpenes by coniferous trees. BVOCs are known to have a considerable impact on atmospheric chemistry and are precursors for secondary organic aerosol, which in turn are important for the aerosol feedback on the Earth's climate. Recently, Bäck et al. (2012) reported a high diversity of the chemical composition of emitted compounds from pine trees growing at the same stand due to genetic variation. This study here uses cloned trees growing naturally in a transect in Europe in order to exclude genetic variation and to assess emission variation between and within selected tree species grown at different climatic conditions. The International Phenological Garden (IPG) network, where cloned trees are used to monitor the long-term phenological observations of representative tree species for Europe provides a specific, cloned set of important tree species, which had been planted throughout Europe starting in 1957. This gives a unique opportunity to study the adaptation to various climatic conditions and field conditions in genetically identical plants in relation to BVOC emissions. During a field campaign in 2013 at the IPG site in Taastrup, Denmark (55°40' N, 12°18' E), seven trees were measured at three heights within the canopy. Measured trees were two English oaks (Quercus robur), one European beech (Fagus sylvatica) and four Norway spruces (Picea abies) of two provenances. For oak and one provenance of spruce, measurements were performed twice, both in June and in August in order to examine any emission pattern change with the progression of the summer. Measurements were performed using a gas-exchange cuvette of a photosynthesis system combined with BVOC adsorbent tubes, which were

  20. Chronic intermittent hypobaric hypoxia protects the heart against ischemia/reperfusion injury through upregulation of antioxidant enzymes in adult guinea pigs

    PubMed Central

    Guo, Hui-cai; Zhang, Zhe; Zhang, Li-nan; Xiong, Chen; Feng, Chen; Liu, Qian; Liu, Xu; Shi, Xiao-lu; Wang, Yong-li

    2009-01-01

    Aim: To investigate the protection and the anti-oxidative mechanism afforded by chronic intermittent hypobaric hypoxia (CIHH) against ischemia/reperfusion (I/R) injury in guinea pig hearts. Methods: Adult male guinea pigs were exposed to CIHH by mimicking a 5000 m high altitude (pB=404 mmHg, pO2=84 mmHg) in a hypobaric chamber for 6 h/day for 28 days. Langendorff-perfused isolated guinea pig hearts were used to measure variables of left ventricular function during baseline perfusion, ischemia and the reperfusion period. The activity and protein expression of antioxidant enzymes in the left myocardium were evaluated using biochemical methods and Western blotting, respectively. Intracellular reactive oxygen species (ROS) were assessed using ROS-sensitive fluorescence. Results: After 30 min of global no-flow ischemia followed by 60 min of reperfusion, myocardial function had better recovery rates in CIHH guinea pig hearts than in control hearts. The activity and protein expression of superoxide dismutase (SOD) and catalase (CAT) were significantly increased in the myocardium of CIHH guinea pigs. Pretreatment of control hearts with an antioxidant mixture containing SOD and CAT exerted cardioprotective effects similar to CIHH. The irreversible CAT inhibitor aminotriazole (ATZ) abolished the cardioprotection of CIHH. Cardiac contractile dysfunction and oxidative stress induced by exogenous hydrogen peroxide (H2O2) were attenuated by CIHH and CAT. Conclusions: These data suggest that CIHH protects the heart against I/R injury through upregulation of antioxidant enzymes in guinea pig. PMID:19543301

  1. Nuclear cloning, epigenetic reprogramming and cellular differentiation.

    PubMed

    Jaenisch, Rudolf; Hochedlinger, Konrad; Eggan, Kevin

    2005-01-01

    The full-term development of sheep, cows, goats, pigs and mice has been achieved through the transfer of somatic cell nuclei into enucleated oocytes. Despite these successes, mammalian cloning remains an inefficient process, with a preponderance of reconstructed embryos failing at early- to mid-gestation stages of development. The small percentage of conceptuses that survive to term are characterized by a high mortality rate and frequently display grossly increased placental and birth weights. It is likely that inappropriate expression of key developmental genes may contribute to lethality of cloned embryos. One of the most interesting issues of nuclear cloning is the question of genomic reprogramming, i.e. whether successful cloning requires the resetting of epigenetic modifications which are characteristic of the adult donor nucleus. Processes such as X-inactivation and genomic imprinting are known to depend on epigenetic modifications of the genome. The classical nuclear transfer experiments with frogs have suggested that the source of the donor nucleus affects the phenotype of the clone. We have, using expression profiling, compared gene expression in clones derived from embryonic stem (ES) cells and from somatic donor cell nuclei and find substantial gene dysregulation. Our results suggest that faulty reprogramming is caused by the nuclear cloning procedure itself. In addition, the type of donor nucleus contributes to the abnormal expression pattern seen in cloned animals. One of the major unresolved issues has been whether nuclei of terminally differentiated cells can be reprogrammed by transfer into the oocyte. To address this question we have derived monoclonal mice from B and T cells and used the genetic rearrangements of the immunoglobulin and T cell receptor genes to retrospectively verify the differentiation state of the donor nucleus. Finally, we discuss our recent studies on the reprogramming of nuclei from terminally differentiated neurons and from

  2. "We Are Guinea Pigs Really": Examining the Realities of ICT-based Adult Learning.

    ERIC Educational Resources Information Center

    Selwyn, Neil; Gorard, Stephen; Williams, Sara

    2002-01-01

    Interviews with 36 adult learners in information-communications technology (ICT) settings, using the concept of learning trajectories, revealed barriers to widening participation through ICT. In addition to technical shortcomings, social, economic, cultural, and political issues hinder the process, including lack of innovative instruction and…

  3. Investigation of Pharmacological Activity of Caralluma penicillata: Anti-Inflammatory Properties and Gastritis Protection against Indomethacin in Adult Guinea Pigs.

    PubMed

    Albaser, Nabil; Ghanem, Najeeb; Shehab, Mohanad; Al-Adhal, Adnan; Amood Al-Kamarany, Mohammed

    2014-01-01

    Caralluma is a plant that possessing a great therapeutic potential in folk medicine in Yemen, namely, Caralluma penicillata (C. penicillata) as antiulcer. The study aims to evaluate the anti-inflammatory properties and gastritis protection activity of C. penicillata against indomethacin in adult guinea pigs. The study was divided into four parts: firstly, the optimum dose of extract as anti-inflammatory effect was determined. Secondly, the acute anti-inflammatory effect of extract were estimated. Thirdly, the repeated doses of extract against chronic inflammation was estimated. The anti-inflammatory activity of extract was compared with indomethacin as a prototype of drug against inflammation. Fourthly, the gastritis protection properties of extract with/without indomethacin were performed. The results showed that a 400 mg/kg of 10% ethanol extract produced the maximum of anti-inflammatory effect. Also, the single dose of extract was equipotent for indomethacin (10 mg/kg), but shorter in duration with regard to acute anti-inflammatory effect. In addition, the repeated doses of extract against chronic inflammation were less potent than indomethacin with regard to ulcerogenic effect. On the other hand, extract-indomethacin combination reduced the gastritis effect of indomethacin based on ulcer index and histological study.

  4. Reinnervation by axon collaterals from single facial motoneurons to multiple muscle targets following axotomy in the adult guinea pig.

    PubMed

    Ito, M; Kudo, M

    1994-01-01

    To study the process of recovery from facial palsy experimentally, the location of cranial motoneurons supplying the posterior belly of the digastric muscle (PDG) and the extratemporal portion of the facial nerve trunk was examined in a double-labeling paradigm using two retrograde tracers in the adult guinea pig of which the facial nerve had been surgically injured. In different stages after the induced facial palsy had recovered functionally (4-13 weeks after the surgical operation), wheat germ-agglutinated horseradish peroxidase (WGA-HRP) was injected into the PDG and Fluoro-Ruby (FR) was applied to the proximal cut end of the extratemporal portion of the facial nerve trunk. Distribution of neurons retrogradely labeled with WGA-HRP and/or FR was plotted in the brainstem and compared with that of the controls. In the intact cases, HRP-labeled neurons were restrictedly seen in the accessory facial nucleus (Acs7), while FR-labeled neurons were found within the main facial nucleus (FMN). In the axotomized cases: (1) HRP-labeled neurons were seen diffusely in the Acs7 as well as in the FMN, where normal myotopical representation no longer seemed to be maintained. (2) FR-labeled neurons were also observed diffusely in the FMN and the Acs7. (3) A considerable number of neurons were doubly labeled with WGA-HRP and FR in both the Acs and the FMN in cases with shorter survival periods (4-7 weeks), but not in cases with longer survival periods (12-13 weeks). Thus, new findings show that connections are temporarily maintained by single, facial motoneurons with axon collaterals to multiple muscle targets in adult mammals.(ABSTRACT TRUNCATED AT 250 WORDS)

  5. Purification and molecular cloning of aspartic proteinases from the stomach of adult Japanese fire belly newts, Cynops pyrrhogaster.

    PubMed

    Nagasawa, Tatsuki; Sano, Kaori; Kawaguchi, Mari; Kobayashi, Ken-Ichiro; Yasumasu, Shigeki; Inokuchi, Tomofumi

    2016-04-01

    Six aspartic proteinase precursors, a pro-cathepsin E (ProCatE) and five pepsinogens (Pgs), were purified from the stomach of adult newts (Cynops pyrrhogaster). On sodium dodecylsulfate-polyacrylamide gel electrophoresis, the molecular weights of the Pgs and active enzymes were 37-38 kDa and 31-34 kDa, respectively. The purified ProCatE was a dimer whose subunits were connected by a disulphide bond. cDNA cloning by polymerase chain reaction and subsequent phylogenetic analysis revealed that three of the purified Pgs were classified as PgA and the remaining two were classified as PgBC belonging to C-type Pg. Our results suggest that PgBC is one of the major constituents of acid protease in the urodele stomach. We hypothesize that PgBC is an amphibian-specific Pg that diverged during its evolutional lineage. PgBC was purified and characterized for the first time. The purified urodele pepsin A was completely inhibited by equal molar units of pepstatin A. Conversely, the urodele pepsin BC had low sensitivity to pepstatin A. In acidic condition, the activation rates of newt pepsin A and BC were similar to those of mammalian pepsin A and C1, respectively. Our results suggest that the enzymological characters that distinguish A- and C-type pepsins appear to be conserved in mammals and amphibians. PMID:26711235

  6. Purification and molecular cloning of aspartic proteinases from the stomach of adult Japanese fire belly newts, Cynops pyrrhogaster.

    PubMed

    Nagasawa, Tatsuki; Sano, Kaori; Kawaguchi, Mari; Kobayashi, Ken-Ichiro; Yasumasu, Shigeki; Inokuchi, Tomofumi

    2016-04-01

    Six aspartic proteinase precursors, a pro-cathepsin E (ProCatE) and five pepsinogens (Pgs), were purified from the stomach of adult newts (Cynops pyrrhogaster). On sodium dodecylsulfate-polyacrylamide gel electrophoresis, the molecular weights of the Pgs and active enzymes were 37-38 kDa and 31-34 kDa, respectively. The purified ProCatE was a dimer whose subunits were connected by a disulphide bond. cDNA cloning by polymerase chain reaction and subsequent phylogenetic analysis revealed that three of the purified Pgs were classified as PgA and the remaining two were classified as PgBC belonging to C-type Pg. Our results suggest that PgBC is one of the major constituents of acid protease in the urodele stomach. We hypothesize that PgBC is an amphibian-specific Pg that diverged during its evolutional lineage. PgBC was purified and characterized for the first time. The purified urodele pepsin A was completely inhibited by equal molar units of pepstatin A. Conversely, the urodele pepsin BC had low sensitivity to pepstatin A. In acidic condition, the activation rates of newt pepsin A and BC were similar to those of mammalian pepsin A and C1, respectively. Our results suggest that the enzymological characters that distinguish A- and C-type pepsins appear to be conserved in mammals and amphibians.

  7. Long-term effect on in vitro cloning efficiency after treatment of somatic cells with Xenopus egg extract in the pig.

    PubMed

    Liu, Ying; Ostrup, Olga; Li, Rong; Li, Juan; Vajta, Gábor; Kragh, Peter M; Schmidt, Mette; Purup, Stig; Hyttel, Poul; Klærke, Dan; Callesen, Henrik

    2014-08-01

    In somatic cell nuclear transfer (SCNT), donor cell reprogramming is considered as a biologically important and vulnerable event. Various donor cell pre-treatments with Xenopus egg extracts can promote reprogramming. Here we investigated if the reprogramming effect of one treatment with Xenopus egg extract on donor cells was maintained for several cell passages. The extract treatment resulted in increased cell-colony formation from early passages in treated porcine fibroblasts (ExTES), and increased development of cloned embryos. Partial dedifferentiation was observed in ExTES cells, shown as a tendency towards upregulation of NANOG, c-MYC and KLF-4 and downregulation of DESMIM compared with ExTES at Passage 2. Compared with our routine SCNT, continuously increased development of cloned embryos was observed in the ExTES group, and ExTES cloned blastocysts displayed hypermethylated DNA patterns and hypermethylation of H3K4me3 and H3K27me3 in ICM compared with TE. All seven recipients became pregnant after transferral of ExTES cloned embryos and gave birth to 7-22 piglets per litter (average 12). In conclusion, our results demonstrate that one treatment of porcine fibroblasts with Xenopus egg extract can result in long-term increased ability of the cells to promote their in vitro function in subsequent SCNT. Finally these cells can also result in successful development of cloned embryos to term.

  8. 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

  9. 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

  10. First report of Metastrongylus pudendotectus by the genetic characterization of mitochondria genome of cox1 in pigs from Tibet, China.

    PubMed

    Li, Kun; Luo, Houqiang; Zhang, Hui; Lan, Yanfang; Han, Zhaoqing; Shahzad, Muhammad; Wang, Xiaoqiang; Qiu, Gang; Huang, Shucheng; Jiang, Wenteng; Li, Jiakui

    2016-06-15

    Lungworms, a world wild distributed parasites cause serious respiratory diseases to the pigs. A high infection rate of Metastrongylus lungworms has been found in Tibetan pigs being slaughtered in different slaughter houses of Tibet autonomous region. The main aim of our study was to detect and confirm the lungworm parasite by the genetic characterization of mitochondrial cox1genome isolated from the lungs of Tibetan pig. The adult lungworms were collected from the lungs of slaughtered pigs and identification was done through morphological examinations. Total genomic DNA of the extracted worms was performed and a fragment (∼450bp) of the cox1 of mitochondrial (mt) gene was amplified. Amplicons were cloned into PGEM(®)-T Easy vector and the positive clones were sequenced from a commercial company. Sequence and phylogenetic analysis were performed by software of DNAMAN and MEGA respectively. The results revealed that the lungworms infecting the Tibetan pigs were Metastrongylus pudendotectus (M. pudendotectus). To our knowledge, this is the first report for the isolation and identification for the genetic characterization of mitochondria (mt) genome of cox1 of M. pudendotectus derived from Tibetan pigs in Tibet, China. PMID:27198783

  11. First report of Metastrongylus pudendotectus by the genetic characterization of mitochondria genome of cox1 in pigs from Tibet, China.

    PubMed

    Li, Kun; Luo, Houqiang; Zhang, Hui; Lan, Yanfang; Han, Zhaoqing; Shahzad, Muhammad; Wang, Xiaoqiang; Qiu, Gang; Huang, Shucheng; Jiang, Wenteng; Li, Jiakui

    2016-06-15

    Lungworms, a world wild distributed parasites cause serious respiratory diseases to the pigs. A high infection rate of Metastrongylus lungworms has been found in Tibetan pigs being slaughtered in different slaughter houses of Tibet autonomous region. The main aim of our study was to detect and confirm the lungworm parasite by the genetic characterization of mitochondrial cox1genome isolated from the lungs of Tibetan pig. The adult lungworms were collected from the lungs of slaughtered pigs and identification was done through morphological examinations. Total genomic DNA of the extracted worms was performed and a fragment (∼450bp) of the cox1 of mitochondrial (mt) gene was amplified. Amplicons were cloned into PGEM(®)-T Easy vector and the positive clones were sequenced from a commercial company. Sequence and phylogenetic analysis were performed by software of DNAMAN and MEGA respectively. The results revealed that the lungworms infecting the Tibetan pigs were Metastrongylus pudendotectus (M. pudendotectus). To our knowledge, this is the first report for the isolation and identification for the genetic characterization of mitochondria (mt) genome of cox1 of M. pudendotectus derived from Tibetan pigs in Tibet, China.

  12. Comparison of liver mitochondrial proteins derived from newborn cloned calves and from cloned adult cattle by two-dimensional differential gel electrophoresis.

    PubMed

    Takeda, Kumiko; Tasai, Mariko; Akagi, Satoshi; Watanabe, Shinya; Oe, Mika; Chikuni, Koichi; Ohnishi-Kameyama, Mayumi; Hanada, Hirofumi; Nakamura, Yoshiaki; Tagami, Takahiro; Nirasawa, Keijiro

    2011-04-01

    Aberrant reprogramming of donor somatic cell nuclei may result in many severe problems in animal cloning. The inability to establish functional interactions between donor nucleus and recipient mitochondria is also likely responsible for such a developmental deficiency. However, detailed knowledge of protein expression during somatic cell nuclear transfer (SCNT) in cattle is lacking. In the present study, variations in mitochondrial protein levels between SCNT-derived and control cattle, and from calves derived by artificial insemination were investigated. Mitochondrial fractions were prepared from frozen liver samples and subjected to two-dimensional (2-D) fluorescence differential gel electrophoresis (DIGE) using CyDye™ dyes. Protein expression changes were confirmed with a volume ratio greater than 2.0 (P < 0.05). 2D-DIGE analysis revealed differential expression of three proteins for SCNT cattle (n = 4) and seven proteins for SCNT calves (n = 6) compared to controls (P < 0.05). Different protein patterning was observed among SCNT animals even if animals were generated from the same donor cell source. No differences were detected in two of the SCNT cattle. Moreover, there was no novel protein identified in any of the SCNT cattle or calves. In conclusion, variation in mitochondrial protein expression concentrations was observed in non-viable, neonatal SCNT calves and among SCNT individuals. This result implicates mitochondrial-related gene expression in early developmental loss of SCNT embryos. Comparative proteomic analysis represents an important tool for further studies on SCNT animals.

  13. An mRNA Vaccine Encoding Rabies Virus Glycoprotein Induces Protection against Lethal Infection in Mice and Correlates of Protection in Adult and Newborn Pigs

    PubMed Central

    Voss, Daniel; Petsch, Benjamin; Baumhof, Patrick; Kramps, Thomas; Stitz, Lothar

    2016-01-01

    Rabies is a zoonotic infectious disease of the central nervous system (CNS). In unvaccinated or untreated subjects, rabies virus infection causes severe neurological symptoms and is invariably fatal. Despite the long-standing existence of effective vaccines, vaccine availability remains insufficient, with high numbers of fatal infections mostly in developing countries. Nucleic acid based vaccines have proven convincingly as a new technology for the fast development of vaccines against newly emerging pathogens, diseases where no vaccine exists or for replacing already existing vaccines. We used an optimized non-replicating rabies virus glycoprotein (RABV-G) encoding messenger RNA (mRNA) to induce potent neutralizing antibodies (VN titers) in mice and domestic pigs. Functional antibody titers were followed in mice for up to one year and titers remained stable for the entire observation period in all dose groups. T cell analysis revealed the induction of both, specific CD4+ as well as CD8+ T cells by RABV-G mRNA, with the induced CD4+ T cells being higher than those induced by a licensed vaccine. Notably, RABV-G mRNA vaccinated mice were protected against lethal intracerebral challenge infection. Inhibition of viral replication by vaccination was verified by qRT-PCR. Furthermore, we demonstrate that CD4+ T cells are crucial for the generation of neutralizing antibodies. In domestic pigs we were able to induce VN titers that correlate with protection in adult and newborn pigs. This study demonstrates the feasibility of a non-replicating mRNA rabies vaccine in small and large animals and highlights the promises of mRNA vaccines for the prevention of infectious diseases. PMID:27336830

  14. An mRNA Vaccine Encoding Rabies Virus Glycoprotein Induces Protection against Lethal Infection in Mice and Correlates of Protection in Adult and Newborn Pigs.

    PubMed

    Schnee, Margit; Vogel, Annette B; Voss, Daniel; Petsch, Benjamin; Baumhof, Patrick; Kramps, Thomas; Stitz, Lothar

    2016-06-01

    Rabies is a zoonotic infectious disease of the central nervous system (CNS). In unvaccinated or untreated subjects, rabies virus infection causes severe neurological symptoms and is invariably fatal. Despite the long-standing existence of effective vaccines, vaccine availability remains insufficient, with high numbers of fatal infections mostly in developing countries. Nucleic acid based vaccines have proven convincingly as a new technology for the fast development of vaccines against newly emerging pathogens, diseases where no vaccine exists or for replacing already existing vaccines. We used an optimized non-replicating rabies virus glycoprotein (RABV-G) encoding messenger RNA (mRNA) to induce potent neutralizing antibodies (VN titers) in mice and domestic pigs. Functional antibody titers were followed in mice for up to one year and titers remained stable for the entire observation period in all dose groups. T cell analysis revealed the induction of both, specific CD4+ as well as CD8+ T cells by RABV-G mRNA, with the induced CD4+ T cells being higher than those induced by a licensed vaccine. Notably, RABV-G mRNA vaccinated mice were protected against lethal intracerebral challenge infection. Inhibition of viral replication by vaccination was verified by qRT-PCR. Furthermore, we demonstrate that CD4+ T cells are crucial for the generation of neutralizing antibodies. In domestic pigs we were able to induce VN titers that correlate with protection in adult and newborn pigs. This study demonstrates the feasibility of a non-replicating mRNA rabies vaccine in small and large animals and highlights the promises of mRNA vaccines for the prevention of infectious diseases. PMID:27336830

  15. Molecular regulation of the hypothalamic-pituitary-adrenal axis in adult male guinea pigs after prenatal stress at different stages of gestation

    PubMed Central

    Kapoor, Amita; Leen, Jason; Matthews, Stephen G

    2008-01-01

    Studies in humans and animals have demonstrated that maternal stress during fetal development can lead to altered hypothalamic-pituitary-adrenal (HPA) axis function and behaviour postnatally. We have previously shown adult male guinea pigs that were born to mothers exposed to a stressor during the phase of rapid fetal brain growth (gestational days (GD) 50, 51 and 52; prenatal stress (PS)50) exhibit significantly increased basal plasma cortisol levels. In contrast, male guinea pig offspring whose mothers were exposed to stress later in gestation (GD60, 61 and 62; PS60) exhibited a significantly higher plasma cortisol response to activation of the HPA axis. In the present study, we hypothesized that the endocrine changes in HPA axis function observed in male guinea pig offspring would be reflected by altered molecular regulation of the HPA axis. Corticosteroid receptors in the hippocampus, hypothalamus and pituitary were measured, as well as corticotropin-releasing hormone (CRH), pro-opiomelanocortin (POMC) and adrenal enzymes in the paraventricular nucleus, pituitary and adrenal cortex, respectively, by in situ hybridization and Western blot. PS50 male offspring exhibited a significant reduction in glucocorticoid receptor (GR) mRNA (P <0.01) in the CA3 region of the hippocampus and significantly increased POMC mRNA (P <0.05) in the pituitary, consistent with the increase in basal HPA axis activity observed. In line with elevated activity of the HPA axis, both PS50 and PS60 male offspring exhibited significantly higher steroidogenic factor (SF)-1 (P <0.001) and melanocortin 2 receptor (MC2-R) mRNA (P <0.001) in the adrenal cortex. This study demonstrates that short periods of prenatal stress during critical windows of neuroendocrine development affect the expression of key regulators of HPA axis activity leading to the changes in endocrine function observed in prenatally stressed male offspring. Further, these changes are dependent on the timing of the maternal

  16. Evolution and genetic diversity of the Spain23F-ST81 clone causing adult invasive pneumococcal disease in Barcelona (1990–2012)

    PubMed Central

    Domenech, A.; Ardanuy, C.; Grau, I.; Calatayud, L.; Pallares, R.; Fenoll, A.; Brueggemann, A. B.; Liñares, J.

    2014-01-01

    Objectives We aimed to analyse the clinical epidemiology and genetic diversity of invasive pneumococcal disease (IPD) episodes attributed to the Spain23F-ST81 (PMEN1) clone. Methods Fifty-eight (2.7%) of 2117 invasive pneumococci isolated from adult patients during the 1990–2012 period shared a PFGE pattern related to the PMEN1 clone. The genotype was confirmed by multilocus sequence typing. The pbp2x, pbp1a, pbp2b and pspA genes were PCR-amplified and sequenced. Polymorphisms in the pspC gene were identified by PCR restriction fragment length polymorphism. The presence of transposons with erythromycin and tetracycline resistance determinants was detected by PCR. Results The prevalence of the PMEN1 clone increased from 0.8% in 1991 to 6.2% in 2001, and decreased to 0% in 2010–12, concomitant with the introduction of the seven-valent pneumococcal conjugate vaccine for children. A total of 93.1% of patients had pneumonia, meningitis or peritonitis; 87.9% of patients had associated underlying diseases, mainly cancer, chronic obstructive pulmonary disease and diabetes. Two closely related sequence types (STs) (ST81, n = 52; ST85, n = 6) were detected, with different serotypes: 23F (n = 42), 19A (n = 9) and 19F (n = 6). All the isolates were resistant to penicillin, co-trimoxazole and chloramphenicol. All the isolates also shared the same pbp1a allele, whereas multiple alleles of pbp2b, pbp2x, pspA and pspC were detected. Of the isolates, 89.7% were tetracycline resistant and 60.3% (n = 35) were macrolide resistant, and resistance was associated with different Tn916-like transposons. Conclusions Adult IPD caused by this clone was mainly detected in patients with underlying conditions, and genetic variability was observed among PMEN1 isolates collected in our area over the past 20 years. PMID:24324223

  17. Hair cell regeneration after ATOH1 gene therapy in the cochlea of profoundly deaf adult guinea pigs.

    PubMed

    Atkinson, Patrick J; Wise, Andrew K; Flynn, Brianna O; Nayagam, Bryony A; Richardson, Rachael T

    2014-01-01

    The degeneration of hair cells in the mammalian cochlea results in permanent sensorineural hearing loss. This study aimed to promote the regeneration of sensory hair cells in the mature cochlea and their reconnection with auditory neurons through the introduction of ATOH1, a transcription factor known to be necessary for hair cell development, and the introduction of neurotrophic factors. Adenoviral vectors containing ATOH1 alone, or with neurotrophin-3 and brain derived neurotrophic factor were injected into the lower basal scala media of guinea pig cochleae four days post ototoxic deafening. Guinea pigs treated with ATOH1 gene therapy, alone, had a significantly greater number of cells expressing hair cell markers compared to the contralateral non-treated cochlea when examined 3 weeks post-treatment. This increase, however, did not result in a commensurate improvement in hearing thresholds, nor was there an increase in synaptic ribbons, as measured by CtBP2 puncta after ATOH1 treatment alone, or when combined with neurotrophins. However, hair cell formation and synaptogenesis after co-treatment with ATOH1 and neurotrophic factors remain inconclusive as viral transduction was reduced due to the halving of viral titres when the samples were combined. Collectively, these data suggest that, whilst ATOH1 alone can drive non-sensory cells towards an immature sensory hair cell phenotype in the mature cochlea, this does not result in functional improvements after aminoglycoside-induced deafness.

  18. Hair Cell Regeneration after ATOH1 Gene Therapy in the Cochlea of Profoundly Deaf Adult Guinea Pigs

    PubMed Central

    Atkinson, Patrick J.; Wise, Andrew K.; Flynn, Brianna O.; Nayagam, Bryony A.; Richardson, Rachael T.

    2014-01-01

    The degeneration of hair cells in the mammalian cochlea results in permanent sensorineural hearing loss. This study aimed to promote the regeneration of sensory hair cells in the mature cochlea and their reconnection with auditory neurons through the introduction of ATOH1, a transcription factor known to be necessary for hair cell development, and the introduction of neurotrophic factors. Adenoviral vectors containing ATOH1 alone, or with neurotrophin-3 and brain derived neurotrophic factor were injected into the lower basal scala media of guinea pig cochleae four days post ototoxic deafening. Guinea pigs treated with ATOH1 gene therapy, alone, had a significantly greater number of cells expressing hair cell markers compared to the contralateral non-treated cochlea when examined 3 weeks post-treatment. This increase, however, did not result in a commensurate improvement in hearing thresholds, nor was there an increase in synaptic ribbons, as measured by CtBP2 puncta after ATOH1 treatment alone, or when combined with neurotrophins. However, hair cell formation and synaptogenesis after co-treatment with ATOH1 and neurotrophic factors remain inconclusive as viral transduction was reduced due to the halving of viral titres when the samples were combined. Collectively, these data suggest that, whilst ATOH1 alone can drive non-sensory cells towards an immature sensory hair cell phenotype in the mature cochlea, this does not result in functional improvements after aminoglycoside-induced deafness. PMID:25036727

  19. Factors influencing the commercialisation of cloning in the pork industry.

    PubMed

    Pratt, S L; Sherrer, E S; Reeves, D E; Stice, S L

    2006-01-01

    Production of cloned pigs using somatic cell nuclear transfer (SCNT) is a repeatable and predictable procedure and multiple labs around the world have generated cloned pigs and genetically modified cloned pigs. Due to the integrated nature of the pork production industry, pork producers are the most likely to benefit and are in the best position to introduce cloning in to production systems. Cloning can be used to amplify superior genetics or be used in conjunction with genetic modifications to produce animals with superior economic traits. Though unproven, cloning could add value by reducing pig-to-pig variability in economically significant traits such as growth rate, feed efficiency, and carcass characteristics. However, cloning efficiencies using SCNT are low, but predictable. The inefficiencies are due to the intrusive nature of the procedure, the quality of oocytes and/or the somatic cells used in the procedure, the quality of the nuclear transfer embryos transferred into recipients, pregnancy rates of the recipients, and neonatal survival of the clones. Furthermore, in commercial animal agriculture, clones produced must be able to grow and thrive under normal management conditions, which include attainment of puberty and subsequent capability to reproduce. To integrate SCNT into the pork industry, inefficiencies at each step of the procedure must be overcome. In addition, it is likely that non-surgical embryo transfer will be required to deliver cloned embryos, and/or additional methods to generate high health clones will need to be developed. This review will focus on the state-of-the-art for SCNT in pigs and the steps required for practical implementation of pig cloning in animal agriculture.

  20. [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.

  1. Identification of porcine polycystic kidney disease 1 (PKD1) gene: molecular cloning, expression profile, and implication in disease model.

    PubMed

    He, Jin; Wang, Qingsong; Ye, Jianhua; Hu, Xiaoxiang; Li, Ning

    2011-12-15

    The polycystic kidney disease 1 (PKD1) gene, which accounts for ~85% of human autosomal dominant polycystic kidney disease (ADPKD) cases, has been extensively studied in human and mouse. Much information about the pathogenesis of and treatments for ADPKD has been gained from the use of mouse models. However, because mouse models pose some limitations, further studies in other model systems are needed to investigate the biological basis of ADPKD. The pig is regarded as an important biomedical model. Thus, we isolated a pig PKD1 homolog and characterized its cDNA sequence, genomic structure, expression profile, alternative splicing, methylation status, protein characteristics, and immunohistochemical features in both neonatal and adult pigs. The pig PKD1 cDNA is 14,209bp long and encodes a 4305-residue polypeptide. The genomic sequence of PKD1 is ~50kb with 46 exons. An alternative splice acceptor site was identified in intron 9. PKD1 is expressed in all tissues tested in both neonatal and adult pigs and exhibits a developmentally regulated expression pattern. Western blotting revealed that the molecular mass of polycystin-1 is ~460kDa, but its expression level is relatively low. Immunohistochemical study of the kidneys shows that polycystin-1 is mainly expressed in the tubular epithelia. Bisulfite methylation analysis of CpG islands in the promoter region does not show a direct correlation between methylation status and expression level among different tissues/cells. The cloning and characterization of pig PKD1 indicates that the pig and human genes are highly similar in length of genomic and cDNA sequences, genomic structure and context, expression patterns, conserved transcription factor binding sites, and the molecular mass of the encoded polycystin-1. These data support our current understanding of PKD1, and suggest that the pig is an ideal candidate for development of an ADPKD disease model. PMID:21945688

  2. Cloning of Homo sapiens? No!

    PubMed

    McKinnell, Robert G

    2002-01-01

    Animal cloning by nuclear transplantation was first developed in the northern leopard frog, Rana pipiens. It was soon extended to other amphibian species and within time, to various mammalian species. The production of a cloned sheep (Dolly) from an adult nuclear donor reawakened interest in human cloning. Nuclear transfer for the production of animal clones has served experimental biology well. Nonetheless, the potential burden of developmental hazards, scientists and funds diverted from more needy causes, as well as the potential assault on the concept of family has led the author to oppose human cloning.

  3. Cloning of Homo sapiens? No!

    PubMed

    McKinnell, Robert G

    2002-01-01

    Animal cloning by nuclear transplantation was first developed in the northern leopard frog, Rana pipiens. It was soon extended to other amphibian species and within time, to various mammalian species. The production of a cloned sheep (Dolly) from an adult nuclear donor reawakened interest in human cloning. Nuclear transfer for the production of animal clones has served experimental biology well. Nonetheless, the potential burden of developmental hazards, scientists and funds diverted from more needy causes, as well as the potential assault on the concept of family has led the author to oppose human cloning. PMID:11841468

  4. Cloning to reproduce desired genotypes.

    PubMed

    Westhusin, M E; Long, C R; Shin, T; Hill, J R; Looney, C R; Pryor, J H; Piedrahita, J A

    2001-01-01

    Cloned sheep, cattle, goats, pigs and mice have now been produced using somatic cells for nuclear transplantation. Animal cloning is still very inefficient with on average less than 10% of the cloned embryos transferred resulting in a live offspring. However successful cloning of a variety of different species and by a number of different laboratory groups has generated tremendous interest in reproducing desired genotypes. Some of these specific genotypes represent animal cell lines that have been genetically modified. In other cases there is a significant demand for cloning animals characterized by their inherent genetic value, for example prize livestock, household pets and rare or endangered species. A number of different variables may influence the ability to reproduce a specific genotype by cloning. These include species, source of recipient ova, cell type of nuclei donor, treatment of donor cells prior to nuclear transfer, and the techniques employed for nuclear transfer. At present, there is no solid evidence that suggests cloning will be limited to only a few specific animals, and in fact, most data collected to date suggests cloning will be applicable to a wide variety of different animals. The ability to reproduce any desired genotype by cloning will ultimately depend on the amount of time and resources invested in research.

  5. Cloning cattle.

    PubMed

    Oback, B; Wells, D N

    2003-01-01

    Over the past six years, hundreds of apparently normal calves have been cloned worldwide from bovine somatic donor cells. However, these surviving animals represent less than 5% of all cloned embryos transferred into recipient cows. Most of the remaining 95% die at various stages of development from a predictable pattern of placental and fetal abnormalities, collectively referred to as the "cloning-syndrome." The low efficiency seriously limits commercial applicability and ethical acceptance of somatic cloning and enforces the development of improved cloning methods. In this paper, we describe our current standard operating procedure (SOP) for cattle cloning using zona-free nuclear transfer. Following this SOP, the output of viable and healthy calves at weaning is about 9% of embryos transferred. Better standardization of cloning protocols across and within research groups is needed to separate technical from biological factors underlying low cloning efficiency.

  6. Why Clone?

    MedlinePlus

    ... How might cloning be used in medicine? Cloning animal models of disease Much of what researchers learn about human disease comes from studying animal models such as mice. Often, animal models are ...

  7. Age differences in the purr call distinguished by units in the adult guinea pig primary auditory cortex.

    PubMed

    Grimsley, J M S; Palmer, A R; Wallace, M N

    2011-07-01

    Many communication calls contain information about the physical characteristics of the calling animal. During maturation of the guinea pig purr call the pitch becomes lower as the fundamental frequency progressively decreases from 476 to 261 Hz on average. Neurons in the primary auditory cortex (AI) often respond strongly to the purr and we postulated that some of them are capable of distinguishing between purr calls of different pitch. Consequently four pitch-shifted versions of a single call were used as stimuli. Many units in AI (79/182) responded to the purr call either with an onset response or with multiple bursts of firing that were time-locked to the phrases of the call. All had a characteristic frequency ≤5 kHz. Both types of unit altered their firing rate in response to pitch-shifted versions of the call. Of the responsive units, 41% (32/79) had a firing rate locked to the stimulus envelope that was at least 50% higher for one version of the call than any other. Some (14/32) had a preference that could be predicted from their frequency response area while others (18/32) were not predictable. We conclude that about 18% of stimulus-driven cells at the low-frequency end of AI are very sensitive to age-related changes in the purr call.

  8. [Cloning - controversies].

    PubMed

    Twardowski, T; Michalska, A

    2001-01-01

    Cloning of the human being is not only highly controversial; in the opinion of the authors it is impossible - we are not able to reproduce human behaviour and character traits. Reproduction through cloning is limited to personal genome resources. The more important is protection of genomic characteristics as private property and taking advantage of cloning for production of the human organs directly or through xenotransplants. In this paper we present the legislation related to cloning in Poland, in the European Union and other countries. We also indicate who and why is interested in cloning.

  9. Localisation of calcium ions and calcium-ATPase activity within myelinated nerve fibres of the adult guinea-pig optic nerve.

    PubMed Central

    Maxwell, W L; Watt, C; Pediani, J D; Graham, D I; Adams, J H; Gennarelli, T A

    1991-01-01

    There is no published description of the distribution of free Ca2+, nor of the distribution of Ca(2+)-ATPase activity associated with the maintenance of low axoplasmic Ca2+ concentrations, in normal central myelinated nerve fibres. We have used the oxalate-pyroantimonate technique to localise free Ca2+, together with the lead-citrate technique to localise Ca(2+)-ATPase activity within myelinated fibres from the adult guinea-pig optic nerve. Pyroantimonate precipitate occurred within the axoplasm at nodes of Ranvier and the internode, at areas of myelin disruption, within Schmidt-Lanterman incisures (SLI) and glial paranodal loops. But precipitate was absent from the axoplasm beneath SLI and at the paranode. Ca(2+)-ATPase activity was localised in axonal smooth endoplasmic reticulum (SER), the outer membrane of mitochondria, the nodal axolemma, the glial membranes of the paranodal loops, the SLI and the external aspect of the myelin sheath. We have demonstrated large domains within the axons of CNS fibres where calcium is present or absent. Moreover, we have shown that, where calcium is absent, there is localisation of Ca(2+)-ATPase activity, which would serve to remove calcium from the adjacent axoplasm. Our results are compared with information obtained from PNS fibres and some differences of distribution discussed. Images Fig. 1 Fig. 2 Fig. 3 Fig. 4 Fig. 5 Fig. 6 Fig. 7 Fig. 8 PMID:1833365

  10. Pattern of carbon dioxide production and retention is similar in adult pigs when fed hourly, but not when fed a single meal

    PubMed Central

    Moehn, Soenke; Bertolo, Robert FP; Pencharz, Paul B; Ball, Ronald O

    2004-01-01

    Background The understanding of bicarbonate kinetics and CO2 retention in the body is necessary to conduct amino acid tracer oxidation studies in both humans and laboratory animals. Significant metabolic activity is associated with eating which can affect bicarbonate steady state kinetics. A study was conducted to assess the impact of feeding regimen on the recovery of labelled bicarbonate and energy expenditure in adult female pigs (sows). Five catheterized sows (235 ± 5 kg) were fed semi-synthetic diets as: a single meal 2 h into the infusion after an overnight fast, or in eight hourly meals starting 2 h before the infusion. Oxygen consumption, CO2 production and 14CO2 recovery (ie fraction not retained) were determined during primed, constant intravenous infusions of NaH14CO3. Results The 14CO2 recovery (%) after fasting (58.1 ± 4.8) was lower than that after single meal feeding (78.8 ± 5.9) or hourly meal feeding (81.0 ± 2.6, P = 0.03). CO2 production correlated with 14CO2 recovery during hourly feeding (r = 0.40, P = 0.01); this relationship was not significant after single meal feeding (P = 0.30), probably due to physical activity-associated CO2 production. Conclusions The correlation of CO2 retention factors with CO2 production during hourly feeding suggests that this regimen should be preferred for future amino acid kinetics studies. PMID:15242516

  11. Hypervirulent Klebsiella pneumoniae clones causing bacteraemia in adults in a teaching hospital in Barcelona, Spain (2007-2013).

    PubMed

    Cubero, M; Grau, I; Tubau, F; Pallarés, R; Dominguez, M A; Liñares, J; Ardanuy, C

    2016-02-01

    Virulent hypermucoviscous Klebsiella pneumoniae strains associated with the magA and rmpA genes have mainly emerged in Asia. We analysed the frequency and the clinical and molecular epidemiology of K. pneumoniae bacteraemia isolates obtained over a 7-year period (2007-2013). Fifty-three of 878 K. pneumoniae invasive isolates (5.4%) showed a hypermucoviscous phenotype (by the string test). Of these, 16 (30.2%) were magA(+)/rmpA(+), 12 (22.6%) were magA(-)/rmpA(+), and the remaining 25 (47.2%) were magA(-)/rmpA(-). After multilocus sequence typing and wzi sequencing, all magA(+)/rmpA(+) isolates were serotype K1 and sequence type (ST)23. Of the 12 magA(-)/rmpA(+) isolates, nine were K2 (ST380, ST86, ST65, ST25 and ST493), and three magA(-)/rmpA(+) isolates had the new wzi allele 122, an unknown serotype, and the new ST1013. The remaining isolates, which were magA(-)/rmpA(-), showed different serotypes and STs. Patients with magA(+)/rmpA(+) or magA(-)/rmpA(+)K. pneumoniae bacteraemia more frequently had pyogenic liver abscesses (PLAs) and pneumonia than patients with magA(-)/rmpA(-)K. pneumoniae bacteraemia (respectively: 21.4% vs. 8%, p 0.26; and 17.9% vs. 0%, p 0.05). In fact, magA(-)/rmpA(-) isolates were similar to the those termed 'classic' K. pneumoniae isolates causing bacteraemia, the urinary and biliary tracts being the main foci of infection. In conclusion, hypervirulent clones (CC23K1, CC86K2, CC65K2, and CC380K2) were infrequent among K. pneumoniae isolates causing bacteraemia in our geographical area. A hypermucoviscous phenotype as determined with the string test is not enough to recognize these clones; additional molecular studies are needed. Patients with magA(+) and/or rmpA(+)K. pneumoniae bacteraemia more frequently had PLAs and pneumonia than patients without hypermucoviscosity genes.

  12. High-Throughput Cloning of Temperature-Sensitive Caenorhabditis elegans Mutants with Adult Syncytial Germline Membrane Architecture Defects

    PubMed Central

    Lowry, Josh; Yochem, John; Chuang, Chien-Hui; Sugioka, Kenji; Connolly, Amy A.; Bowerman, Bruce

    2015-01-01

    The adult Caenorhabditis elegans hermaphrodite gonad consists of two mirror-symmetric U-shaped arms, with germline nuclei located peripherally in the distal regions of each arm. The nuclei are housed within membrane cubicles that are open to the center, forming a syncytium with a shared cytoplasmic core called the rachis. As the distal germline nuclei progress through meiotic prophase, they move proximally and eventually cellularize as their compartments grow in size. The development and maintenance of this complex and dynamic germline membrane architecture are relatively unexplored, and we have used a forward genetic screen to identify 20 temperature-sensitive mutations in 19 essential genes that cause defects in the germline membrane architecture. Using a combined genome-wide SNP mapping and whole genome sequencing strategy, we have identified the causal mutations in 10 of these mutants. Four of the genes we have identified are conserved, with orthologs known to be involved in membrane biology, and are required for proper development or maintenance of the adult germline membrane architecture. This work provides a starting point for further investigation of the mechanisms that control the dynamics of syncytial membrane architecture during adult oogenesis. PMID:26311651

  13. In Vitro and In Vivo Development of Horse Cloned Embryos Generated with iPSCs, Mesenchymal Stromal Cells and Fetal or Adult Fibroblasts as Nuclear Donors

    PubMed Central

    Olivera, Ramiro; Moro, Lucia Natalia; Jordan, Roberto; Luzzani, Carlos; Miriuka, Santiago; Radrizzani, Martin; Donadeu, F. Xavier; Vichera, Gabriel

    2016-01-01

    The demand for equine cloning as a tool to preserve high genetic value is growing worldwide; however, nuclear transfer efficiency is still very low. To address this issue, we first evaluated the effects of time from cell fusion to activation (<1h, n = 1261; 1-2h, n = 1773; 2-3h, n = 1647) on in vitro and in vivo development of equine embryos generated by cloning. Then, we evaluated the effects of using different nuclear donor cell types in two successive experiments: I) induced pluripotent stem cells (iPSCs) vs. adult fibroblasts (AF) fused to ooplasts injected with the pluripotency-inducing genes OCT4, SOX2, MYC and KLF4, vs. AF alone as controls; II) umbilical cord-derived mesenchymal stromal cells (UC-MSCs) vs. fetal fibroblasts derived from an unborn cloned foetus (FF) vs. AF from the original individual. In the first experiment, both blastocyst production and pregnancy rates were higher in the 2-3h group (11.5% and 9.5%, respectively), respect to <1h (5.2% and 2%, respectively) and 1-2h (5.6% and 4.7%, respectively) groups (P<0.05). However, percentages of born foals/pregnancies were similar when intervals of 2-3h (35.2%) or 1-2h (35.7%) were used. In contrast to AF, the iPSCs did not generate any blastocyst-stage embryos. Moreover, injection of oocytes with the pluripotency-inducing genes did not improve blastocyst production nor pregnancy rates respect to AF controls. Finally, higher blastocyst production was obtained using UC-MSC (15.6%) than using FF (8.9%) or AF (9.3%), (P<0.05). Despite pregnancy rates were similar for these 3 groups (17.6%, 18.2% and 22%, respectively), viable foals (two) were obtained only by using FF. In summary, optimum blastocyst production rates can be obtained using a 2-3h interval between cell fusion and activation as well as using UC-MSCs as nuclear donors. Moreover, FF line can improve the efficiency of an inefficient AF line. Overall, 24 healthy foals were obtained from a total of 29 born foals. PMID:27732616

  14. To clone or not to clone--whither the law?

    PubMed

    Lupton, M L

    1999-01-01

    The cloning of Dolly the lamb from adult cells by scientists at the Roslin Laboratories near Edinburgh in February 1997 has startled the world because it now opens the way to clone adult human beings. The reaction to Ian Wilmut's breakthrough has been instant and largely negative. Bills were rushed into both the US Senate and House of Representatives aimed at banning the cloning of human beings. Human cloning is premature at this stage, but there are many positive spin-offs of cloning in the field of genetic engineering, such as the production of human proteins such as blood clotting factors which aid in healing wounds. Progress by means of cloning can also be made into devising a cure for Parkinson's Disease amongst others. No lesser ethicist than John C. Fletcher of the University of Virginia foresees circumstances in which human cloning is acceptable e.g. to enable a couple to replace a dying child, to enable a couple, one of whom is infertile, to clone a child from either partner. Extensive regulation of cloning by the law is inevitable but, in doing so, the legislation should be careful not to outlaw research in this area which could be beneficial to mankind. PMID:10436743

  15. To clone or not to clone--whither the law?

    PubMed

    Lupton, M L

    1999-01-01

    The cloning of Dolly the lamb from adult cells by scientists at the Roslin Laboratories near Edinburgh in February 1997 has startled the world because it now opens the way to clone adult human beings. The reaction to Ian Wilmut's breakthrough has been instant and largely negative. Bills were rushed into both the US Senate and House of Representatives aimed at banning the cloning of human beings. Human cloning is premature at this stage, but there are many positive spin-offs of cloning in the field of genetic engineering, such as the production of human proteins such as blood clotting factors which aid in healing wounds. Progress by means of cloning can also be made into devising a cure for Parkinson's Disease amongst others. No lesser ethicist than John C. Fletcher of the University of Virginia foresees circumstances in which human cloning is acceptable e.g. to enable a couple to replace a dying child, to enable a couple, one of whom is infertile, to clone a child from either partner. Extensive regulation of cloning by the law is inevitable but, in doing so, the legislation should be careful not to outlaw research in this area which could be beneficial to mankind.

  16. A comparative study on efficiency of adult fibroblasts and amniotic fluid-derived stem cells as donor cells for production of hand-made cloned buffalo (Bubalus bubalis) embryos.

    PubMed

    Em, Sadeesh; Kataria, Meena; Shah, Fozia; Yadav, P S

    2016-08-01

    The efficiency of two cell types, namely adult fibroblasts, and amniotic fluid stem (AFS) cells as nuclear donor cells for somatic cell nuclear transfer by hand-made cloning in buffalo (Bubalus bubalis) was compared. The in vitro expanded buffalo adult fibroblast cells showed a typical "S" shape growth curve with a doubling time of 40.8 h and stained positive for vimentin. The in vitro cultured undifferentiated AFS cells showed a doubling time of 33.2 h and stained positive for alkaline phosphatase, these cells were also found positive for undifferentiated embryonic stem cell markers like OCT-4, NANOG and SOX-2, which accentuate their pluripotent property. Further, when AFS cells were exposed to corresponding induction conditions, these cells differentiated into osteogenic, adipogenic and chondrogenic lineages which was confirmed through alizaran, oil red O and alcian blue staining, respectively. Cultured adult fibroblasts and AFS cells of passages 10-15 and 8-12, respectively, were used as nuclear donors. A total of 94 embryos were reconstructed using adult fibroblast as donor cells with cleavage and blastocyst production rate of 62.8 ± 1.8 and 19.1 ± 1.5, respectively. An overall cleavage and blastocyst formation rate of 71.1 ± 1.2 and 29.9 ± 2.2 was obtained when 97 embryos were reconstructed using AFS cells as donor cells. There were no significant differences (P > 0.05) in reconstructed efficiency between the cloned embryos derived from two donor cells, whereas the results showed that there were significant differences (P < 0.05) in cleavage and blastocyst rates between the cloned embryos derived from two donor cell groups. Average total cell numbers for blastocyst generated using AFS cells (172.4 ± 5.8) was significantly (P < 0.05) higher than from adult fibroblasts (148.2 ± 6.1). This study suggests that the in vitro developmental potential of the cloned embryos derived from AFS cells were higher than that of the cloned embryos

  17. Detection of an early adult T-cell leukemia-lymphoma clone in lymph nodes with anaplastic lymphoma kinase-negative anaplastic large cell lymphoma involvement.

    PubMed

    Tokunaga, Masahito; Yoshida, Noriaki; Nakano, Nobuaki; Kubota, Ayumu; Takeuchi, Shogo; Takatsuka, Yoshifusa; Seto, Masao; Utsunomiya, Atae

    2016-04-01

    A 58-year-old man was admitted to our hospital with systemic lymphadenopathy and was diagnosed with anaplastic lymphoma kinase-negative anaplastic large cell lymphoma (ALCL) by lymph node biopsy. Although he was a human T-cell leukemia virus type I (HTLV-1) carrier, Southern blot analysis of the lymph node did not show monoclonal integration of HTLV-1 provirus deoxyribonucleic acid (DNA). He achieved complete remission after chemotherapy and subsequently, autologous peripheral blood stem cell transplantation (auto-PBSCT) was performed. Fifteen months after the auto-PBSCT, abnormal lymphocytes in the peripheral blood gradually increased. Southern blot analysis revealed monoclonal integration of HTLV-1 provirus DNA and monoclonal rearrangement of TRB. He was diagnosed with chronic type adult T-cell leukemia-lymphoma (ATL), which immediately progressed to the acute type. He died of tumor progression despite intensive chemotherapy. We analyzed genomic alterations of the ALCL and ATL cells using array comparative genomic hybridization. We found that the genomic alteration pattern differed between the two diseases. T-cell receptor clonality analysis using polymerase chain reaction (PCR) showed that the T-cell clone of the ATL was present in the lymph nodes with ALCL involvement, but not in peripheral blood. This finding suggests that lymph nodes can serve as a niche for ATL development.

  18. Science and technology of farm animal cloning: state of the art.

    PubMed

    Vajta, Gábor; Gjerris, Mickey

    2006-05-01

    Details of the first mammal born after nuclear transfer cloning were published by Steen Malte Willadsen in 1986. In spite of its enormous scientific significance, this discovery failed to trigger much public concern, possibly because the donor cells were derived from pre-implantation stage embryos. The major breakthrough in terms of public recognition has happened when Ian Wilmut et al. [Wilmut, I., Schnieke, A.E., McWhir, J., Kind, A.J., Campbell, K.H., 1997. Viable offspring derived from fetal és adult mammalian cells. Nature 385, 810-813] described the successful application of almost exactly the same method, but using the nuclei of somatic cells from an adult mammal, to create Dolly the sheep. It has become theoretically possible to produce an unlimited number of genetic replicates from an adult animal or a post-implantation foetus. Since 1997 a number of different species including pigs, goats, horses, cats, etc. have been cloned with the somatic cell nuclear transfer technique. Although the technology still has relatively low success rates and there seems to be substantial problems with the welfare of some of the cloned animals, cloning is used both within basic research and the biomedical sector. The next step seems to be to implement cloning in the agricultural production system and several animals have been developed in this direction. This article reviews the current state of the art of farm animal cloning from a scientific and technological perspective, describes the animal welfare problems and critically assess different applications of farm animal cloning. The scope is confined to animal biotechnologies in which the use of cell nuclear transfer is an essential part and extends to both biomedical and agricultural applications of farm animal cloning. These applications include the production of genetically identical animals for research purposes, and also the creation of genetically modified animals. In the agricultural sector, cloning can be used as a

  19. Science and technology of farm animal cloning: state of the art.

    PubMed

    Vajta, Gábor; Gjerris, Mickey

    2006-05-01

    Details of the first mammal born after nuclear transfer cloning were published by Steen Malte Willadsen in 1986. In spite of its enormous scientific significance, this discovery failed to trigger much public concern, possibly because the donor cells were derived from pre-implantation stage embryos. The major breakthrough in terms of public recognition has happened when Ian Wilmut et al. [Wilmut, I., Schnieke, A.E., McWhir, J., Kind, A.J., Campbell, K.H., 1997. Viable offspring derived from fetal és adult mammalian cells. Nature 385, 810-813] described the successful application of almost exactly the same method, but using the nuclei of somatic cells from an adult mammal, to create Dolly the sheep. It has become theoretically possible to produce an unlimited number of genetic replicates from an adult animal or a post-implantation foetus. Since 1997 a number of different species including pigs, goats, horses, cats, etc. have been cloned with the somatic cell nuclear transfer technique. Although the technology still has relatively low success rates and there seems to be substantial problems with the welfare of some of the cloned animals, cloning is used both within basic research and the biomedical sector. The next step seems to be to implement cloning in the agricultural production system and several animals have been developed in this direction. This article reviews the current state of the art of farm animal cloning from a scientific and technological perspective, describes the animal welfare problems and critically assess different applications of farm animal cloning. The scope is confined to animal biotechnologies in which the use of cell nuclear transfer is an essential part and extends to both biomedical and agricultural applications of farm animal cloning. These applications include the production of genetically identical animals for research purposes, and also the creation of genetically modified animals. In the agricultural sector, cloning can be used as a

  20. Academic Cloning.

    ERIC Educational Resources Information Center

    Sikula, John P.; Sikula, Andrew F.

    1980-01-01

    The authors define "cloning" as an integral feature of all educational systems, citing teaching practices which reward students for closely reproducing the teacher's thoughts and/or behaviors and administrative systems which tend to promote like-minded subordinates. They insist, however, that "academic cloning" is not a totally negative practice.…

  1. A high utility integrated map of the pig genome

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Background: The domestic pig is being increasingly exploited as a system for modeling human disease. It also has substantial economic importance for meat-based protein production. Physical clone maps have underpinned large-scale genomic sequencing and enabled focused cloning efforts for many genome...

  2. Unique circovirus-like genome detected in pig feces.

    PubMed

    Cheung, Andrew K; Ng, Terry Fei-Fan; Lager, Kelly M; Alt, David P; Delwart, Eric L; Pogranichniy, Roman M

    2014-01-01

    Using a metagenomic approach and molecular cloning methods, we identified, cloned, and sequenced the complete genome of a novel circular DNA virus, porcine stool-associated virus (PoSCV4), from pig feces. Phylogenetic analysis of the deduced replication initiator protein showed that PoSCV4 is most related to a fur seal feces-associated circular DNA virus. PMID:24723710

  3. Cloning the laboratory mouse.

    PubMed

    Wakayama, T; Yanagimachi, R

    1999-06-01

    A brief account is given of early attempts to clone mammals (mice) by transferring cells (nuclei) of preimplantation embryos into enucleated oocytes, zygotes or blastomeres of two-cell embryos. This is followed by a brief review of recent successes using adult somatic cells: mammary gland cells for sheep, muscle cells for cattle and cumulus cells for mice. We have developed a technique for cloning the laboratory mouse by transferring cumulus cell nuclei into enucleated oocytes. With this technique, we have produced a population of over 80 cloned animals, and have carried the process over four generations. Development and fertility of these appear normal. However, the yield is very low; only approximately 1% of injected oocytes are carried to term. The challenge is now to understand the reason for this high loss. Is it a problem of technique, genomic reprogramming, somatic mutation, imprinting or incompatible cell cycle phases?

  4. Malignant transformation of guinea pig cells after exposure to ultraviolet-irradiated guinea pig cytomegalovirus

    SciTech Connect

    Isom, H.C.; Mummaw, J.; Kreider, J.W.

    1983-04-30

    Guinea pig cells were malignantly transformed in vitro by ultraviolet (uv)-irradiated guinea pig cytomegalovirus (GPCMV). When guinea pig hepatocyte monolayers were infected with uv-irradiated GPCMV, three continuous epithelioid cell lines which grew in soft agarose were established. Two independently derived GPCMV-transformed liver cells and a cell line derived from a soft agarose clone of one of these lines induced invasive tumors when inoculated subcutaneously or intraperitoneally into nude mice. The tumors were sarcomas possibly derived from hepatic stroma or sinusoid. Transformed cell lines were also established after infection of guinea pig hepatocyte monolayers with human cytomegalovirus (HCMV) or simian virus 40 (SV40). These cell lines also formed colonies in soft agarose and induced sarcomas in nude mice. It is concluded that (i) GPCMV can malignantly transform guinea pig cells; (ii) cloning of GPCMV-transformed cells in soft agarose produced cells that induced tumors with a shorter latency period but with no alteration in growth rate or final tumor size; and (iii) the tumors produced by GPCMV-and HCMV-transformed guinea pig cells were more similar to each other in growth rate than to those induced by SV40-transformed guinea pig cells.

  5. Cloning: questions answered and unsolved.

    PubMed

    Latham, Keith E

    2004-02-01

    Cloning by the transfer of adult somatic cell nuclei to oocytes has produced viable offspring in a variety of mammalian species. The technology is still in its initial stages of development. Studies to date have answered several basic questions related to such issues as genome potency, life expectancy of clones, mitochondrial fates, and feasibility of inter-species nuclear transfer. They have also raised new questions related to the control of nuclear reprogramming and function. These questions are reviewed here.

  6. Comparison of in vitro developmental competence of cloned caprine embryos using donor karyoplasts from adult bone marrow mesenchymal stem cells vs ear fibroblast cells.

    PubMed

    Kwong, P J; Nam, H Y; Wan Khadijah, W E; Kamarul, T; Abdullah, R B

    2014-04-01

    The aim of this study was to produce cloned caprine embryos using either caprine bone marrow-derived mesenchymal stem cells (MSCs) or ear fibroblast cells (EFCs) as donor karyoplasts. Caprine MSCs were isolated from male Boer goats of an average age of 1.5 years. To determine the pluripotency of MSCs, the cells were induced to differentiate into osteocytes, chondrocytes and adipocytes. Subsequently, MSCs were characterized through cell surface antigen profiles using specific markers, prior to their use as donor karyoplasts for nuclear transfer. No significant difference (p > 0.05) in fusion rates was observed between MSCs (87.7%) and EFCs (91.3%) used as donor karyoplasts. The cleavage rate of cloned embryos derived with MSCs (87.0%) was similar (p > 0.05) to those cloned using EFCs (84.4%). However, the in vitro development of MSCs-derived cloned embryos (25.3%) to the blastocyst stage was significantly higher (p < 0.05) than those derived with EFCs (20.6%). In conclusion, MSCs could be reprogrammed by caprine oocytes, and production of cloned caprine embryos with MSCs improved their in vitro developmental competence, but not in their fusion and cleavage rate as compared to cloning using somatic cells such as EFCs. PMID:24456113

  7. The cloning and sequence analysis of the aspC and tyrB genes from Escherichia coli K12. Comparison of the primary structures of the aspartate aminotransferase and aromatic aminotransferase of E. coli with those of the pig aspartate aminotransferase isoenzymes.

    PubMed Central

    Fotheringham, I G; Dacey, S A; Taylor, P P; Smith, T J; Hunter, M G; Finlay, M E; Primrose, S B; Parker, D M; Edwards, R M

    1986-01-01

    In this paper we describe the cloning and sequence analysis of the tyrB and aspC genes from Escherichia coli K12, which encode the aromatic aminotransferase and aspartate aminotransferase respectively. The tyrB gene was isolated from a cosmid carrying the nearby dnaB gene, identified by its ability to complement a dnaB lesion. Deletion and linker insertion analysis located the tyrB gene to a 1.7-kilobase NruI-HindIII-digest fragment. Sequence analysis revealed a gene encoding a 43 000 Da polypeptide. The gene starts with a GTG codon and is closely followed by a structure resembling a rho independent terminator. The aspC gene was cloned by screening gene banks, prepared from a prototrophic E. coli K12 strain, for plasmids able to complement the aspC tyrB lesions in the aminotransferase-deficient strain HW225. Sub-cloning and deletion analysis located the aspC gene on a 1.8-kilobase HincII-StuI-digest fragment. Sequence analysis revealed the presence of a gene encoding a 43 000 Da protein, the sequence of which is identical with that previously obtained for the aspartate aminotransferase from E. coli B. Considerable overproduction of the two enzymes was demonstrated. We compared the deduced protein sequences with those of the pig mitochondrial and cytoplasmic aspartate aminotransferases. From the extensive homology observed we are able to propose that the two E. coli enzymes possess subunit structures, subunit interactions and coenzyme-binding and substrate-binding sites that are very similar both to each other and to those of the mammalian enzymes and therefore must also have very similar catalytic mechanisms. Comparison of the aspC and tyrB gene sequences reveals that they appear to have diverged as much as is possible within the constraints of functionality and codon usage. PMID:3521591

  8. Genetic analysis of Trichuris suis and Trichuris trichiura recovered from humans and pigs in a sympatric setting in Uganda.

    PubMed

    Nissen, Sofie; Al-Jubury, Azmi; Hansen, Tina V A; Olsen, Annette; Christensen, Henrik; Thamsborg, Stig M; Nejsum, Peter

    2012-08-13

    The whipworms Trichuris trichiura and Trichuris suis in humans and pigs, respectively, are believed to be two different species yet closely related. Morphologically, adult worms, eggs and larvae of the two species are indistinguishable. The aim of this study was to examine the genetic variation of Trichuris sp. mainly recovered from natural infected pigs and humans. Worm material isolated from humans and pigs living in the same geographical region in Uganda were analyzed by PCR, cloning and sequencing. Measurements of morphometric characters were also performed. The analysis of the ITS-2 (internal transcribed spacer) region showed a high genetic variation in the human-derived worms with two sequence types, designated type 1 and type 2, differing with up to 45%, the type 2 being identical to the sequence found in pig-derived worms. A single human-derived worm showed exclusively the type 2-genotype (T. suis-type) and three cases of 'heterozygote' worms in humans were identified. However, the analysis showed that sympatric Trichuris primarily assorted with host origin. Sequence analysis of a part of the genetically conserved β-tubulin gene confirmed two separate populations/species but also showed that the 'heterozygote' worms had a T. suis-like β-tubulin gene. A PCR-RFLP on the ITS-2 region was developed, that could distinguish between worms of the pig, human and 'heterozygote' type. The data suggest that Trichuris in pigs and humans belong to two different populations (i.e. are two different species). However, the data presented also suggest that cross-infections of humans with T. suis takes place. Further studies on sympatric Trichuris populations are highly warranted in order to explore transmission dynamics and unravel the zoonotic potential of T. suis.

  9. Distribution patterns of the glucose transporters GLUT4 and GLUT1 in skeletal muscles of rats (Rattus norvegicus), pigs (Sus scrofa), cows (Bos taurus), adult goats, goat kids (Capra hircus), and camels (Camelus dromedarius).

    PubMed

    Duehlmeier, R; Sammet, K; Widdel, A; von Engelhardt, W; Wernery, U; Kinne, J; Sallmann, H-P

    2007-02-01

    Earlier studies demonstrated that forestomach herbivores are less insulin sensitive than monogastric omnivores. The present study was carried out to determine if different distribution patterns of the glucose transporters GLUT1 and GLUT4 may contribute to these different insulin sensitivities. Western blotting was used to measure GLUT1 and GLUT4 protein contents in oxidative (masseter, diaphragm) and glycolytic (longissimus lumborum, semitendinosus) skeletal muscle membranes of monogastric omnivores (rats and pigs), and of forestomach herbivores (cows, adult goats, goat kids, and camels). Muscles were characterized biochemically. Comparing red and white muscles, the isocitrate dehydrogenase (ICDH) activity was 1.5-15-times higher in oxidative muscles of all species, whereas lactate dehydrogenase (LDH) activity was 1.4-4.4-times higher in glycolytic muscles except in adult goats. GLUT4 levels were 1.5-6.3-times higher in oxidative muscles. GLUT1 levels were 2.2-8.3-times higher in glycolytic muscles in forestomach herbivores but not in monogastric animals. We conclude that GLUT1 may be the predominant glucose transporter in glycolytic muscles of ruminating animals. The GLUT1 distribution patterns were identical in adult and pre-ruminant goats, indicating that GLUT1 expression among these muscles is determined genetically. The high blood glucose levels of camels cited in literature may be due to an "NIDDM-like" impaired GLUT4 activity in skeletal muscle.

  10. Normal telomere lengths found in cloned cattle.

    PubMed

    Tian, X C; Xu, J; Yang, X

    2000-11-01

    Success of cloning using adult somatic cells has been reported in sheep, mice and cattle. The report that 'Dolly' the sheep, the first clone from an adult mammal, inherited shortened telomeres from her cell donor and that her telomeres were further shortened by the brief culture of donor cells has raised serious scientific and public concerns about the 'genetic age' and potential developmental problems of cloned animals. This observation was challenged by a recent report that showed calves cloned from fetal cells have longer telomeres than their age-matched controls. The question remains whether Dolly's short telomeres were an exception or a general fact, which would differ from the telomeres of fetal-derived clones. PMID:11062462

  11. Oxfendazole flukicidal activity in pigs.

    PubMed

    Ortiz, Pedro; Terrones, Susana; Cabrera, María; Hoban, Cristian; Ceballos, Laura; Moreno, Laura; Canton, Candela; Donadeu, Meritxell; Lanusse, Carlos; Alvarez, Luis

    2014-08-01

    Although oxfendazole (OFZ) is a well know broad-spectrum benzimidazole anthelmintic, the assessment of its potential trematodicidal activity remains unexplored. OFZ administration at single high doses has been recommended to control Taenia solium cysticercus in pigs. The current study investigated the flukicidal activity obtained after a single high (30mg/kg) oral dose of OFZ in pigs harbouring a natural Fasciola hepatica infection. Sixteen (16) local ecotype pigs were randomly allocated into two (2) experimental groups of 8 animals each named as follow: Untreated control and OFZ treated, in which animals received OFZ (Synanthic(®), Merial Ltd., 9.06% suspension) orally at 30mg/kg. At seven (7) days post-treatment, all the animals were sacrificed and direct adult liver fluke counts were performed following the WAAVP guidelines. None of the animals involved in this experiment showed any adverse event during the study. OFZ treatment as a single 30mg/kg oral dose showed a 100% efficacy against F. hepatica. In conclusion, the trial described here demonstrated an excellent OFZ activity against F. hepatica in naturally infected pigs, after its administration at a single oral dose of 30mg/kg.

  12. The Pig Olfactory Brain: A Primer

    PubMed Central

    Feldman, Sanford; Osterberg, Stephen K.

    2016-01-01

    Despite the fact that pigs are reputed to have excellent olfactory abilities, few studies have examined regions of the pig brain involved in the sense of smell. The present study provides an overview of the olfactory bulb, anterior olfactory nucleus, and piriform cortex of adult pigs using several approaches. Nissl, myelin, and Golgi stains were used to produce a general overview of the organization of the regions and confocal microscopy was employed to examine 1) projection neurons, 2) GABAergic local circuit neurons that express somatostatin, parvalbumin, vasoactive intestinal polypeptide, or calretinin, 3) neuromodulatory fibers (cholinergic and serotonergic), and 4) glia (astrocytes and microglia). The findings revealed that pig olfactory structures are quite large, highly organized and follow the general patterns observed in mammals. PMID:26936231

  13. The Pig Olfactory Brain: A Primer.

    PubMed

    Brunjes, Peter C; Feldman, Sanford; Osterberg, Stephen K

    2016-06-01

    Despite the fact that pigs are reputed to have excellent olfactory abilities, few studies have examined regions of the pig brain involved in the sense of smell. The present study provides an overview of the olfactory bulb, anterior olfactory nucleus, and piriform cortex of adult pigs using several approaches. Nissl, myelin, and Golgi stains were used to produce a general overview of the organization of the regions and confocal microscopy was employed to examine 1) projection neurons, 2) GABAergic local circuit neurons that express somatostatin, parvalbumin, vasoactive intestinal polypeptide, or calretinin, 3) neuromodulatory fibers (cholinergic and serotonergic), and 4) glia (astrocytes and microglia). The findings revealed that pig olfactory structures are quite large, highly organized and follow the general patterns observed in mammals. PMID:26936231

  14. Cloning cattle: the methods in the madness.

    PubMed

    Oback, Björn; Wells, David N

    2007-01-01

    Somatic cell nuclear transfer (SCNT) is much more widely and efficiently practiced in cattle than in any other species, making this arguably the most important mammal cloned to date. While the initial objective behind cattle cloning was commercially driven--in particular to multiply genetically superior animals with desired phenotypic traits and to produce genetically modified animals-researchers have now started to use bovine SCNT as a tool to address diverse questions in developmental and cell biology. In this paper, we review current cattle cloning methodologies and their potential technical or biological pitfalls at any step of the procedure. In doing so, we focus on one methodological parameter, namely donor cell selection. We emphasize the impact of epigenetic and genetic differences between embryonic, germ, and somatic donor cell types on cloning efficiency. Lastly, we discuss adult phenotypes and fitness of cloned cattle and their offspring and illustrate some of the more imminent commercial cattle cloning applications.

  15. The pig X and Y Chromosomes: structure, sequence, and evolution.

    PubMed

    Skinner, Benjamin M; Sargent, Carole A; Churcher, Carol; Hunt, Toby; Herrero, Javier; Loveland, Jane E; Dunn, Matt; Louzada, Sandra; Fu, Beiyuan; Chow, William; Gilbert, James; Austin-Guest, Siobhan; Beal, Kathryn; Carvalho-Silva, Denise; Cheng, William; Gordon, Daria; Grafham, Darren; Hardy, Matt; Harley, Jo; Hauser, Heidi; Howden, Philip; Howe, Kerstin; Lachani, Kim; Ellis, Peter J I; Kelly, Daniel; Kerry, Giselle; Kerwin, James; Ng, Bee Ling; Threadgold, Glen; Wileman, Thomas; Wood, Jonathan M D; Yang, Fengtang; Harrow, Jen; Affara, Nabeel A; Tyler-Smith, Chris

    2016-01-01

    We have generated an improved assembly and gene annotation of the pig X Chromosome, and a first draft assembly of the pig Y Chromosome, by sequencing BAC and fosmid clones from Duroc animals and incorporating information from optical mapping and fiber-FISH. The X Chromosome carries 1033 annotated genes, 690 of which are protein coding. Gene order closely matches that found in primates (including humans) and carnivores (including cats and dogs), which is inferred to be ancestral. Nevertheless, several protein-coding genes present on the human X Chromosome were absent from the pig, and 38 pig-specific X-chromosomal genes were annotated, 22 of which were olfactory receptors. The pig Y-specific Chromosome sequence generated here comprises 30 megabases (Mb). A 15-Mb subset of this sequence was assembled, revealing two clusters of male-specific low copy number genes, separated by an ampliconic region including the HSFY gene family, which together make up most of the short arm. Both clusters contain palindromes with high sequence identity, presumably maintained by gene conversion. Many of the ancestral X-related genes previously reported in at least one mammalian Y Chromosome are represented either as active genes or partial sequences. This sequencing project has allowed us to identify genes--both single copy and amplified--on the pig Y Chromosome, to compare the pig X and Y Chromosomes for homologous sequences, and thereby to reveal mechanisms underlying pig X and Y Chromosome evolution.

  16. The pig X and Y Chromosomes: structure, sequence, and evolution

    PubMed Central

    Skinner, Benjamin M.; Sargent, Carole A.; Churcher, Carol; Hunt, Toby; Herrero, Javier; Loveland, Jane E.; Dunn, Matt; Louzada, Sandra; Fu, Beiyuan; Chow, William; Gilbert, James; Austin-Guest, Siobhan; Beal, Kathryn; Carvalho-Silva, Denise; Cheng, William; Gordon, Daria; Grafham, Darren; Hardy, Matt; Harley, Jo; Hauser, Heidi; Howden, Philip; Howe, Kerstin; Lachani, Kim; Ellis, Peter J.I.; Kelly, Daniel; Kerry, Giselle; Kerwin, James; Ng, Bee Ling; Threadgold, Glen; Wileman, Thomas; Wood, Jonathan M.D.; Yang, Fengtang; Harrow, Jen; Affara, Nabeel A.; Tyler-Smith, Chris

    2016-01-01

    We have generated an improved assembly and gene annotation of the pig X Chromosome, and a first draft assembly of the pig Y Chromosome, by sequencing BAC and fosmid clones from Duroc animals and incorporating information from optical mapping and fiber-FISH. The X Chromosome carries 1033 annotated genes, 690 of which are protein coding. Gene order closely matches that found in primates (including humans) and carnivores (including cats and dogs), which is inferred to be ancestral. Nevertheless, several protein-coding genes present on the human X Chromosome were absent from the pig, and 38 pig-specific X-chromosomal genes were annotated, 22 of which were olfactory receptors. The pig Y-specific Chromosome sequence generated here comprises 30 megabases (Mb). A 15-Mb subset of this sequence was assembled, revealing two clusters of male-specific low copy number genes, separated by an ampliconic region including the HSFY gene family, which together make up most of the short arm. Both clusters contain palindromes with high sequence identity, presumably maintained by gene conversion. Many of the ancestral X-related genes previously reported in at least one mammalian Y Chromosome are represented either as active genes or partial sequences. This sequencing project has allowed us to identify genes—both single copy and amplified—on the pig Y Chromosome, to compare the pig X and Y Chromosomes for homologous sequences, and thereby to reveal mechanisms underlying pig X and Y Chromosome evolution. PMID:26560630

  17. An emerging zoonotic clone in the Netherlands provides clues to virulence and zoonotic potential of Streptococcus suis

    PubMed Central

    Willemse, N.; Howell, K. J.; Weinert, L. A.; Heuvelink, A.; Pannekoek, Y.; Wagenaar, J. A.; Smith, H. E.; van der Ende, A.; Schultsz, C.

    2016-01-01

    Streptococcus suis is a zoonotic swine pathogen and a major public health concern in Asia, where it emerged as an important cause of bacterial meningitis in adults. While associated with food-borne transmission in Asia, zoonotic S. suis infections are mainly occupational hazards elsewhere. To identify genomic differences that can explain zoonotic potential, we compared whole genomes of 98 S. suis isolates from human patients and pigs with invasive disease in the Netherlands, and validated our observations with 18 complete and publicly available sequences. Zoonotic isolates have smaller genomes than non-zoonotic isolates, but contain more virulence factors. We identified a zoonotic S. suis clone that diverged from a non-zoonotic clone by means of gene loss, a capsule switch, and acquisition of a two-component signalling system in the late 19th century, when foreign pig breeds were introduced. Our results indicate that zoonotic potential of S. suis results from gene loss, recombination and horizontal gene transfer events. PMID:27381348

  18. 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

  19. 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.

  20. Transgenesis for pig models

    PubMed Central

    Yum, Soo-Young; Yoon, Ki-Young; Lee, Choong-Il; Lee, Byeong-Chun

    2016-01-01

    Animal models, particularly pigs, have come to play an important role in translational biomedical research. There have been many pig models with genetically modifications via somatic cell nuclear transfer (SCNT). However, because most transgenic pigs have been produced by random integration to date, the necessity for more exact gene-mutated models using recombinase based conditional gene expression like mice has been raised. Currently, advanced genome-editing technologies enable us to generate specific gene-deleted and -inserted pig models. In the future, the development of pig models with gene editing technologies could be a valuable resource for biomedical research. PMID:27030199

  1. On classical cloning and no-cloning

    NASA Astrophysics Data System (ADS)

    Teh, Nicholas J.

    2012-02-01

    It is part of information theory folklore that, while quantum theory prohibits the generic (or universal) cloning of states, such cloning is allowed by classical information theory. Indeed, many take the phenomenon of no-cloning to be one of the features that distinguishes quantum mechanics from classical mechanics. In this paper, we argue that pace conventional wisdom, in the case where one does not include a machine system, there is an analog of the no-cloning theorem for classical systems. However, upon adjoining a non-trivial machine system (or ancilla) one finds that, pace the quantum case, the obstruction to cloning disappears for pure states. We begin by discussing some conceptual points and category-theoretic generalities having to do with cloning, and proceed to discuss no-cloning in both the case of (non-statistical) classical mechanics and classical statistical mechanics.

  2. Keith's MAGIC: Cloning and the Cell Cycle.

    PubMed

    Wells, D N

    2013-10-01

    Abstract Professor Keith Campbell's critical contribution to the discovery that a somatic cell from an adult animal can be fully reprogrammed by oocyte factors to form a cloned individual following nuclear transfer (NT)(Wilmut et al., 1997 ) overturned a dogma concerning the reversibility of cell fate that many scientists had considered to be biologically impossible. This seminal experiment proved the totipotency of adult somatic nuclei and finally confirmed that adult cells could differentiate without irreversible changes to the genetic material.

  3. "The Spotty Cow Tickled the Pig with a Curly Tail": How Do Sentence Position, Preferred Argument Structure, and Referential Complexity Affect Children's and Adults' Choice of Referring Expression?

    ERIC Educational Resources Information Center

    Theakston, Anna L.

    2012-01-01

    In this study, 5-year-olds and adults described scenes that differed according to whether (a) the subject or object of a transitive verb represented an accessible or inaccessible referent, consistent or inconsistent with patterns of preferred argument structure, and (b) a simple noun was sufficient to uniquely identify an inaccessible referent.…

  4. The Clone Factory

    ERIC Educational Resources Information Center

    Stoddard, Beryl

    2005-01-01

    Have humans been cloned? Is it possible? Immediate interest is sparked when students are asked these questions. In response to their curiosity, the clone factory activity was developed to help them understand the process of cloning. In this activity, students reenact the cloning process, in a very simplified simulation. After completing the…

  5. Human cloning 2001.

    PubMed

    Healy, David L; Weston, Gareth; Pera, Martin F; Rombauts, Luk; Trounson, Alan O

    2002-05-01

    This review summaries human cloning from a clinical perspective. Natural human clones, that is, monozygotic twins, are increasing in the general community. Iatrogenic human clones have been produced for decades in infertile couples given fertility treatment such as ovulation induction. A clear distinction must be made between therapeutic cloning using embryonic stem cells and reproductive cloning attempts. Unlike the early clinical years of in vitro fertilization, with cloning there is no animal model that is safe and dependable. Until there is such a model, 'Dolly'-style human cloning is medically unacceptable.

  6. Phenotype of cloned mice: development, behavior, and physiology.

    PubMed

    Tamashiro, Kellie L K; Wakayama, Teruhiko; Yamazaki, Yukiko; Akutsu, Hidenori; Woods, Stephen C; Kondo, Sylvia; Yanagimachi, Ryuzo; Sakai, Randall R

    2003-11-01

    Cloning technology has potential to be a valuable tool in basic research, clinical medicine, and agriculture. However, it is critical to determine the consequences of this technique in resulting offspring before widespread use of the technology. Mammalian cloning using somatic cells was first demonstrated in sheep in 1997 and since then has been extended to a number of other species. We examined development, behavior, physiology, and longevity in B6C3F1 female mice cloned from adult cumulus cells. Control mice were naturally fertilized embryos subjected to the same in vitro manipulation and culture conditions as clone embryos. Clones attained developmental milestones similar to controls. Activity level, motor ability and coordination, and learning and memory skills of cloned mice were comparable with controls. Interestingly, clones gained more body weight than controls during adulthood. Increased body weight was attributable to higher body fat and was associated with hyperleptinemia and hyperinsulinemia indicating that cloned mice are obese. Cloned mice were not hyperphagic as adults and had hypersensitive leptin and melanocortin signaling systems. Longevity of cloned mice was comparable with that reported by the National Institute on Aging and the causes of death were typical for this strain of mouse. These studies represent the first comprehensive set of data to characterize cloned mice and provide critical information about the long-term effects of somatic cell cloning. PMID:14610260

  7. Who is the parent in cloning?

    PubMed

    Elster, N

    1999-01-01

    In July 1996, a sheep named Dolly was born in Scotland. What makes Dolly's birth noteworthy is that she is the result of the first successful cloning attempt using the nucleus of an adult cell. The technique that led to Dolly's birth involved transferring the nucleus of a mammary cell from an adult sheep to the enucleated egg cell of an unrelated sheep with gestation occurring in a third sheep. The possibility of applying this technique to human reproduction raised concerns worldwide with several countries moving for an immediate bans on human cloning. In the United States, President Clinton requested that the National Bioethics Advisory Commission ("NBAC"), a multidisciplinary group composed of scientists, lawyers, educators, theologians, and ethicists study the implications of cloning and issue recommendations. The Commission consulted other scientists, ethicists, theologians, lawyers, and citizens with interests in this advancing technology and concluded that, "at this time it is morally unacceptable for anyone in the public or private sector, whether in a research or clinical setting, to attempt to create a child using somatic cell nuclear transfer cloning." This Article was included in a larger work prepared at the request of, and submitted to the Commission by, law professor Lori B. Andrews. Cloning through nuclear transfer will change the way we create and define families. This Article explores how existing law relating to parentage, surrogacy, egg donation, and artificial insemination may apply in the cloning context to clarify the parent-child relationship established through cloning.

  8. Who is the parent in cloning?

    PubMed

    Elster, N

    1999-01-01

    In July 1996, a sheep named Dolly was born in Scotland. What makes Dolly's birth noteworthy is that she is the result of the first successful cloning attempt using the nucleus of an adult cell. The technique that led to Dolly's birth involved transferring the nucleus of a mammary cell from an adult sheep to the enucleated egg cell of an unrelated sheep with gestation occurring in a third sheep. The possibility of applying this technique to human reproduction raised concerns worldwide with several countries moving for an immediate bans on human cloning. In the United States, President Clinton requested that the National Bioethics Advisory Commission ("NBAC"), a multidisciplinary group composed of scientists, lawyers, educators, theologians, and ethicists study the implications of cloning and issue recommendations. The Commission consulted other scientists, ethicists, theologians, lawyers, and citizens with interests in this advancing technology and concluded that, "at this time it is morally unacceptable for anyone in the public or private sector, whether in a research or clinical setting, to attempt to create a child using somatic cell nuclear transfer cloning." This Article was included in a larger work prepared at the request of, and submitted to the Commission by, law professor Lori B. Andrews. Cloning through nuclear transfer will change the way we create and define families. This Article explores how existing law relating to parentage, surrogacy, egg donation, and artificial insemination may apply in the cloning context to clarify the parent-child relationship established through cloning. PMID:12650149

  9. Cloning the Dmrt1 and DmrtA2 genes of ayu (Plecoglossus altivelis) and mapping their expression in adult, larval, and embryonic stages

    PubMed Central

    WANG, Jin-Hua; MIAO, Liang; LI, Ming-Yun; GUO, Xiao-Fei; PAN, Na; CHEN, Ying-Ying; ZHAO, Liang

    2014-01-01

    The Dmrt family of genes are involved in sex differentiation in different species of invertebrates, and some vertebrates including human. In this study, we cloned the full-length cDNA of ayu (Plecoglossus altivelis) Dmrt1 and DmrtA2. Sequence and phylogenetic tree analyses showed ayu Dmrt1 showed highest similarity to that of Oncorhynchus mykiss while ayu DmrtA2 is most similar to that of Oryzias latipes. Fluorescence-based quantitative reverse transcription PCR (qRT-PCR) revealed the Dmrt1 was predominantly expressed in the testis. At the larval stages, Dmrt1 mRNA expression level was highest during 52-64 days post hatching (dph) and at the gastrula stage during embryonic development. DmrtA2, meanwhile, was specifically expressed in the ovary and was highly expressed in the female brain tissue, but not male brain tissue. During the larval stages, DmrtA2 expression remained high before day 34, and then fluctuated while generally decreasing. During embryonic development, DmrtA2 expression increased gradually and peaked at the hatching stage. Our data suggest that ayu Dmrt1 might participate in the differentiation and maintenance of testis while DmrtA2 may play a role in ovary-differentiation and mature-ovary maintenance. DmrtA2 might also participate in brain development. PMID:24668652

  10. WILD PIG ATTACKS ON HUMANS

    SciTech Connect

    Mayer, J.

    2013-04-12

    Attacks on humans by wild pigs (Sus scrofa) have been documented since ancient times. However, studies characterizing these incidents are lacking. In an effort to better understand this phenomenon, information was collected from 412 wild pig attacks on humans. Similar to studies of large predator attacks on humans, data came from a variety of sources. The various attacks compiled occurred in seven zoogeographic realms. Most attacks occurred within the species native range, and specifically in rural areas. The occurrence was highest during the winter months and daylight hours. Most happened under non-hunting circumstances and appeared to be unprovoked. Wounded animals were the chief cause of these attacks in hunting situations. The animals involved were typically solitary, male and large in size. The fate of the wild pigs involved in these attacks varied depending upon the circumstances, however, most escaped uninjured. Most human victims were adult males traveling on foot and alone. The most frequent outcome for these victims was physical contact/mauling. The severity of resulting injuries ranged from minor to fatal. Most of the mauled victims had injuries to only one part of their bodies, with legs/feet being the most frequent body part injured. Injuries were primarily in the form of lacerations and punctures. Fatalities were typically due to blood loss. In some cases, serious infections or toxemia resulted from the injuries. Other species (i.e., pets and livestock) were also accompanying some of the humans during these attacks. The fates of these animals varied from escaping uninjured to being killed. Frequency data on both non-hunting and hunting incidents of wild pig attacks on humans at the Savannah River Site, South Carolina, showed quantitatively that such incidents are rare.

  11. Impact of cloning on cattle breeding systems.

    PubMed

    McClintock, A E

    1998-01-01

    The concept of clone-family testing is compared with existing progeny testing systems. The critical factors that will decide how cloning is utilized are the potential size of cloned families, and the cost per embryo (or per calf born). If family sizes of 100,000 become routinely achievable (cheaply), then clone testing becomes viable. In rough figures, cloned embryos costing $30 with a 50% calving rate would be attractive to farmers and would be cheap enough that farmers would buy more (crossbred) embryos in order to breed further replacement cows. At $300 per embryo, farmers would be more inclined to buy a number of cloned pure-bred female embryos and then to use conventional artificial insemination to breed further replacements from these superior cows. At $3000 per embryo, farmers would probably only be interested in very small numbers of cloned animals, most of which would be males. The relative importance of adult versus fetal cloning is discussed. The need for gene banks to preserve genetic variation is emphasized; both gametes and somatic tissue cultures should be considered.

  12. Genome-wide epigenetic alterations in cloned bovine fetuses.

    PubMed

    Cezar, Gabriela Gebrin; Bartolomei, Marisa S; Forsberg, Erik J; First, Neal L; Bishop, Michael D; Eilertsen, Kenneth J

    2003-03-01

    To gain a better understanding of global methylation differences associated with development of nuclear transfer (NT)-generated cattle, we analyzed the genome-wide methylation status of spontaneously aborted cloned fetuses, cloned fetuses, and adult clones that were derived from transgenic and nontransgenic cumulus, genital ridge, and body cell lines. Cloned fetuses were recovered from ongoing normal pregnancies and were morphologically normal. Fetuses generated by artificial insemination (AI) were used as controls. In vitro fertilization (IVF) fetuses were compared with AI controls to assess effects of in vitro culture on the 5-methylcytosine content of fetal genomes. All of the fetuses were female. Skin biopsies were obtained from cloned and AI-generated adult cows. All of the adult clones were phenotypically normal and lactating and had no history of health or reproductive disorders. Genome-wide cytosine methylation levels were monitored by reverse-phase HPLC, and results indicated reduced levels of methylated cytosine in NT-generated fetuses. In contrast, no differences were observed between adult, lactating clones and similarly aged lactating cows produced by AI. These data imply that survivability of cloned cattle may be closely related to the global DNA methylation status. This is the first report to indicate that global methylation losses may contribute to the developmental failure of cloned bovine fetuses. PMID:12604655

  13. Innexins in the lobster stomatogastric nervous system: cloning, phylogenetic analysis, developmental changes and expression within adult identified dye and electrically coupled neurons.

    PubMed

    Ducret, E; Alexopoulos, H; Le Feuvre, Y; Davies, J A; Meyrand, P; Bacon, J P; Fénelon, V S

    2006-12-01

    Gap junctions play a key role in the operation of neuronal networks by enabling direct electrical and metabolic communication between neurons. Suitable models to investigate their role in network operation and plasticity are invertebrate motor networks, which are built of comparatively few identified neurons, and can be examined throughout development; an excellent example is the lobster stomatogastric nervous system. In invertebrates, gap junctions are formed by proteins that belong to the innexin family. Here, we report the first molecular characterization of two crustacean innexins: the lobster Homarus gammarus innexin 1 (Hg-inx1) and 2 (Hg-inx2). Phylogenetic analysis reveals that innexin gene duplication occurred within the arthropod clade before the separation of insect and crustacean lineages. Using in situ hybridization, we find that each innexin is expressed within the adult and developing lobster stomatogastric nervous system and undergoes a marked down-regulation throughout development within the stomatogastric ganglion (STG). The number of innexin expressing neurons is significantly higher in the embryo than in the adult. By combining in situ hybridization, dye and electrical coupling experiments on identified neurons, we demonstrate that adult neurons that express at least one innexin are dye and electrically coupled with at least one other STG neuron. Finally, two STG neurons display no detectable amount of either innexin mRNAs but may express weak electrical coupling with other STG neurons, suggesting the existence of other forms of innexins. Altogether, we provide evidence that innexins are expressed within small neuronal networks built of dye and electrically coupled neurons and may be developmentally regulated. PMID:17156373

  14. Cysticercosis: identification and cloning of protective recombinant antigens.

    PubMed

    Manoutcharian, K; Rosas, G; Hernandez, M; Fragoso, G; Aluja, A; Villalobos, N; Rodarte, L F; Sciutto, E

    1996-04-01

    We describe the cloning and the evaluation of the protective capacity of 5 recombinant antigens expressed during the cysticercus stage of both Taenia crassiceps and Taenia solium. A cDNA library was constructed in bacteriophage lambda ZAP using mRNA isolated from larvae of T. crassiceps of the ORF strain. The recombinant phage library was screened with polyclonal antibodies against 56- and 74-kDa protective antigen fractions. This screening identified 13 recombinant clones, 5 of which were also strongly recognized by pooled sera from pigs experimentally infected with T. solium. The native antigens are proteins of 56 (clones KETcl, 4, 7) and 74 and 78 kDa (clones KETc11, 12) of T. crassiceps cysticerci. Vaccination experiments using these 5 recombinant clones against murine cysticercosis point to the relevance of KETcl, 4, 7, and 12 in host protection, whereas immunization with the clone KETc11 does not modify the parasite load in females and facilitates the parasitosis in males. We report here the DNA and the deduced amino acid sequence (100 amino acids) of the first protective antigen (KETc7) of potential interest for T. solium pig cysticercosis prevention.

  15. Cleaning pipelines: a pigging primer

    SciTech Connect

    Kipin, P.

    1985-02-04

    The ''pig'', a cleaning device currently used to clear out pipes, is discussed here. Types of pigs are described and include styrofoam, rubber, and soft foam. The limitations to the use of pigs are discussed. Unless all valves are fully open, a pig can get stuck. Ball-type tees may cause a short pig to drop and bypass. Generally, no pig is able to traverse a one-cut miter.

  16. Mammalian cloning: possibilities and threats.

    PubMed

    Mitalipov, S M; Wolf, D P

    2000-10-01

    The cloning of mammals originated with the production of limited numbers of genetically identical offspring by blastomere separation or embryo splitting. In the past few years, remarkable progress has been reported in cloning by nuclear transfer (NT) with donor nuclei recovered from embryonic, fetal or adult cells. Factors that contribute to the successful reprogramming of the transferred nucleus and the normal term development of the newly reconstructed embryo include the cell cycle stage of both the donor nucleus and recipient cytoplast, the timing of fusion and cytoplast activation, and the source of donor nuclei. The possibility of producing live offspring by somatic cell NT carries potential applications in animal husbandry, biotechnology, transgenic and pharmaceutical production, biomedical research, and the preservation of endangered species. However, the low efficiencies of cloning by NT coupled with high embryonic, fetal and neonatal losses may restrict immediate commercial applications in agriculture. These limitations notwithstanding, the greatest benefits and practical implications of this new technology could be in transplantation medicine and therapeutic cloning.

  17. When pigs fly, UCP1 makes heat.

    PubMed

    Jastroch, Martin; Andersson, Leif

    2015-05-01

    Brown and beige adipose tissue may represent important therapeutic targets for the treatment of diabetes and obesity as these organs dissipate nutrient energy as heat through the thermogenic uncoupling protein 1 (UCP1). While mice are commonly used to mimic the potential effects of brown/beige adipose tissue that may act in human metabolism, new animal models are edging into the market for translational medicine. Pigs reflect human metabolism better than mice in multiple parameters such as obesity-induced hyperglycemia, cholesterol profiles and energy metabolism. Recently, it was reported that energy expenditure and body temperature in pigs is induced by the hormone leptin, and that leptin's action is mediated by UCP1 in adipose tissue. Given the tremendous importance of identifying molecular mechanisms for targeting therapeutics, we critically examine the evidence supporting the presence of UCP1 in pigs and conclude that methodological shortcomings prevent an unequivocal claim for the presence of UCP1 in pigs. Despite this, we believe that leptin's effects on energy expenditure in pigs are potentially more transformative to human medicine in the absence of UCP1, as adult and obese humans possess only minor amounts of UCP1. In general, we propose that the biology of new animal models requires attention to comparative studies with humans given the increasing amount of genomic information for various animal species.

  18. Multipartite asymmetric quantum cloning

    SciTech Connect

    Iblisdir, S.; Gisin, N.; Acin, A.; Cerf, N.J.; Filip, R.; Fiurasek, J.

    2005-10-15

    We investigate the optimal distribution of quantum information over multipartite systems in asymmetric settings. We introduce cloning transformations that take N identical replicas of a pure state in any dimension as input and yield a collection of clones with nonidentical fidelities. As an example, if the clones are partitioned into a set of M{sub A} clones with fidelity F{sup A} and another set of M{sub B} clones with fidelity F{sup B}, the trade-off between these fidelities is analyzed, and particular cases of optimal N{yields}M{sub A}+M{sub B} cloning machines are exhibited. We also present an optimal 1{yields}1+1+1 cloning machine, which is an example of a tripartite fully asymmetric cloner. Finally, it is shown how these cloning machines can be optically realized.

  19. Ethical issues in cloning.

    PubMed

    Satris, S

    2000-01-01

    There is great public concern with the ethics of human cloning. This paper briefly examines some of what I identify as pseudo-problems or myths associated with cloning, and some of the more substantial ethical concerns.

  20. The identity of clones.

    PubMed

    Evers, K

    1999-02-01

    A common concern with respect to cloning is based on the belief that cloning produces identical individuals. This is a fundamental misunderstanding of what type of identity-relation cloning involves. The concept "identity" is ambiguous, and the statement that cloning produces "identical" individuals is not meaningful unless the notion of identity is clarified. This paper distinguishes between numerical and qualitative; relational and intrinsic: logical and empirical identity, and discusses the empirical individuation of clones in terms of genetics, physiology, perception, cognition and personality. I argue that the only relation of identity cloning involves is qualitative, intrinsic and empirical: genetic indiscernibility, unlikely to include identity under other aspects mentioned. A popular argument against cloning claims our "right" to a "unique identity". This objection either implies (absurdly) the right not to be an identical twin, or assumes (incorrectly) that cloning involves identity other than genetic. Either way, the argument is untenable.

  1. Aristotle and headless clones.

    PubMed

    Mosteller, Timothy

    2005-01-01

    Cloned organisms can be genetically altered so that they do not exhibit higher brain functioning. This form of therapeutic cloning allows for genetically identical organs and tissues to be harvested from the clone for the use of the organism that is cloned. "Spare parts" cloning promises many opportunities for future medical advances. What is the ontological and ethical status of spare parts, headless clones? This paper attempts to answer this question from the perspective of Aristotle's view of the soul. Aristotle's metaphysics as applied to his view of biological essences generates an ethic that can contribute to moral reasoning regarding the use of headless spare parts clones. The task of this paper is to show the implications that Aristotle's view of the soul, if it is true, would have on the ethics of headless, spare parts cloning. PMID:16180113

  2. Aristotle and headless clones.

    PubMed

    Mosteller, Timothy

    2005-01-01

    Cloned organisms can be genetically altered so that they do not exhibit higher brain functioning. This form of therapeutic cloning allows for genetically identical organs and tissues to be harvested from the clone for the use of the organism that is cloned. "Spare parts" cloning promises many opportunities for future medical advances. What is the ontological and ethical status of spare parts, headless clones? This paper attempts to answer this question from the perspective of Aristotle's view of the soul. Aristotle's metaphysics as applied to his view of biological essences generates an ethic that can contribute to moral reasoning regarding the use of headless spare parts clones. The task of this paper is to show the implications that Aristotle's view of the soul, if it is true, would have on the ethics of headless, spare parts cloning.

  3. Chromatin remodeling in nuclear cloning.

    PubMed

    Wade, Paul A; Kikyo, Nobuaki

    2002-05-01

    Nuclear cloning is a procedure to create new animals by injecting somatic nuclei into unfertilized oocytes. Recent successes in mammalian cloning with differentiated adult nuclei strongly indicate that oocyte cytoplasm contains unidentified remarkable reprogramming activities with the capacity to erase the previous memory of cell differentiation. At the heart of this nuclear reprogramming lies chromatin remodeling as chromatin structure and function define cell differentiation through regulation of the transcriptional activities of the cells. Studies involving the modification of chromatin elements such as selective uptake or release of binding proteins, covalent histone modifications including acetylation and methylation, and DNA methylation should provide significant insight into the molecular mechanisms of nuclear dedifferentiation and redifferentiation in oocyte cytoplasm.

  4. Generation of α1,3-galactosyltransferase and cytidine monophospho-N-acetylneuraminic acid hydroxylase gene double-knockout pigs.

    PubMed

    Miyagawa, Shuji; Matsunari, Hitomi; Watanabe, Masahito; Nakano, Kazuaki; Umeyama, Kazuhiro; Sakai, Rieko; Takayanagi, Shuko; Takeishi, Toki; Fukuda, Tooru; Yashima, Sayaka; Maeda, Akira; Eguchi, Hiroshi; Okuyama, Hiroomi; Nagaya, Masaki; Nagashima, Hiroshi

    2015-01-01

    Zinc-finger nucleases (ZFNs) and transcription activator-like effector nucleases (TALENs) are new tools for producing gene knockout (KO) animals. The current study reports produced genetically modified pigs, in which two endogenous genes were knocked out. Porcine fibroblast cell lines were derived from homozygous α1,3-galactosyltransferase (GalT) KO pigs. These cells were subjected to an additional KO for the cytidine monophospho-N-acetylneuraminic acid hydroxylase (CMAH) gene. A pair of ZFN-encoding mRNAs targeting exon 8 of the CMAH gene was used to generate the heterozygous CMAH KO cells, from which cloned pigs were produced by somatic cell nuclear transfer (SCNT). One of the cloned pigs obtained was re-cloned after additional KO of the remaining CMAH allele using the same ZFN-encoding mRNAs to generate GalT/CMAH-double homozygous KO pigs. On the other hand, the use of TALEN-encoding mRNAs targeting exon 7 of the CMAH gene resulted in efficient generation of homozygous CMAH KO cells. These cells were used for SCNT to produce cloned pigs homozygous for a double GalT/CMAH KO. These results demonstrate that the combination of TALEN-encoding mRNA, in vitro selection of the nuclear donor cells and SCNT provides a robust method for generating KO pigs. PMID:26227017

  5. Generation of α1,3-galactosyltransferase and cytidine monophospho-N-acetylneuraminic acid hydroxylase gene double-knockout pigs

    PubMed Central

    MIYAGAWA, Shuji; MATSUNARI, Hitomi; WATANABE, Masahito; NAKANO, Kazuaki; UMEYAMA, Kazuhiro; SAKAI, Rieko; TAKAYANAGI, Shuko; TAKEISHI, Toki; FUKUDA, Tooru; YASHIMA, Sayaka; MAEDA, Akira; EGUCHI, Hiroshi; OKUYAMA, Hiroomi; NAGAYA, Masaki; NAGASHIMA, Hiroshi

    2015-01-01

    Zinc-finger nucleases (ZFNs) and transcription activator-like effector nucleases (TALENs) are new tools for producing gene knockout (KO) animals. The current study reports produced genetically modified pigs, in which two endogenous genes were knocked out. Porcine fibroblast cell lines were derived from homozygous α1,3-galactosyltransferase (GalT) KO pigs. These cells were subjected to an additional KO for the cytidine monophospho-N-acetylneuraminic acid hydroxylase (CMAH) gene. A pair of ZFN-encoding mRNAs targeting exon 8 of the CMAH gene was used to generate the heterozygous CMAH KO cells, from which cloned pigs were produced by somatic cell nuclear transfer (SCNT). One of the cloned pigs obtained was re-cloned after additional KO of the remaining CMAH allele using the same ZFN-encoding mRNAs to generate GalT/CMAH-double homozygous KO pigs. On the other hand, the use of TALEN-encoding mRNAs targeting exon 7 of the CMAH gene resulted in efficient generation of homozygous CMAH KO cells. These cells were used for SCNT to produce cloned pigs homozygous for a double GalT/CMAH KO. These results demonstrate that the combination of TALEN-encoding mRNA, in vitro selection of the nuclear donor cells and SCNT provides a robust method for generating KO pigs. PMID:26227017

  6. A comparative study on expression profile of developmentally important genes during pre-implantation stages in buffalo hand-made cloned embryos derived from adult fibroblasts and amniotic fluid derived stem cells.

    PubMed

    Em, Sadeesh; Shah, Fozia; Kataria, Meena; Yadav, P S

    2016-08-01

    Abnormal gene expression in somatic cell nuclear transfer embryos due to aberrant epigenetic modifications of the donor nucleus may account for much of the observed diminished viability and developmental abnormalities. The present study compared the developmentally important gene expression pattern at 4-cell, 8- to 16-cell, morula, and blastocyst stages of buffalo nuclear transfer (NT) embryos from adult fibroblasts (AFs) and amniotic fluid stem cells (AFSCs). In vitro fertilized embryos were used as control embryos. Alterations in the expression pattern of genes implicated in transcription and pluripotency (OCT4, STAT3, NANOG), DNA methylation (DNMT1, DNMT3A), histone deacetylation (HDAC2), growth factor signaling, and imprinting (IGF2, IGF2R), apoptosis (BAX, BCL2), oxidative stress (MnSOD), metabolism (GLUT1) regulation were observed in cloned embryos. The expression of transcripts in AFSC-NT embryos more closely followed that of the in vitro fertilized embryos compared with AF-NT embryos. It is concluded that AFSCs with a relatively undifferentiated genome may serve as suitable donors which could be reprogrammed more efficiently to reactivate expression of early embryonic genes in buffalo NT.

  7. Pig in the Middle.

    ERIC Educational Resources Information Center

    Mills, Sophie

    2000-01-01

    Explores themes relating to human transition as they appear in "Charlotte's Web" and four other stories using pigs as a subject. Discusses the motifs common to all these texts that recur in the film "Babe." Considers how the cycle of life and death is ceaseless, and pigs symbolize the necessary transitions that people must all make. (NH)

  8. Tail biting in pigs.

    PubMed

    Schrøder-Petersen, D L; Simonsen, H B

    2001-11-01

    One of the costly and welfare-reducing problems in modern pig production is tail biting. Tail biting is an abnormal behaviour, characterized by one pig's dental manipulation of another pig's tail. Tail biting can be classified into two groups: the pre-injury stage, before any wound on the tail is present, and the injury stage, where the tail is wounded and bleeding. Tail biting in the injury stage will reduce welfare of the bitten pig and the possible spread of infection is a health as well as welfare problem. The pigs that become tail biters may also suffer, because they are frustrated due to living in a stressful environment. This frustration may result in an excessive motivation for biting the tails of pen mates. This review aims to summarize recent research and theories in relation to tail biting. PMID:11681870

  9. Production of diabetic offspring using cryopreserved epididymal sperm by in vitro fertilization and intrafallopian insemination techniques in transgenic pigs.

    PubMed

    Umeyama, Kazuhiro; Honda, Kasumi; Matsunari, Hitomi; Nakano, Kazuaki; Hidaka, Tatsuro; Sekiguchi, Keito; Mochizuki, Hironori; Takeuchi, Yasuhiro; Fujiwara, Tsukasa; Watanabe, Masahito; Nagaya, Masaki; Nagashima, Hiroshi

    2013-12-17

    Somatic cell nuclear transfer (SCNT) is a useful technique for creating pig strains that model human diseases. However, production of numerous cloned disease model pigs by SCNT for large-scale experiments is impractical due to its complexity and inefficiency. In the present study, we aimed to establish an efficient procedure for proliferating the diabetes model pig carrying the mutant human hepatocyte nuclear factor-1α gene. A founder diabetes transgenic cloned pig was generated by SCNT and treated with insulin to allow for normal growth to maturity, at which point epididymal sperm could be collected for cryopreservation. In vitro fertilization and intrafallopian insemination using the cryopreserved epididymal sperm resulted in diabetes model transgenic offspring. These results suggest that artificial reproductive technology using cryopreserved epididymal sperm could be a practical option for proliferation of genetically modified disease model pigs. PMID:23979397

  10. Production of Diabetic Offspring Using Cryopreserved Epididymal Sperm by In Vitro Fertilization and Intrafallopian Insemination Techniques in Transgenic Pigs

    PubMed Central

    UMEYAMA, Kazuhiro; HONDA, Kasumi; MATSUNARI, Hitomi; NAKANO, Kazuaki; HIDAKA, Tatsuro; SEKIGUCHI, Keito; MOCHIZUKI, Hironori; TAKEUCHI, Yasuhiro; FUJIWARA, Tsukasa; WATANABE, Masahito; NAGAYA, Masaki; NAGASHIMA, Hiroshi

    2013-01-01

    Abstract Somatic cell nuclear transfer (SCNT) is a useful technique for creating pig strains that model human diseases. However, production of numerous cloned disease model pigs by SCNT for large-scale experiments is impractical due to its complexity and inefficiency. In the present study, we aimed to establish an efficient procedure for proliferating the diabetes model pig carrying the mutant human hepatocyte nuclear factor-1α gene. A founder diabetes transgenic cloned pig was generated by SCNT and treated with insulin to allow for normal growth to maturity, at which point epididymal sperm could be collected for cryopreservation. In vitro fertilization and intrafallopian insemination using the cryopreserved epididymal sperm resulted in diabetes model transgenic offspring. These results suggest that artificial reproductive technology using cryopreserved epididymal sperm could be a practical option for proliferation of genetically modified disease model pigs. PMID:23979397

  11. Cloning: can it be good for us? An overview of cloning technology and its moral implications.

    PubMed

    FitzGerald, K

    2001-01-01

    Adequate answers to moral questions about cloning require a working knowledge of the science and technology involved, both present and anticipated. This essay presents an overview of the current state of somatic cell nuclear transfer technology (SCNT), the type of cloning that now permits whole organism reproduction from adult DNA. This essay explains the basic science and technology of SCNT and explores its potential uses. Next, this essay notes remaining scientific obstacles and unanswered moral questions that must be resolved before SCNT can be used for human reproduction. Attention is given to aspects related to cloning for therapeutic and research purposes.

  12. Production of transgenic miniature pigs by pronuclear microinjection.

    PubMed

    Uchida, M; Shimatsu, Y; Onoe, K; Matsuyama, N; Niki, R; Ikeda, J E; Imai, H

    2001-12-01

    Miniature pig is an attractive animal for a wide range of research fields, such as medicine and pharmacology, because of its small size, the possibility of breeding it under minimum environmental controls and the physiology that is potentially similar to that of human. Although transgenic technology is useful for the analysis of gene function and for the development of model animals for various diseases, there have not yet been any reports on producing transgenic miniature pig. This study is the first successful report concerning the production of transgenic miniature pig by pronuclear microinjection. The huntingtin gene cloned from miniature pig, which is a homologue of candidate gene for Huntington's disease, connected with rat neuron-specific enolase promoter region, was injected into a pronucleus of fertilized eggs with micromanipulator. The eggs were transferred into the oviduct of recipient miniature pigs, whose estrus cycles were previously synchronized with a progesterone analogue. A total of 402 injected eggs from 171 donors were transferred to 23 synchronized recipients. Sixteen of them maintained pregnancy and delivered 65 young, and one resulted in abortion. Five of the 68 offspring (three of which were aborted) were determined to have transgene by PCR and Southern analysis. The overall rate of transgenic production was 1.24% (transgenic/injected eggs). This study provides the first success and useful information regarding production of transgenic miniature pig for biomedical research.

  13. Cryptosporidium scrofarum n. sp. (Apicomplexa: Cryptosporidiidae) in domestic pigs (Sus scrofa)

    PubMed Central

    Kváč, Martin; Kestřánová, Michaela; Pinková, Martina; Květoňová, Dana; Kalinová, Jana; Wagnerová, Pavla; Kotková, Michaela; Vítovec, Jiří; Ditrich, Oleg; McEvoy, John; Stenger, Brianna; Sak, Bohumil

    2012-01-01

    We describe the morphological, biological, and molecular characteristics of Cryptosporidium pig genotype II and propose the species name Cryptosporidium scrofarum n. sp. to reflect its prevalence in adult pigs worldwide. Oocysts of C. scrofarum are morphologically indistinguishable from C. parvum, measuring 4.81–5.96 µm (mean = 5.16) × 4.23–5.29 µm (mean = 4.83) with a length to width ratio of 1.07 ± 0.06 (n = 400). Oocysts of C. scrofarum obtained from a naturally infected pig were infectious for 8-week-old pigs but not 4-week-old pigs. The prepatent period in 8-week-old Cryptosporidium-naive pigs was 4–6 days and the patent period was longer than 30 days. The infection intensity of C. scrofarum in pigs was generally low, in the range 250-4000 oocysts per gram of faeces. Infected pigs showed no clinical signs of cryptosporidiosis and no pathology was detected. Cryptosporidium scrofarum was not infectious for adult SCID mice, adult BALB c mice, Mongolian gerbils (Meriones unguiculatus), southern multimammate mice (Mastomys coucha), yellow-necked mice (Apodemus flavicollis), or guinea pigs (Cavia porcellus). Phylogenetic analyses based on Small subunit rRNA, actin, and heat shock protein 70 gene sequences revealed that C. scrofarum is genetically distinct from all known Cryptosporidium species. PMID:23021264

  14. Widespread distribution of hepatitis E virus in Spanish pig herds

    PubMed Central

    2011-01-01

    Background Hepatitis E virus (HEV) infection is a serious health problem in developing countries and is also increasingly reported in industrialized regions. HEV is considered a zoonotic agent and strains isolated from swine and human sources are genetically similar. Thus, HEV is of increasing importance to both public and animal health. The aim of the present study was to evaluate the distribution of HEV in a large population of pigs from herds located in different autonomous regions throughout Spain. Results The presence of anti-HEV IgG antibodies was analyzed in 1141 swine serum samples (corresponding to 381 pigs younger than 6 months and 760 pigs older than 6 months) collected from 85 herds. Herds were located in 6 provinces in 4 autonomous regions throughout Spain. At least one pig tested positive for anti-HEV IgG in over 80% of herds. Of individual pigs, 20.4% (233/1141) were positive for anti-HEV IgG, with the prevalence being higher in adult pigs than in those under 6 months (30.2% vs. 15.5%). A subset of serum samples taken at 2- to 5-week intervals showed that seroprevalence dropped between 3 and 11 weeks of age, and then rose significantly by the 15th week. Pigs were also examined for the presence of HEV-RNA by RT-PCR. Of pigs tested for the presence of HEV-RNA 18.8% (64/341) were positive, with at least one pig in almost half of the herds testing positive. HEV-RNA amplicons from several positive pigs were sequenced and all were of genotype 3. Conclusions HEV was found to be widely distributed among swine farms across Spain, with the prevalence being highest among animals older than 6 months. These results indicate that HEV infection either is or is likely to become endemic in the Spanish swine population. PMID:21999141

  15. Induction of protection against porcine cysticercosis in growing pigs by DNA vaccination.

    PubMed

    Guo, Aijiang; Jin, Zhizhong; Zheng, Yadong; Hai, Gang; Yuan, Gailing; Li, Hailong; Cai, Xuepeng

    2007-01-01

    A DNA vaccine, pcDNA3-B, was developed by using the nucleotide sequence of Taenia solium B antigen and cloning into pcDNA3.1 plasmid. The growing pigs were vaccinated by one intramuscular infection of 200 or 1000 microg pcDNA3-B. The immunization with 1000 microg of pcDNA3-B showed 92.6% protection when the pigs were challenged by T. solium eggs and four of the five pigs vaccinated had no viable cysts. The results provide encouraging information on the use of pcDNA3-B vaccination for the prevention of cysticercosis.

  16. Production of α1,3-Galactosyltransferase–Deficient Pigs

    PubMed Central

    Phelps, Carol J.; Koike, Chihiro; Vaught, Todd D.; Boone, Jeremy; Wells, Kevin D.; Chen, Shu-Hung; Ball, Suyapa; Specht, Susan M.; Polejaeva, Irina A.; Monahan, Jeff A.; Jobst, Pete M.; Sharma, Sugandha B.; Lamborn, Ashley E.; Garst, Amy S.; Moore, Marilyn; Demetris, Anthony J.; Rudert, William A.; Bottino, Rita; Bertera, Suzanne; Trucco, Massimo; Starzl, Thomas E.; Dai, Yifan; Ayares, David L.

    2011-01-01

    The enzyme α1,3-galactosyltransferase (α1,3GT or GCTA1) synthesizes α1,3-galactose (α1,3Gal) epitopes (Galα1,3Galβ1,4GlcNAc-R), which are the major xenoantigens causing hyperacute rejection in pig-to-human xenotransplantation. Complete removal of α1,3Gal from pig organs is the critical step toward the success of xenotransplantation. We reported earlier the targeted disruption of one allele of the α1,3GT gene in cloned pigs. A selection procedure based on a bacterial toxin was used to select for cells in which the second allele of the gene was knocked out. Sequencing analysis demonstrated that knockout of the second allele of the α1,3GT gene was caused by a T-to-G single point mutation at the second base of exon 9, which resulted in inactivation of the α1,3GT protein. Four healthy α1,3GT double-knockout female piglets were produced by three consecutive rounds of cloning. The piglets carrying a point mutation in the α1,3GT gene hold significant value, as they would allow production of α1,3Gal-deficient pigs free of antibiotic-resistance genes and thus have the potential to make a safer product for human use. PMID:12493821

  17. To what extent can the law control human cloning?

    PubMed

    Wood, P G

    1999-01-01

    This paper explores the legal ramifications of human reproductive cloning in response to 'Dolly'--the first animal cloned from an adult cell. No attempt is made to address the complex moral and ethical dilemmas that will inevitably be consequential on future successes in human reproductive cloning. Some of the potential benefits of cloning are briefly summarized but discussion is focused primarily on the current state of the law in the UK and some other European jurisdictions. Attempts to legislate on human cloning in the US, the emerging role of the EU and amendments to the European Convention on Human Rights are outlined. The potential problems likely to be encountered in the enforcement of any global treaties or international regulations are highlighted. It is argued that attempting to control human cloning by imposing legal prohibition is futile and a pragmatic solution to this impending problem is required forthwith.

  18. Trap style influences wild pig behavior and trapping success

    USGS Publications Warehouse

    Williams, B.L.; Holtfreter, R.W.; Ditchkoff, S.S.; Grand, J.B.

    2011-01-01

    Despite the efforts of many natural resource professionals, wild pig (Sus scrofa) populations are expanding in many areas of the world. Although many creative techniques for controlling pig populations are being explored, trapping has been and still is themost commonly usedmethod of population control formany public and private land managers. We conducted an observational study to examine the efficiency of 2 frequently used trap styles: a small, portable box-style trap and a larger, semi-permanent, corral-style trap.We used game cameras to examine patterns of trap entry by wild pigs around each style of trap, and we conducted a trapping session to compare trapping success between trap styles. Adult female and juvenile wild pigs entered both styles of trap more readily than did adult males, and adult males seemed particularly averse to entering box traps. Less than 10% of adult male visits to box traps resulted in entries, easily the least percentage of any class at any style of trap. Adult females entered corral traps approximately 2.2 times more often per visit than box traps and re-entered corral traps >2 times more frequently. Juveniles entered and reentered both box and corral traps at similar rates. Overall (all-class) entry-per-visit rates at corral traps (0.71) were nearly double that of box traps (0.37). Subsequent trapping data supported these preliminary entry data; the capture rate for corral traps was >4 times that of box traps. Our data suggest that corral traps are temporally and economically superior to box traps with respect to efficiency; that is, corral traps effectively trap more pigs per trap night at a lower cost per pig than do box traps. ?? 2011 The Wildlife Society.

  19. Mechanical ventilation and sepsis impair protein metabolism in the diaphragm of neonatal pigs

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Mechanical ventilation (MV) impairs diaphragmatic function and diminishes the ability to wean from ventilatory support in adult humans. In normal neonatal pigs, animals that are highly anabolic, endotoxin (LPS) infusion induces sepsis, reduces peripheral skeletal muscle protein synthesis rates, but ...

  20. Expression of NMS and NMU2R in the pig reproductive axis during the estrus cycle and the effect of NMS on the reproductive axis in vitro.

    PubMed

    Yang, Guihong; Su, Juan; Li, Xun; Yao, Yuan; Lei, Zhihai; Yang, Xizhi; Kou, Rui; Liu, Yanpeng

    2009-12-01

    Evidence has revealed that neuromedin S (NMS) and neuromedin U-receptor type-2 (NMU2R) mRNAs are expressed in the central nervous system and reproductive organs. Previous data indicated that variation of NMS and NMU2R was due to the phases of the adult rat hypothalamus estrus cycle. However, the expression and function of NMS in the pig reproductive axis remains unexplored. In this study, 16 virginal gilts were classified into four groups: proestrus, estrus, diestrus 1, and diestrus 2; the expression of NMS and NMU2R in the cyclic pig hypothalamus-pituitary-ovary axis was studied by reverse transcriptaion-polymerase chain reaction (RT-PCR), and the effect of NMS on the reproductive axis in vitro was detected by radioimmunoassay (RIA). The cloned pig NMS and NMU2R sequences were 82% and 90.2% identical to those of the corresponding human homologues, respectively. RT-PCR showed that NMS and NMU2R mRNA expression in the hypothalamus and pituitary changed with the estrus cycle, i.e., with the highest level in the proestrus group and the lowest in the estrus group. In the ovary, NMS and NMU2R expression was highest in the diestrus 2 group and the lowest in the proestrus group. In the in vitro study, different concentrations of NMS induced the release of gonadotropin-releasing hormone, luteinizing hormone, and estradiol at different levels of the reproductive axis. Taken together, the expression pattern of NMS during the estrus cycle and its role in reproductive hormones in vitro provide novel evidences of the potential roles of NMS in the regulation of pig reproduction.

  1. Procreative liberty, enhancement and commodification in the human cloning debate.

    PubMed

    Shapshay, Sandra

    2012-12-01

    The aim of this paper is to scrutinize a contemporary standoff in the American debate over the moral permissibility of human reproductive cloning in its prospective use as a eugenic enhancement technology. I shall argue that there is some significant and under-appreciated common ground between the defenders and opponents of human cloning. Champions of the moral and legal permissibility of cloning support the technology based on the right to procreative liberty provided it were to become as safe as in vitro fertilization and that it be used only by adults who seek to rear their clone children. However, even champions of procreative liberty oppose the commodification of cloned embryos, and, by extension, the resulting commodification of the cloned children who would be produced via such embryos. I suggest that a Kantian moral argument against the use of cloning as an enhancement technology can be shown to be already implicitly accepted to some extent by champions of procreative liberty on the matter of commodification of cloned embryos. It is in this argument against commodification that the most vocal critics of cloning such as Leon Kass and defenders of cloning such as John Robertson can find greater common ground. Thus, I endeavor to advance the debate by revealing a greater degree of moral agreement on some fundamental premises than hitherto recognized.

  2. Procreative liberty, enhancement and commodification in the human cloning debate.

    PubMed

    Shapshay, Sandra

    2012-12-01

    The aim of this paper is to scrutinize a contemporary standoff in the American debate over the moral permissibility of human reproductive cloning in its prospective use as a eugenic enhancement technology. I shall argue that there is some significant and under-appreciated common ground between the defenders and opponents of human cloning. Champions of the moral and legal permissibility of cloning support the technology based on the right to procreative liberty provided it were to become as safe as in vitro fertilization and that it be used only by adults who seek to rear their clone children. However, even champions of procreative liberty oppose the commodification of cloned embryos, and, by extension, the resulting commodification of the cloned children who would be produced via such embryos. I suggest that a Kantian moral argument against the use of cloning as an enhancement technology can be shown to be already implicitly accepted to some extent by champions of procreative liberty on the matter of commodification of cloned embryos. It is in this argument against commodification that the most vocal critics of cloning such as Leon Kass and defenders of cloning such as John Robertson can find greater common ground. Thus, I endeavor to advance the debate by revealing a greater degree of moral agreement on some fundamental premises than hitherto recognized. PMID:22983766

  3. Method for cloning genes

    SciTech Connect

    Weissman, S.M.; Pereira, D.; Sood, A.

    1988-04-19

    This patent describes a recombinant cloning vehicle comprising an inserted human gene, the improvement wherein the cloning vehicle is isolated from a recombinant clone which is isolated and identified by a process comprising the steps of: (a) effecting cDNA synthesis on a mixture of mRNAs containing a target mRNA coding for a major hisitocompatibility antigen, and isolating the resultant cDNA mixture; (b) inserting the resultant cDNA into recombinant cloning vehicles, and transforming hosts with the vehicles; and (c) separating the transformants and isolating and identifying a recombinant clone containing a DNA segment which is homologous over at least a portion thereof to at least one oligonucleotide probe specific for the DNA segment.

  4. Cloning for human reproduction: one American perspective.

    PubMed

    Chester, R

    2001-09-01

    The author, an American law professor, believes that whole-body cloning of adult humans will be possible in the near future. He does not believe the procedure should be banned when used as a form of assisted reproduction, but that it should be regulated by the government to ensure proper testing and application. After raising a number of scientific, ethical, religious and legal issues, Professor Chester addresses parentage in light of both old and new concepts of the 'family.' Finally, he focuses on the problem of women as surrogate mothers of clones, arguing in the process that the surrogate, having no real genetic tie to the clone, would have less of a claim to parentage than at least some of the surrogates currently gestating foetuses.

  5. A divergent clade of circular single-stranded DNA viruses from pig feces

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Using metagenomics and molecular cloning methods, we characterized five novel small circular viral genomes from pig feces distantly related to chimpanzee and porcine stool-associated circular viruses, (ChiSCV and PoSCV1). Phylogenetic analysis placed these viruses into a new, highly divergent, clade...

  6. On cloning human beings.

    PubMed

    de Melo-Martin, Inmaculada

    2002-06-01

    The purpose of this paper is to show that arguments for and against cloning fail to make their case because of one or both of the following reasons: 1) they take for granted customary beliefs and assumptions that are far from being unquestionable; 2) they tend to ignore the context in which human cloning is developed. I will analyze some of the assumptions underlying the main arguments that have been offered for and against cloning. Once these assumptions are critically analyzed, arguments both rejecting and supporting human cloning seem to lose weight. I will first briefly present the main arguments that have been proposed against cloning and I will argue that they fail to establish their case. In the next section I will evaluate some of the positive arguments that have been offered supporting such technology. This analysis will show that the case for cloning also fails. Finally, I will maintain that because critics and especially supporters of this technology neglect the context in which human cloning is developed and might be implemented, their arguments are far from compelling.

  7. Urolithiasis in finishing pigs.

    PubMed

    Maes, D G D; Vrielinck, J; Millet, S; Janssens, G P J; Deprez, P

    2004-11-01

    Urolithiasis in sows and neonatal pigs is well-known, but information on its occurrence and impact in finishing pigs is sparse. This study reports three outbreaks of urolithiasis in finishing pigs. In one herd, no symptoms were observed, whereas in the other herds the presence of calculi caused obstruction of the urinary tract resulting in death. Using infra-red spectroscopy, the predominant mineral-type found in the uroliths was calcium carbonate (calcite). Only small amounts of calcium oxalate (< 1%) could be detected. A high urinary pH, small abnormalities in the mineral composition of the feed and insufficient drinking water were the most important risk factors identified. To prevent urolithiasis, it is important to ensure adequate water intake, to provide a balanced mineral diet, and to avoid urinary tract infections.

  8. A somatic cell hybrid panel for pig regional gene mapping characterized by molecular cytogenetics.

    PubMed

    Yerle, M; Echard, G; Robic, A; Mairal, A; Dubut-Fontana, C; Riquet, J; Pinton, P; Milan, D; Lahbib-Mansais, Y; Gellin, J

    1996-01-01

    A panel of 27 pig x rodent somatic cell hybrids was produced and characterized cytogenetically. The first step of this study consisted of hybridizing a SINE probe to GTG-banded metaphases of each hybrid clone in order to count and identify the normal pig chromosomes and to detect rearranged ones. The second step consisted of using the DNA of each clone as a probe after pIRS-PCR (porcine interspersed repetitive sequence-polymerase chain reaction) amplification to highly enrich it in pig sequences. These probes, hybridized to normal pig metaphase chromosomes, enabled the identification of the complete porcine complement in the hybrid lines. Whole chromosomes and fragments were characterized quickly and precisely, and results were compared. In addition to this cytogenetic characterization, molecular verification was also carried out by using primers specific to six microsatellites and to one gene previously mapped to pig chromosomes. The results obtained allow us to conclude that we have produced a panel that is informative for all porcine chromosomes. This panel constitutes a highly efficient tool to establish not only assignments of genes and markers but also regional localizations on pig chromosomes. PMID:8697807

  9. Benefits and problems with cloning animals.

    PubMed

    Smith, L C; Bordignon, V; Babkine, M; Fecteau, G; Keefer, C

    2000-12-01

    Animal cloning is becoming a useful technique for producing transgenic farm animals and is likely to be used to produce clones from valuable adults. Other applications will also undoubtedly be discovered in the near future, such as for preserving endangered breeds and species. Although cloning promises great advantages for commerce and research alike, its outcome is not always certain due to high pregnancy losses and high morbidity and mortality during the neonatal period. Research into the mechanisms involved in the reprogramming of the nucleus is being conducted throughout the world in an attempt to better understand the molecular and cellular mechanisms involved in correcting these problems. Although the cause of these anomalies remains mostly unknown, similar phenotypes have been observed in calves derived through in vitro fertilization, suggesting that culture conditions are involved in these phenomena. In the meantime, veterinarians and theriogenologists have an important role to play in improving the efficiency of cloning by finding treatments to assure normal gestation to term and to develop preventative and curative care for cloned neonates.

  10. Investigating the geometry of pig airways using computed tomography

    NASA Astrophysics Data System (ADS)

    Mansy, Hansen A.; Azad, Md Khurshidul; McMurray, Brandon; Henry, Brian; Royston, Thomas J.; Sandler, Richard H.

    2015-03-01

    Numerical modeling of sound propagation in the airways requires accurate knowledge of the airway geometry. These models are often validated using human and animal experiments. While many studies documented the geometric details of the human airways, information about the geometry of pig airways is scarcer. In addition, the morphology of animal airways can be significantly different from that of humans. The objective of this study is to measure the airway diameter, length and bifurcation angles in domestic pigs using computed tomography. After imaging the lungs of 3 pigs, segmentation software tools were used to extract the geometry of the airway lumen. The airway dimensions were then measured from the resulting 3 D models for the first 10 airway generations. Results showed that the size and morphology of the airways of different animals were similar. The measured airway dimensions were compared with those of the human airways. While the trachea diameter was found to be comparable to the adult human, the diameter, length and branching angles of other airways were noticeably different from that of humans. For example, pigs consistently had an early airway branching from the trachea that feeds the superior (top) right lung lobe proximal to the carina. This branch is absent in the human airways. These results suggested that the human geometry may not be a good approximation of the pig airways and may contribute to increasing the errors when the human airway geometric values are used in computational models of the pig chest.

  11. A Yeast Artificial Chromosome Clone Map of the Drosophila Genome

    PubMed Central

    Cai, H.; Kiefel, P.; Yee, J.; Duncan, I.

    1994-01-01

    We describe the mapping of 979 randomly selected large yeast artificial chromosome (YAC) clones of Drosophila DNA by in situ hybridization to polytene chromosomes. Eight hundred and fifty-five of the clones are euchromatic and have primary hybridization sites in the banded portions of the polytene chromosomes, whereas 124 are heterochromatic and label the chromocenter. The average euchromatic clone contains about 211 kb and, at its primary site, labels eight or nine contiguous polytene bands. Thus, the extent as well as chromosomal position of each clone has been determined. By direct band counts, we estimate our clones provide about 76% coverage of the euchromatin of the major autosomes, and 63% coverage of the X. When previously reported YAC mapping data are combined with ours, euchromatic coverage is extended to about 90% for the autosomes and 82% for the X. The distribution of gap sizes in our map and the coverage achieved are in good agreement with expectations based on the assumption of random coverage, indicating that euchromatic clones are essentially randomly distributed. However, certain gaps in coverage, including the entire fourth chromosome euchromatin, may be significant. Heterochromatic sequences are underrepresented among the YAC clones by two to three fold. This may result, at least in part, from underrepresentation of heterochromatic sequences in adult DNA (the source of most of the clones analyzed), or from clone instability. PMID:8013915

  12. Do Managers Clone Themselves?

    ERIC Educational Resources Information Center

    Baron, Alma S.

    1981-01-01

    A recent questionnaire survey provides statistics on male managers' views of female managers. The author recommends that male managers break out of their cloning behavior and that the goal ought to be a plurality in management. (Author/WD)

  13. Statement on Human Cloning

    MedlinePlus

    ... form Search American Association for the Advancement of Science Statement on Human Cloning Print Email Tweet The American Association for the Advancement of Science (AAAS) recognizes the intense debates within our society ...

  14. Twins: A cloning experience.

    PubMed

    Prainsack, Barbara; Spector, Tim D

    2006-11-01

    Drawing upon qualitative interviews with monozygotic (identical) twins sharing 100% of their genes, and with dizygotic (fraternal) twins and singletons as control groups, this paper explores what it means to be genetically identical. (The twins interviewed were from the TwinsUK register in London.) In the context of the ongoing debate on human reproductive cloning, it examines questions such as: To what extent do identical twins perceive their emotional and physical bond to be a result of their genetic makeup? What would they think if they had been deliberately created genetically identical? How would they feel about being genetically identical to a person who was born a few years earlier or later? First, our respondents ascribed no great significance to the role of genes in their understanding of what it means to be identical twins. Second, the opinion that human reproductive cloning would "interfere with nature", or "contradict God's will", was expressed by our respondents exclusively on the abstract level. The more our respondents were able to relate a particular invented cloning scenario to their own life-worlds, the lower the prevalence of the argument. Third, for all three groups of respondents, the scenario of having been born in one of the other groups was perceived as strange. Fourth, the aspect that our respondents disliked about cloning scenarios was the potential motives of the cloners. Without equating monozygotic twins directly with "clones", these results from "naturally" genetically identical individuals add a new dimension to what a future cloning situation could entail: The cloned person might possibly (a) perceive a close physical and emotional connection to the progenitor as a blessing; (b) suffer from preconceptions of people who regard physical likeness as a sign of incomplete individuality; and (c) perceive the idea of not having been born a clone of a particular person as unpleasant.

  15. Senecavirus A: An Emerging Vesicular Infection in Brazilian Pig Herds.

    PubMed

    Leme, R A; Zotti, E; Alcântara, B K; Oliveira, M V; Freitas, L A; Alfieri, A F; Alfieri, A A

    2015-12-01

    Vesicular diseases are clinically and economically important infections that affect farm animals. North American studies have suggested that Senecavirus A infection might be associated with a vesicular disease in pigs known as porcine idiopathic vesicular disease (PIVD). In the beginning of 2015, outbreaks of porcine vesicular disease have occurred in six Brazilian states from three geographical regions. Official diagnostic tests were performed with negative results for classical vesicular diseases of compulsory reporting. This study investigated Senecavirus A infection in PIVD outbreaks in which other aetiological agents were ruled out. A primer set was designed to amplify a 542-bp product size of VP3/VP1 region of Senecavirus A genome in RT-PCR assay. Primer specificity was analysed in silico and in porcine biological specimens. For this, clinical specimens were collected from eight pig herds affected with PIVD, including vesicular fluid (n = 4) and swabs (n = 7) and scrapings of ruptured vesicles and ulcerative lesions (n = 5) from weaned and adult pigs. Clinically healthy animals (n = 52) of PIVD-affected and non-affected pig herds also were evaluated for Senecavirus A infection. The 16 samples from PIVD-affected animals were positive for Senecavirus A in the RT-PCR assay, while none of the clinically healthy pigs were detected with the virus. Sequencing analysis revealed high nucleotide (87.6-98.5%) and amino acid (95-99.4%) similarities to SVV-01 prototype and other Senecavirus A strains from North American pigs. Primer set presented herein was suitable for molecular characterization of Senecavirus A. The results suggest that Senecavirus A was the aetiological agent of the vesicular disease outbreaks in the evaluated pig herds. This is the first study to report the Senecavirus A infection in clinically affected pigs outside of North America. Senecavirus A was considered a novel emerging pathogen associated with an important vesicular disease in Brazil

  16. A Simple "Pig" Game

    ERIC Educational Resources Information Center

    Johnson, Roger W.

    2008-01-01

    Our pig game involves a series of tosses of a die with the possibility of a player's score improving with each additional toss. With each additional toss, however, there is also the chance of losing the entire score accumulated so far. Two different strategies for deciding how many tosses a player should attempt are developed and then compared in…

  17. St. Paul's Pig Pack.

    ERIC Educational Resources Information Center

    Miller, Penny Folley

    1982-01-01

    Describes a guinea pig (cavy) breeding and management program developed as part of an elementary school science curriculum. Includes comments on show competitions (sponsored by the American Rabbit Breeders Association) to measure the success of the breeding program and to enable children to experience the business world. (Author/JN)

  18. Cryptosporidium oocysts and Balantidium coli cysts in pigs reared semi-intensively in Zuru, Nigeria.

    PubMed

    Yatswako, S; Faleke, O O; Gulumbe, M L; Daneji, A I

    2007-10-01

    A cross-sectional study was conducted on the presence of Cryptosporidium oocysts and Balantidium coli cysts in fecal samples from semi-intensively managed pigs in Zuru Local Government Area of Kebbi State, Nigeria between November 2005 and July, 2006. A total of 632 households with human population of 5905 were identified in seven pig- rearing locations in the study area while 105 (16.6%) of these households with human population of 1105 (18.7%) reared a total number of 3895 pigs. Physical randomization was used to select 50% representative samples of pig-rearing households and pigs for this study. Out of the 402 pigs from 55 households, 207 (51.5%) pigs were positive for Balantidium coli cysts, 56 (13.9%) for Cryptosporidium oocysts while mixed infection was observed in 29 (7.2%). There is significance difference in the distribution of the two parasites in the pigs surveyed (p < 0.05). Young pigs were most affected with infection rates of 33 (58.9%) Cryptosporidium oocysts, 113 (54.6%) Balantidium coli cysts and 17(58.6%) mix infection. Human fecal samples collected from 53 individuals revealed 3 (5.7%) positive cases of Cryptosporidium oocysts all in young ones while a positive case of Balantidium coli cyst infection was found in an adult female. Water and soil samples from two areas were also found to contain both organisms. The semi-intensive system of pig rearing which allowed pigs to scavenge and defecate about, defective personal and environmental hygiene couple with the usage of untreated pig feaces as manure on vegetable farms in the study area can enhance the spread of these zoonotic diseases in human population. PMID:19090165

  19. RAG1/2 knockout pigs with severe combined immunodeficiency.

    PubMed

    Huang, Jiao; Guo, Xiaogang; Fan, Nana; Song, Jun; Zhao, Bentian; Ouyang, Zhen; Liu, Zhaoming; Zhao, Yu; Yan, Quanmei; Yi, Xiaoling; Schambach, Axel; Frampton, Jon; Esteban, Miguel A; Yang, Dongshan; Yang, Huaqiang; Lai, Liangxue

    2014-08-01

    Pigs share many physiological, biochemical, and anatomical similarities with humans and have emerged as valuable large animal models for biomedical research. Considering the advantages in immune system resemblance, suitable size, and longevity for clinical practical and monitoring purpose, SCID pigs bearing dysfunctional RAG could serve as important experimental tools for regenerative medicine, allograft and xenograft transplantation, and reconstitution experiments related to the immune system. In this study, we report the generation and phenotypic characterization of RAG1 and RAG2 knockout pigs using transcription activator-like effector nucleases. Porcine fetal fibroblasts were genetically engineered using transcription activator-like effector nucleases and then used to provide donor nuclei for somatic cell nuclear transfer. We obtained 27 live cloned piglets; among these piglets, 9 were targeted with biallelic mutations in RAG1, 3 were targeted with biallelic mutations in RAG2, and 10 were targeted with a monoallelic mutation in RAG2. Piglets with biallelic mutations in either RAG1 or RAG2 exhibited hypoplasia of immune organs, failed to perform V(D)J rearrangement, and lost mature B and T cells. These immunodeficient RAG1/2 knockout pigs are promising tools for biomedical and translational research. PMID:24973446

  20. Extremal quantum cloning machines

    SciTech Connect

    Chiribella, G.; D'Ariano, G. M.; Perinotti, P.; Cerf, N.J.

    2005-10-15

    We investigate the problem of cloning a set of states that is invariant under the action of an irreducible group representation. We then characterize the cloners that are extremal in the convex set of group covariant cloning machines, among which one can restrict the search for optimal cloners. For a set of states that is invariant under the discrete Weyl-Heisenberg group, we show that all extremal cloners can be unitarily realized using the so-called double-Bell states, whence providing a general proof of the popular ansatz used in the literature for finding optimal cloners in a variety of settings. Our result can also be generalized to continuous-variable optimal cloning in infinite dimensions, where the covariance group is the customary Weyl-Heisenberg group of displacement000.

  1. Mechanical ventilation alone, and in the presence of sepsis, impair protein metabolism in the diaphragm of neonatal pigs

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Mechanical ventilation (MV) impairs diaphragmatic function and diminishes the ability to wean from ventilatory support in adult humans. In normal neonatal pigs, animals that are highly anabolic, endotoxin (LPS) infusion induces sepsis, reduces peripheral skeletal muscle protein synthesis rates, but ...

  2. Successful mouse cloning of an outbred strain by trichostatin A treatment after somatic nuclear transfer.

    PubMed

    Kishigami, Satoshi; Bui, Hong-Thuy; Wakayama, Sayaka; Tokunaga, Kenzo; Van Thuan, Nguyen; Hikichi, Takafusa; Mizutani, Eiji; Ohta, Hiroshi; Suetsugu, Rinako; Sata, Tetsutaro; Wakayama, Teruhiko

    2007-02-01

    Although the somatic cloning technique has been used for numerous applications and basic research of reprogramming in various species, extremely low success rates have plagued this technique for a decade. Further in mice, the "clonable" strains have been limited to mainly hybrid F1 strains such as B6D2F1. Recently, we established a new efficient cloning technique using trichostatin A (TSA) which leads to a 2-5 fold increase in success rates for mouse cloning of B6D2F1 cumulus cells. To further test the validity of this TSA cloning technique, we tried to clone the adult ICR mouse, an outbred strain, which has never been directly cloned before. Only when TSA was used did we obtain both male and female cloned mice from cumulus and fibroblast cells of adult ICR mice with 4-5% success rates, which is comparable to 5-7% of B6D2F1. Thus, the TSA treatment is the first cloning technique to allow us to successfully clone outbred mice, demonstrating that this technique not only improves the success rates of cloning from hybrid strains, but also enables mouse cloning from normally "unclonable" strains.

  3. Profile of new green fluorescent protein transgenic Jinhua pigs as an imaging source

    NASA Astrophysics Data System (ADS)

    Kawarasaki, Tatsuo; Uchiyama, Kazuhiko; Hirao, Atsushi; Azuma, Sadahiro; Otake, Masayoshi; Shibata, Masatoshi; Tsuchiya, Seiko; Enosawa, Shin; Takeuchi, Koichi; Konno, Kenjiro; Hakamata, Yoji; Yoshino, Hiroyuki; Wakai, Takuya; Ookawara, Shigeo; Tanaka, Hozumi; Kobayashi, Eiji; Murakami, Takashi

    2009-09-01

    Animal imaging sources have become an indispensable material for biological sciences. Specifically, gene-encoded biological probes serve as stable and high-performance tools to visualize cellular fate in living animals. We use a somatic cell cloning technique to create new green fluorescent protein (GFP)-expressing Jinhua pigs with a miniature body size, and characterized the expression profile in various tissues/organs and ex vivo culture conditions. The born GFP-transgenic pig demonstrate an organ/tissue-dependent expression pattern. Strong GFP expression is observed in the skeletal muscle, pancreas, heart, and kidney. Regarding cellular levels, bone-marrow-derived mesenchymal stromal cells, hepatocytes, and islet cells of the pancreas also show sufficient expression with the unique pattern. Moreover, the cloned pigs demonstrate normal growth and fertility, and the introduced GFP gene is stably transmitted to pigs in subsequent generations. The new GFP-expressing Jinhua pigs may be used as new cellular/tissue light resources for biological imaging in preclinical research fields such as tissue engineering, experimental regenerative medicine, and transplantation.

  4. Molecular cloning of a gene encoding a Chlamydia psittaci 57-kDa protein that shares antigenic determinants with ca. 60-kDa proteins present in many gram-negative bacteria.

    PubMed

    Menozzi, F D; Menozzi-Dejaiffe, C; Nano, F E

    1989-03-01

    In order to develop reagents to study the immune response of guinea pigs to infection by Chlamydia psittaci guinea pig inclusion conjunctivitis strain (GPIC), we constructed a plasmid clone bank with C. psittaci DNA. One of the recombinant clones isolated produced large amounts of a 57-kilodalton (kDa) protein that was immunoreactive with sera from GPIC infected guinea pigs. While investigating this recombinant protein, we discovered that all the Gram-negative bacteria analyzed so far have immunoreactive proteins of similar size. This protein seems to be a 'common antigen' already described in various Gram-negative bacteria.

  5. Comparison of Cervical Spine Anatomy in Calves, Pigs and Humans

    PubMed Central

    Sheng, Sun-Ren; Xu, Hua-Zi; Wang, Yong-Li; Zhu, Qing-An; Mao, Fang-Min; Lin, Yan; Wang, Xiang-Yang

    2016-01-01

    Background Context Animals are commonly used to model the human spine for in vitro and in vivo experiments. Many studies have investigated similarities and differences between animals and humans in the lumbar and thoracic vertebrae. However, a quantitative anatomic comparison of calf, pig, and human cervical spines has not been reported. Purpose To compare fundamental structural similarities and differences in vertebral bodies from the cervical spines of commonly used experimental animal models and humans. Study Design Anatomical morphometric analysis was performed on cervical vertebra specimens harvested from humans and two common large animals (i.e., calves and pigs). Methods Multiple morphometric parameters were directly measured from cervical spine specimens of twelve pigs, twelve calves and twelve human adult cadavers. The following anatomical parameters were measured: vertebral body width (VBW), vertebral body depth (VBD), vertebral body height (VBH), spinal canal width (SCW), spinal canal depth (SCD), pedicle width (PW), pedicle depth (PD), pedicle inclination (PI), dens width (DW), dens depth (DD), total vertebral width (TVW), and total vertebral depth (TVD). Results The atlantoaxial (C1–2) joint in pigs is similar to that in humans and could serve as a human substitute. The pig cervical spine is highly similar to the human cervical spine, except for two large transverse processes in the anterior regions ofC4–C6. The width and depth of the calf odontoid process were larger than those in humans. VBW and VBD of calf cervical vertebrae were larger than those in humans, but the spinal canal was smaller. Calf C7 was relatively similar to human C7, thus, it may be a good substitute. Conclusion Pig cervical vertebrae were more suitable human substitutions than calf cervical vertebrae, especially with respect to C1, C2, and C7. The biomechanical properties of nerve vascular anatomy and various segment functions in pig and calf cervical vertebrae must be

  6. Functional expression of pig renal organic anion transporter 3 (pOAT3).

    PubMed

    Hagos, Yohannes; Braun, Isabella M; Krick, Wolfgang; Burckhardt, Gerhard; Bahn, Andrew

    2005-05-01

    With the cloning of pig renal organic anion transporter 1 (pOAT1) (Biochimie 84 (2002) 1219) we set up a model system for comparative studies of cloned and natively isolated membrane located transport proteins. Meanwhile, another transport protein involved in p-aminohippurate (PAH) uptake on the basolateral side of the proximal tubule cells was identified, designated organic anion transporter 3 (OAT3). To explore the contribution of pOAT1 to the PAH clearance in comparison to OAT3, it was the aim of this study to extend our model by cloning of the pig ortholog of OAT3. Sequence comparisons of human organic anion transporter 3 (hOAT3) with the expressed sequence tag (EST) database revealed a clone and partial sequence of the pig renal organic anion transporter 3 (pOAT3) ortholog. Sequencing of the entire open reading frame resulted in a protein of 543 amino acid residues encoded by 1632 base pairs (EMBL Acc. No. AJ587003). It showed high homologies of 81%, 80%, 76%, and 77% to the human, rabbit, rat, and mouse OAT3, respectively. A functional characterization of pOAT3 in Xenopus laevis oocytes yielded an apparent Km (Kt) for [3H]estrone sulfate of 7.8 +/- 1.3 microM. Moreover, pOAT3 mediated [3H]estrone sulfate uptake was almost abolished by 0.5 mM of glutarate, dehydroepiandosterone sulfate, or probenecid consistent with the hallmarks of OAT3 function.

  7. Applications of quantum cloning

    NASA Astrophysics Data System (ADS)

    Pomarico, E.; Sanguinetti, B.; Sekatski, P.; Zbinden, H.; Gisin, N.

    2011-10-01

    Quantum Cloning Machines (QCMs) allow for the copying of information, within the limits imposed by quantum mechanics. These devices are particularly interesting in the high-gain regime, i.e., when one input qubit generates a state of many output qubits. In this regime, they allow for the study of certain aspects of the quantum to classical transition. The understanding of these aspects is the root of the two recent applications that we will review in this paper: the first one is the Quantum Cloning Radiometer, a device which is able to produce an absolute measure of spectral radiance. This device exploits the fact that in the quantum regime information can be copied with only finite fidelity, whereas when a state becomes macroscopic, this fidelity gradually increases to 1. Measuring the fidelity of the cloning operation then allows to precisely determine the absolute spectral radiance of the input optical source. We will then discuss whether a Quantum Cloning Machine could be used to produce a state visible by the naked human eye, and the possibility of a Bell Experiment with humans playing the role of detectors.

  8. The Cloning of America.

    ERIC Educational Resources Information Center

    Dobson, Judith E.; Dobson, Russell L.

    1981-01-01

    Proposes that the U.S. school system purports to prize human variability, but many educators are engaged in activities that seek to homogenize students. Describes these activities, including diagnosis, labeling, ability grouping, and positive reinforcement. Presents suggestions for counselors to combat sources of cloning and self-validation. (RC)

  9. Human therapeutic cloning.

    PubMed

    Lanza, R P; Cibelli, J B; West, M D

    1999-09-01

    Somatic cell nuclear 'reprogramming' in livestock species is now routine in many laboratories. Here, Robert Lanza, Jose Cibelli and Michael West discuss how these techniques may soon be used to clone genetically matched cells and tissues for transplantation into patients suffering from a wide range of disorders that result from tissue loss or dysfunction.

  10. The First Human Cloned Embryo.

    ERIC Educational Resources Information Center

    Cibelli, Jose B.; Lanza, Robert P.; West, Michael D.; Ezzell, Carol

    2002-01-01

    Describes a process known as parthenogenesis which produces cloned, early-stage embryos and human embryos generated only from eggs. Speculates that this technology puts therapeutic cloning within reach. (DDR)

  11. Sequential cloning of chromosomes

    SciTech Connect

    Lacks, S.A.

    1991-12-31

    A method for sequential cloning of chromosomal DNA and chromosomal DNA cloned by this method are disclosed. The method includes the selection of a target organism having a segment of chromosomal DNA to be sequentially cloned. A first DNA segment, having a first restriction enzyme site on either side. homologous to the chromosomal DNA to be sequentially cloned is isolated. A first vector product is formed by ligating the homologous segment into a suitably designed vector. The first vector product is circularly integrated into the target organism`s chromosomal DNA. The resulting integrated chromosomal DNA segment includes the homologous DNA segment at either end of the integrated vector segment. The integrated chromosomal DNA is cleaved with a second restriction enzyme and ligated to form a vector-containing plasmid, which is replicated in a host organism. The replicated plasmid is then cleaved with the first restriction enzyme. Next, a DNA segment containing the vector and a segment of DNA homologous to a distal portion of the previously isolated DNA segment is isolated. This segment is then ligated to form a plasmid which is replicated within a suitable host. This plasmid is then circularly integrated into the target chromosomal DNA. The chromosomal DNA containing the circularly integrated vector is treated with a third, retrorestriction enzyme. The cleaved DNA is ligated to give a plasmid that is used to transform a host permissive for replication of its vector. The sequential cloning process continues by repeated cycles of circular integration and excision. The excision is carried out alternately with the second and third enzymes.

  12. Probabilistic Cloning and Quantum Computation

    NASA Astrophysics Data System (ADS)

    Gao, Ting; Yan, Feng-Li; Wang, Zhi-Xi

    2004-06-01

    We discuss the usefulness of quantum cloning and present examples of quantum computation tasks for which the cloning offers an advantage which cannot be matched by any approach that does not resort to quantum cloning. In these quantum computations, we need to distribute quantum information contained in the states about which we have some partial information. To perform quantum computations, we use a state-dependent probabilistic quantum cloning procedure to distribute quantum information in the middle of a quantum computation.

  13. [Expression analysis of green fluorescent protein in tissues and organs in α-1,3 galactosyltransferase knockout pigs].

    PubMed

    Zhifang, Li; Chong, Feng; Huili, Ji; Ningning, Shi; Xiaofeng, Song; Qinli, Zhao; Chuan, Long; Dengke, Pan; Xiaogan, Yang

    2015-12-01

    The pig is an ideal source to provide organs because its organ size and physiology are similar to humans. However, an acute rejection will ensue after pig-to-human xenotransplantation. The α-1,3 galactosyltransferase gene knockout (GTKO) pigs were generated in recent years, and could solve the problem of hyperacute rejection. But due to lack of reporting genes, the rejection status of cells and organs post pig-to-human xenotransplantation cannot be visualized. In this study, we introduced the enhanced green fluorescent protein (EGFP) gene driven by the CAG promoter into GTKO porcine ear fibroblasts. Then we produced transgenic pigs expressing the EGFP gene by nuclear transfer technology. Expression levels of EGFP in different tissues and organs of the cloned pig were investigated by Nightsea DFP-1 Fluorescent Protein Flashlight, fluorescence microscope and quantitative PCR assays. The results showed that the protein and transcript of EGFP were expressed in all tissues and organs of the GTKO pig, but the expression was weak in the liver and central nervous system. In conclusion, we have successfully produced the transgenic GTKO pigs expressing EGFP in all tested tissues and organs, which builds up a good basis to track transplanted cells or tissues.

  14. Establishment of a novel, eco-friendly transgenic pig model using porcine pancreatic amylase promoter-driven fungal cellulase transgenes.

    PubMed

    Lin, Y S; Yang, C C; Hsu, C C; Hsu, J T; Wu, S C; Lin, C J; Cheng, W T K

    2015-02-01

    Competition between humans and livestock for cereal and legume grains makes it challenging to provide economical feeds to livestock animals. Recent increases in corn and soybean prices have had a significant impact on the cost of feed for pig producers. The utilization of byproducts and alternative ingredients in pig diets has the potential to reduce feed costs. Moreover, unlike ruminants, pigs have limited ability to utilize diets with high fiber content because they lack endogenous enzymes capable of breaking down nonstarch polysaccharides into simple sugars. Here, we investigated the feasibility of a transgenic strategy in which expression of the fungal cellulase transgene was driven by the porcine pancreatic amylase promoter in pigs. A 2,488 bp 5'-flanking region of the porcine pancreatic amylase gene was cloned by the genomic walking technique, and its structural features were characterized. Using GFP as a reporter, we found that this region contained promoter activity and had the potential to control heterologous gene expression. Transgenic pigs were generated by pronuclear microinjection. Founders and offspring were identified by PCR and Southern blot analyses. Cellulase mRNA and protein showed tissue-specific expression in the pancreas of F1 generation pigs. Cellulolytic enzyme activity was also identified in the pancreas of transgenic pigs. These results demonstrated the establishment of a tissue-specific promoter of the porcine pancreatic amylase gene. Transgenic pigs expressing exogenous cellulase may represent a way to increase the intake of low-cost, fiber-rich feeds.

  15. [Expression analysis of green fluorescent protein in tissues and organs in α-1,3 galactosyltransferase knockout pigs].

    PubMed

    Zhifang, Li; Chong, Feng; Huili, Ji; Ningning, Shi; Xiaofeng, Song; Qinli, Zhao; Chuan, Long; Dengke, Pan; Xiaogan, Yang

    2015-12-01

    The pig is an ideal source to provide organs because its organ size and physiology are similar to humans. However, an acute rejection will ensue after pig-to-human xenotransplantation. The α-1,3 galactosyltransferase gene knockout (GTKO) pigs were generated in recent years, and could solve the problem of hyperacute rejection. But due to lack of reporting genes, the rejection status of cells and organs post pig-to-human xenotransplantation cannot be visualized. In this study, we introduced the enhanced green fluorescent protein (EGFP) gene driven by the CAG promoter into GTKO porcine ear fibroblasts. Then we produced transgenic pigs expressing the EGFP gene by nuclear transfer technology. Expression levels of EGFP in different tissues and organs of the cloned pig were investigated by Nightsea DFP-1 Fluorescent Protein Flashlight, fluorescence microscope and quantitative PCR assays. The results showed that the protein and transcript of EGFP were expressed in all tissues and organs of the GTKO pig, but the expression was weak in the liver and central nervous system. In conclusion, we have successfully produced the transgenic GTKO pigs expressing EGFP in all tested tissues and organs, which builds up a good basis to track transplanted cells or tissues. PMID:26704946

  16. Recombinational Cloning Using Gateway and In-Fusion Cloning Schemes

    PubMed Central

    Throop, Andrea L.; LaBaer, Joshua

    2015-01-01

    The comprehensive study of protein structure and function, or proteomics, depends on the obtainability of full-length cDNAs in species-specific expression vectors and subsequent functional analysis of the expressed protein. Recombinational cloning is a universal cloning technique based on site-specific recombination that is independent of the insert DNA sequence of interest, which differentiates this method from the classical restriction enzyme-based cloning methods. Recombinational cloning enables rapid and efficient parallel transfer of DNA inserts into multiple expression systems. This unit summarizes strategies for generating expression-ready clones using the most popular recombinational cloning technologies, including the commercially available Gateway® (Life Technologies) and In-Fusion® (Clontech) cloning technologies. PMID:25827088

  17. Technology And Pregnant Pigs

    NASA Technical Reports Server (NTRS)

    1978-01-01

    One of the interesting things about aerospace spinoff is the way it keeps cropping up in uncommon applications unimaginably remote from the original technology. For example, the pig pregnancy detector. The pig pregnancy detector? City folk may be surprised to learn that there is such a thing-and wonder why. The why is because it is a sow's job to produce piglets and farmers can't afford to keep those who don't; it costs about a half-dollar a day in feed, labor and facilities, and even in small herds that's intolerable. So the barren sow must go. Until recently, the best method of determining pig pregnancy was "eyeballing," daily visual examination over a period of time. The problem with eyeballing is that pregnancy is not evident until well advanced; when there is no pregnancy, the farmer learns too late that he has been feeding a sow that won't give him a litter. Advancing technology provided an answer: the quick, easy-to-use, accurate automatic detector for early evaluation of pregnancy status. Among the most popular of these devices are Scanopreg and Scanoprobe, to whose development NASA technology contributed. Scanopreg is an ultrasonic system which detects pregnancy about 30 days after breeding, long before eyeballing can provide an answer. The companion Scanoprobe is a dual-function unit which not only determines pregnancy but also gives farmers an analysis of a hog's meat-fat ratio, an important factor in breeding. Only a short time on the market, Scanopreg and Scanoprobe have already found wide acceptance among meat producers because they rapidly repay their cost.

  18. 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

  19. 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.

  20. A review of Greek law on human cloning.

    PubMed

    Mavroforou, Anna; Giannoukas, Athanasios; Michalodimitrakis, Emmanuel

    2003-01-01

    The creation of Dolly, a cloned lamb from adult cells was a major scientific breakthrough, which opened new avenues for many research fields such as reproductive medicine, transplantation and biotechnology. However this achievement brought to public attention the theoretical possibility of human reproductive cloning. Inevitably heated debate occurred on several ethical and legal consequences of the prospect of human cloning. At the present time there is no legal framework in any country to respond to this challenge in a pragmatic way in order to protect human rights and at the same time to allow science to work for the best interests of mankind. Greece is a European Union country with its own traditions, history, culture and beliefs but without political and legislative experience in the handling of medical and biotechnological matters. This paper aims to discuss the legal issues likely to be raised by the prospect of human reproductive cloning in relation to the current state of the Greek legal system.

  1. A review of Greek law on human cloning.

    PubMed

    Mavroforou, Anna; Giannoukas, Athanasios; Michalodimitrakis, Emmanuel

    2003-01-01

    The creation of Dolly, a cloned lamb from adult cells was a major scientific breakthrough, which opened new avenues for many research fields such as reproductive medicine, transplantation and biotechnology. However this achievement brought to public attention the theoretical possibility of human reproductive cloning. Inevitably heated debate occurred on several ethical and legal consequences of the prospect of human cloning. At the present time there is no legal framework in any country to respond to this challenge in a pragmatic way in order to protect human rights and at the same time to allow science to work for the best interests of mankind. Greece is a European Union country with its own traditions, history, culture and beliefs but without political and legislative experience in the handling of medical and biotechnological matters. This paper aims to discuss the legal issues likely to be raised by the prospect of human reproductive cloning in relation to the current state of the Greek legal system. PMID:12809342

  2. Production of alpha 1,3-galactosyltransferase gene knockout pigs expressing both human decay-accelerating factor and N-acetylglucosaminyltransferase III.

    PubMed

    Takahagi, Yoichi; Fujimura, Tatsuya; Miyagawa, Shuji; Nagashima, Hiroshi; Shigehisa, Tamotsu; Shirakura, Ryota; Murakami, Hiroshi

    2005-07-01

    Heterozygous alpha 1,3-galactosyltransferase (GT) gene knockout pigs were produced with transgenic pig fetal cells expressing both human decay-accelerating factor (hDAF) and N-acetylglucosaminyltransferase III (GnT-III). In this study, we assessed the gene targeting efficiency in the transgenic pig fetal cells derived from different fetal tissues such as brain, skin, heart, and liver, or fetal carcass. Targeted cell colonies were selected by hygromycin B. The GT-knockout colonies (KO colonies) were obtained equally from the cells derived from all tissues except liver. Staining with five antibodies against intermediate filaments, all examined KO cell lines stained positive for vimentin with the exception of a colony that stained positive for both vimentin and glial fibrillary acidic protein simultaneously. This is the first study to produce KO cells from the astrocytes. Some of these KO cell lines were used for nuclear transfer (NT) to obtain KO pig fetuses. Fourteen fetuses were obtained from two recipients of the embryo transfer and eight of them had normal ploidy. The cells from the KO pig fetuses were also used for NT to produce cloned KO pigs. Two healthy clone pigs were born. These pigs were determined to have a heterozygous knockout GT gene and the two transgenes. The cells collected from the KO pigs were shown to have similar expression levels of hDAF and GnT-III compared to their original transgenic pigs and less than a half levels of the alphaGal epitopes existed in wild-type pig cells.

  3. Further signs of postnatal sexual differentiation in pigs.

    PubMed

    Ventanas, J; López-Bote, C J; García, C

    1992-01-01

    Fourteen newborn female, and ten male pigs were injected subcutaneously with 100 mg of testosterone propionate within 24 h of birth, and ten male and nine females were used as controls and injected with vehicle alone (Sesame oil). All animals were slaughtered at 175 days of age. Reduction of testicular weight, tubuli size and testosterone concentration, together with alteration of the intertubular area was found in adult treated males. In adult treated females, the histological study of ovaries showed the parenchyma fully occupied by multiple Graaf follicles, but no corpus luteum was found. In addition, treated females never had serum progesterone levels above 2 ng/ml, whilst control females, in some cases had values close to 2 ng/ml, and in some others near to 10 ng/ml. Carcass weight and fatness were not markedly different between groups. The observations presented in this report provide further support for a neonatal component of testosterone-dependent masculinization process in pigs.

  4. The Pig--Pet, Pork or Sacrifice?

    ERIC Educational Resources Information Center

    Arnold, Arthur

    1988-01-01

    Discusses the various roles of the pig in children's books, including E. B. White's CHARLOTTE'S WEB and Nina Bawden's PEPPERMINT PIG. Notes that, although pigs are often used as metaphors for greed, gluttony, and squalor, the portrayal of pigs in children's literature is typically positive. (MM)

  5. [Media, cloning, and bioethics].

    PubMed

    Costa, S I; Diniz, D

    2000-01-01

    This article was based on an analysis of three hundred articles from mainstream Brazilian periodicals over a period of eighteen months, beginning with the announcement of the Dolly case in February 1997. There were two main objectives: to outline the moral constants in the press associated with the possibility of cloning human beings and to identify some of the moral assumptions concerning scientific research with non-human animals that were published carelessly by the media. The authors conclude that there was a haphazard spread of fear concerning the cloning of human beings rather than an ethical debate on the issue, and that there is a serious gap between bioethical reflections and the Brazilian media.

  6. Overlap extension PCR cloning.

    PubMed

    Bryksin, Anton; Matsumura, Ichiro

    2013-01-01

    Rising demand for recombinant proteins has motivated the development of efficient and reliable cloning methods. Here we show how a beginner can clone virtually any DNA insert into a plasmid of choice without the use of restriction endonucleases or T4 DNA ligase. Chimeric primers encoding plasmid sequence at the 5' ends and insert sequence at the 3' ends are designed and synthesized. Phusion(®) DNA polymerase is utilized to amplify the desired insert by PCR. The double-stranded product is subsequently employed as a pair of mega-primers in a PCR-like reaction with circular plasmids. The original plasmids are then destroyed in restriction digests with Dpn I. The product of the overlap extension PCR is used to transform competent Escherichia coli cells. Phusion(®) DNA polymerase is used for both the amplification and fusion reactions, so both steps can be monitored and optimized in the same way. PMID:23996437

  7. Oxamflatin Treatment Enhances Cloned Porcine Embryo Development and Nuclear Reprogramming*

    PubMed Central

    Mao, Jiude; Zhao, Ming-Tao; Whitworth, Kristin M.; Spate, Lee D.; Walters, Eric M.; O'Gorman, Chad; Lee, Kiho; Samuel, Melissa S.; Murphy, Clifton N.; Wells, Kevin; Rivera, Rocio M.

    2015-01-01

    Abstract Faulty epigenetic reprogramming of somatic nuclei is thought to be the main reason for low cloning efficiency by somatic cell nuclear transfer (SCNT). Histone deacetylase inhibitors (HDACi), such as Scriptaid, improve developmental competence of SCNT embryos in several species. Another HDACi, Oxamflatin, is about 100 times more potent than Scriptaid in the ability to inhibit nuclear-specific HDACs. The present study determined the effects of Oxamflatin treatment on embryo development, DNA methylation, and gene expression. Oxamflatin treatment enhanced blastocyst formation of SCNT embryos in vitro. Embryo transfer produced more pigs born and fewer mummies from the Oxamflatin-treated group compared to the Scriptaid-treated positive control. Oxamflatin also decreased DNA methylation of POU5F1 regulatory elements and centromeric repeat elements in day-7 blastocysts. When compared to in vitro–fertilized (IVF) embryos, the methylation status of POU5F1, NANOG, and centromeric repeat was similar in the cloned embryos, indicating these genes were successfully reprogrammed. However, compared to the lack of methylation of XIST in day-7 IVF embryos, a higher methylation level in day-7 cloned embryos was observed, implying that X chromosomes were activated in day-7 IVF blastocysts, but were not fully activated in cloned embryos, i.e., reprogramming of XIST was delayed. A time-course analysis of XIST DNA methylation on day-13, -15, -17, and -19 in vivo embryos revealed that XIST methylation initiated at about day 13 and was not completed by day 19. The methylation of the XIST gene in day-19 control cloned embryos was delayed again when compared to in vivo embryos. However, methylation of XIST in Oxamflatin-treated embryos was comparable with in vivo embryos, which further demonstrated that Oxamflatin could accelerate the delayed reprogramming of XIST gene and thus might improve cloning efficiency. PMID:25548976

  8. Probabilistic cloning of equidistant states

    SciTech Connect

    Jimenez, O.; Roa, Luis; Delgado, A.

    2010-08-15

    We study the probabilistic cloning of equidistant states. These states are such that the inner product between them is a complex constant or its conjugate. Thereby, it is possible to study their cloning in a simple way. In particular, we are interested in the behavior of the cloning probability as a function of the phase of the overlap among the involved states. We show that for certain families of equidistant states Duan and Guo's cloning machine leads to cloning probabilities lower than the optimal unambiguous discrimination probability of equidistant states. We propose an alternative cloning machine whose cloning probability is higher than or equal to the optimal unambiguous discrimination probability for any family of equidistant states. Both machines achieve the same probability for equidistant states whose inner product is a positive real number.

  9. Ethical issues in livestock cloning.

    PubMed

    Thompson, P B

    1999-01-01

    Although cloning may eventually become an important technology for livestock production, four ethical issues must be addressed before the practice becomes widespread. First, researchers must establish that the procedure is not detrimental to the health or well-being of affected animals. Second, animal research institutions should evaluate the net social benefits to livestock producers by weighing the benefits to producers against the opportunity cost of research capacity lost to biomedical projects. Third, scientists should consider the indirect effects of cloning research on the larger ethical issues surrounding human cloning. Finally, the market structure for products of cloned animals should protect individual choice, and should recognize that many individuals find the prospect of cloning (or consuming cloned animals) repugnant. Analysis of these four issues is complicated by spurious arguments alleging that cloning will have a negative impact on environment and genetic diversity.

  10. Ethical issues in livestock cloning.

    PubMed

    Thompson, P B

    1999-01-01

    Although cloning may eventually become an important technology for livestock production, four ethical issues must be addressed before the practice becomes widespread. First, researchers must establish that the procedure is not detrimental to the health or well-being of affected animals. Second, animal research institutions should evaluate the net social benefits to livestock producers by weighing the benefits to producers against the opportunity cost of research capacity lost to biomedical projects. Third, scientists should consider the indirect effects of cloning research on the larger ethical issues surrounding human cloning. Finally, the market structure for products of cloned animals should protect individual choice, and should recognize that many individuals find the prospect of cloning (or consuming cloned animals) repugnant. Analysis of these four issues is complicated by spurious arguments alleging that cloning will have a negative impact on environment and genetic diversity. PMID:15719505

  11. Sequential cloning of chromosomes

    DOEpatents

    Lacks, Sanford A.

    1995-07-18

    A method for sequential cloning of chromosomal DNA of a target organism is disclosed. A first DNA segment homologous to the chromosomal DNA to be sequentially cloned is isolated. The first segment has a first restriction enzyme site on either side. A first vector product is formed by ligating the homologous segment into a suitably designed vector. The first vector product is circularly integrated into the target organism's chromosomal DNA. The resulting integrated chromosomal DNA segment includes the homologous DNA segment at either end of the integrated vector segment. The integrated chromosomal DNA is cleaved with a second restriction enzyme and ligated to form a vector-containing plasmid, which is replicated in a host organism. The replicated plasmid is then cleaved with the first restriction enzyme. Next, a DNA segment containing the vector and a segment of DNA homologous to a distal portion of the previously isolated DNA segment is isolated. This segment is then ligated to form a plasmid which is replicated within a suitable host. This plasmid is then circularly integrated into the target chromosomal DNA. The chromosomal DNA containing the circularly integrated vector is treated with a third, retrorestriction (class IIS) enzyme. The cleaved DNA is ligated to give a plasmid that is used to transform a host permissive for replication of its vector. The sequential cloning process continues by repeated cycles of circular integration and excision. The excision is carried out alternately with the second and third enzymes.

  12. Sequential cloning of chromosomes

    DOEpatents

    Lacks, S.A.

    1995-07-18

    A method for sequential cloning of chromosomal DNA of a target organism is disclosed. A first DNA segment homologous to the chromosomal DNA to be sequentially cloned is isolated. The first segment has a first restriction enzyme site on either side. A first vector product is formed by ligating the homologous segment into a suitably designed vector. The first vector product is circularly integrated into the target organism`s chromosomal DNA. The resulting integrated chromosomal DNA segment includes the homologous DNA segment at either end of the integrated vector segment. The integrated chromosomal DNA is cleaved with a second restriction enzyme and ligated to form a vector-containing plasmid, which is replicated in a host organism. The replicated plasmid is then cleaved with the first restriction enzyme. Next, a DNA segment containing the vector and a segment of DNA homologous to a distal portion of the previously isolated DNA segment is isolated. This segment is then ligated to form a plasmid which is replicated within a suitable host. This plasmid is then circularly integrated into the target chromosomal DNA. The chromosomal DNA containing the circularly integrated vector is treated with a third, retrorestriction (class IIS) enzyme. The cleaved DNA is ligated to give a plasmid that is used to transform a host permissive for replication of its vector. The sequential cloning process continues by repeated cycles of circular integration and excision. The excision is carried out alternately with the second and third enzymes. 9 figs.

  13. Cloning-free CRISPR

    PubMed Central

    Arbab, Mandana; Srinivasan, Sharanya; Hashimoto, Tatsunori; Geijsen, Niels; Sherwood, Richard I.

    2015-01-01

    Summary We present self-cloning CRISPR/Cas9 (scCRISPR), a technology that allows for CRISPR/Cas9-mediated genomic mutation and site-specific knockin transgene creation within several hours by circumventing the need to clone a site-specific single-guide RNA (sgRNA) or knockin homology construct for each target locus. We introduce a self-cleaving palindromic sgRNA plasmid and a short double-stranded DNA sequence encoding the desired locus-specific sgRNA into target cells, allowing them to produce a locus-specific sgRNA plasmid through homologous recombination. scCRISPR enables efficient generation of gene knockouts (∼88% mutation rate) at approximately one-sixth the cost of plasmid-based sgRNA construction with only 2 hr of preparation for each targeted site. Additionally, we demonstrate efficient site-specific knockin of GFP transgenes without any plasmid cloning or genome-integrated selection cassette in mouse and human embryonic stem cells (2%–4% knockin rate) through PCR-based addition of short homology arms. scCRISPR substantially lowers the bar on mouse and human transgenesis. PMID:26527385

  14. Progression of Mycoplasma hyosynoviae infection in three pig herds. Development of tonsillar carrier state, arthritis and antibodies in serum and synovial fluid in pigs from birth to slaughter.

    PubMed

    Hagedorn-Olsen, T; Nielsen, N C; Friis, N F; Nielsen, J

    1999-11-01

    In this investigation, natural infection with Mycoplasma hyosynoviae was followed in groups of individual pigs in three different herds with regard to occurrence of tonsillar carrier state, clinical arthritis and development of antibodies in serum and in synovial fluid. Antibodies were detected by a polyclonal enzyme-linked immunosorbent assay (ELISA) developed for experimental use. The infection with M. hyosynoviae progressed very differently in the three herds investigated. In one herd, the infection was apparently limited to adult pigs. In a second herd, all pigs became tonsillar carriers of M. hyosynoviae, but no mycoplasma-related arthritis nor any serological response was demonstrated within the growing-finishing period. In the third herd investigated, tonsillar infection was detected in all pigs, clinical cases of M. hyosynoviae arthritis followed and a moderate serological response was observed in some, but not all, pigs. In all three herds, M. hyosynoviae infection was carried in the tonsils of the adult pigs, but it was only occasionally transmitted from sows to piglets. Maternal antibodies were transferred to the piglets and persisted for approximately 8-12 weeks. After weaning, some pigs became infected before 20 weeks of age, while others did not. In the majority of cases, the tonsillar infection was established from 11 weeks of age or older. A latent tonsillar infection was present for a period of several weeks within the group of investigated pigs before cases of generalized infection and arthritis were seen. In some cases, generalization of M. hyosynoviae infection in the blood and in joints was observed in spite of the detection of an active serological response a few weeks earlier. The present work suggests that generalization of the infection and development of arthritis may depend on age, immunity, virulence factors and/or infection pressure; in some herds maybe combined with certain triggering mechanisms such as stress and lowered general

  15. Cloning and characterization of a cDNA clone encoding calreticulin from Haemaphysalis qinghaiensis (Acari: Ixodidae).

    PubMed

    Gao, Jinliang; Luo, Jianxun; Fan, Ruiquan; Fingerle, Volker; Guan, Guiquan; Liu, Zhijie; Li, Youquan; Zhao, Haiping; Ma, Miling; Liu, Junlong; Liu, Aihong; Ren, Qiaoyun; Dang, Zhisheng; Sugimoto, Chihiro; Yin, Hong

    2008-03-01

    The application of anti-tick vaccine has been shown to be the most promising alternative strategy compared to the current use of acaricides that suffer from a number of serious limitations. The success of this method is dependent upon identification and cloning of potential tick vaccine antigens. Previously, we have cloned 21 positive clones (named from Hq02 to Hq22) by immunoscreening complimentary DNA (cDNA) libraries of Haemaphysalis qinghaiensis; however, some of those clones did not contain open reading frames (ORF). In this study, we amplified the entire sequence of Hq07 by using rapid amplification of the cDNA ends. Hq07 contains an ORF of 1,233 bp that encodes for 410 amino acid residues with a coding capacity of 47 kDa. Search of the cloned sequences against GenBank revealed that Hq07 is a calreticulin (CRT)-similar clone and designated HqCRT. Expression analysis by reverse transcription-polymerase chain reaction showed that this gene is ubiquitously expressed at different developmental stages and in different tissues of H. qinghaiensis. The gene was expressed as glutathione S-transferase-fused proteins in a prokaryotic system. Western blot analysis revealed that native HqCRT was secreted into their hosts by ticks during blood sucking. Vaccination of sheep with rHqCRT conferred protective immunity against ticks, resulting in 54.3% mortality in adult ticks, compared to the 38.7% death rate in the control group. These results demonstrated that rHqCRT might be a useful vaccine candidate antigen for biological control of H. qinghaiensis.

  16. Nucleus transfer efficiency of ear fibroblast cells isolated from Bama miniature pigs at various ages.

    PubMed

    Wang, Qing-Hua; Peng, Yun; Cai, Xin-Yong; Wan, Meng; Liu, Yu; Wei, Hong

    2015-08-01

    Somatic cell nucleus transfer (SCNT) has been considered the most effective method for conserving endangered animals and expanding the quantity of adult animal models. Bama miniature pigs are genetically stable and share similar biological features to humans. These pigs have been used to establish animal models for human diseases, and for many other applications. However, there is a paucity of studies on the effect of ear fibroblasts derived from different age of adult Bama miniature pigs on nucleus transfer (NT). The present study examined the NT efficiency of ear fibroblasts from fetal, newborn, 1-, 2-, 4-, 6-, 12-month-old miniature pigs by using trypan blue staining, flow cytometry and NT technique, etc., and the cell biological function and SCNT efficiency were compared between groups. The results showed that ear fibroblasts grew well after passage in each group. Spindle-shaped cells initially predominated, and gradually declined with increase of culture time and replaced by polygonal cells. Irregular cell growth occurred in the 2-month-old group and the elder groups. The growth curves of the ear fibroblasts were "S-shaped" in different age groups. The cell proliferation of postnatal ear fibroblasts, especially those from 2-, 4-, 6-, 12-month-old miniature pigs was significantly different from that of fetus ear fibroblasts (P<0.05 or P<0.01). Two-month- and 4-month-old ear fibroblasts had a significantly higher proportion of G1 stage cells (85% to 91%) than those at 6 and 12 months (66% to 74%, P<0.01). The blastocyst rate of reconstructed embryos originating from newborn, 1-, 2-, 4-month-old donor pigs was 6.06% to 7.69% with no significant difference from that in fetus fibroblast group (8.06%). It was concluded that <4-month-old adult Bama miniature pigs represent a better donor cell resource than elder pigs.

  17. Universal cytotoxic activity of a HTLV-1 Tax-specific T cell clone from an HLA-A*24:02⁺ patient with adult T-cell leukemia against a variety of HTLV-I-infected T-cells.

    PubMed

    Tanaka, Yukie; Yamazaki, Rie; Terasako-Saito, Kiriko; Nakasone, Hideki; Akahoshi, Yu; Nakano, Hirofumi; Ugai, Tomotaka; Wada, Hidenori; Yamasaki, Ryoko; Ishihara, Yuko; Kawamura, Koji; Sakamoto, Kana; Ashizawa, Masahiro; Sato, Miki; Kimura, Shun-ichi; Kikuchi, Misato; Kako, Shinichi; Kanda, Junya; Tanihara, Aki; Nishida, Junji; Kanda, Yoshinobu

    2014-01-01

    Adult T cell leukemia/lymphoma (ATL) is an aggressive mature T cell malignancy that is causally associated with human T cell lymphotropic virus type 1 (HTLV-1) infection. The HTLV-1 regulatory protein Tax aggressively accelerates the proliferation of host cells and is also an important target antigen for CD8(+) cytotoxic T cells (CTLs). We previously reported that several predominant HLA-A*24:02-restricted HTLV-1 Tax301-309-specific CTL clones commonly expressed a particular amino acid sequence motif (P-D-R) in complementarity-determining region 3 of T-cell receptor (TCR)-β chain among unrelated ATL patients who underwent allogeneic stem cell transplantation (allo-HSCT). Furthermore, a PDR-motif(+) CTL clone persistently existed in a long-term survivor as a central CTL clone with strong CTL activities after HSCT. Although a larger analysis of the relationship between PDR-motif(+) CTLs and the clinical course is required, the expression of PDR-motif(+) TCR on CD8(+) T cells may play a critical role in the management of anti-HTLV-1 activities for HLA-A24:02(+) ATL patients. Therefore, in this study, we prepared an HTLV-1 Tax301-309 peptide-specific CTL clone (HT-9) expressing PDR-motif(+) TCR isolated from a long-term survivor after HSCT, and evaluated its CTL activity against a variety of HTLV-1-infected T-cells from HLA-A*24:02(+) ATL patients. Before the assay of CTL function, we confirmed that HT-9 expressed less-differentiated effector-memory phenotypes (CD45RA(-)CCR7(-)CD27(+)CD28(+/-)CD57(+/-)) and T-cell exhaustion marker PD-1(+). In assays of CTL function, HT-9 recognized HTLV-1 Tax in an HLA-restricted fashion and demonstrated strong CTL activities against a variety of HTLV-1-infected T-cells from HLA-A*24:02(+) ATL patients regardless of whether the sources were autologous or allogeneic, but not normal cells. These data indicate that PDR-motif(+) TCR could be an important TCR candidate for TCR-gene immunotherapy for HLA-A24:02(+) ATL patients, provided

  18. Identification of full-length proviral DNA of porcine endogenous retrovirus from Chinese Wuzhishan miniature pigs inbred.

    PubMed

    Ma, Yuyuan; Lv, Maomin; Xu, Shu; Wu, Jianmin; Tian, Kegong; Zhang, Jingang

    2010-07-01

    Existence of porcine endogenous retrovirus (PERV) hinders pigs to be used in clinical xenotransplantation to alleviate the shortage of human transplants. Chinese miniature pigs are potential organ donors for xenotransplantation in China. However, so far, an adequate level of information on the molecular characteristics of PERV from Chinese miniature pigs has not been available. We described here the cloning and characterization of full-length proviral DNA of PERV from Chinese Wuzhishan miniature pigs inbred (WZSP). Full-length nucleotide sequences of PERV-WZSP and other PERVs were aligned and phylogenetic tree was constructed from deduced amino-acid sequences of env. The results demonstrated that the full-length proviral DNA of PERV-WZSP belongs to gammaretrovirus and shares high similarity with other PERVs. Sequence analysis also suggested that different patterns of LTR existed in the same porcine germ line and partial PERV-C sequence may recombine with PERV-A sequence in LTR.

  19. Preclinical electrogastrography in experimental pigs

    PubMed Central

    Květina, Jaroslav; Varayil, Jithinraj Edakkanambeth; Ali, Shahzad Marghoob; Kuneš, Martin; Bureš, Jan; Tachecí, Ilja; Rejchrt, Stanislav; Kopáčová, Marcela

    2010-01-01

    Surface electrogastrography (EGG) is a non-invasive means of recording gastric myoelectric activity or slow waves from cutaneous leads placed over the stomach. This paper provides a comprehensive review of preclinical EGG. Our group recently set up and worked out the methods for EGG in experimental pigs. We gained our initial experience in the use of EGG in assessment of porcine gastric myoelectric activity after volume challenge and after intragastric administration of itopride and erythromycin. The mean dominant frequency in pigs is comparable with that found in humans. EGG in experimental pigs is feasible. Experimental EGG is an important basis for further preclinical projects in pharmacology and toxicology. PMID:21217873

  20. To clone or not to clone--a Jewish perspective.

    PubMed Central

    Lipschutz, J H

    1999-01-01

    Many new reproductive methods such as artificial insemination, in vitro fertilisation, freezing of human embryos, and surrogate motherhood were at first widely condemned but are now seen in Western society as not just ethically and morally acceptable, but beneficial in that they allow otherwise infertile couples to have children. The idea of human cloning was also quickly condemned but debate is now emerging. This article examines cloning from a Jewish perspective and finds evidence to support the view that there is nothing inherently wrong with the idea of human cloning. A hypothesis is also advanced suggesting that even if a body was cloned, the brain, which is the essence of humanity, would remain unique. This author suggests that the debate should be changed from "Is cloning wrong?" to "When is cloning wrong?". PMID:10226913

  1. Ethical issues in animal cloning.

    PubMed

    Fiester, Autumn

    2005-01-01

    The issue of human reproductive cloning has recently received a great deal attention in public discourse. Bioethicists, policy makers, and the media have been quick to identify the key ethical issues involved in human reproductive cloning and to argue, almost unanimously, for an international ban on such attempts. Meanwhile, scientists have proceeded with extensive research agendas in the cloning of animals. Despite this research, there has been little public discussion of the ethical issues raised by animal cloning projects. Polling data show that the public is decidedly against the cloning of animals. To understand the public's reaction and fill the void of reasoned debate about the issue, we need to review the possible objections to animal cloning and assess the merits of the anti-animal cloning stance. Some objections to animal cloning (e.g., the impact of cloning on the population of unwanted animals) can be easily addressed, while others (e.g., the health of cloned animals) require more serious attention by the public and policy makers.

  2. Effects of resistant starch on behaviour, satiety-related hormones and metabolites in growing pigs.

    PubMed

    Souza da Silva, C; Haenen, D; Koopmans, S J; Hooiveld, G J E J; Bosch, G; Bolhuis, J E; Kemp, B; Müller, M; Gerrits, W J J

    2014-09-01

    Resistant starch (RS) has been suggested to prolong satiety in adult pigs. The present study investigated RS-induced changes in behaviour, satiety-related hormones and metabolites in catheterized growing pigs to explore possible underlying mechanisms for RS-induced satiety. In a cross-over design with two 14-day periods, 10 pigs (initial BW: 58 kg) were assigned to two treatments comprising diets containing either 35% pregelatinized starch (PS) or 34% retrograded starch (RS). Diets were isoenergetic on gross energy. Pigs were fed at 2.8× maintenance. Postprandial plasma response of satiety-related hormones and metabolites was measured at the end of each period using frequent blood sampling. Faecal and urinary energy losses were measured at the end of each period. Behaviour was scored 24 h from video recordings using scan sampling. Energy digestibility and metabolizability were ~6% lower in RS compared with PS diet (P<0.001), and metabolizable energy (ME) intake was ~3% lower in RS-fed than in PS-fed pigs (P<0.001). RS-fed pigs showed less feeder-directed (P=0.001) and drinking (P=0.10) behaviours than PS-fed pigs throughout the day. Postprandial peripheral short-chain fatty acid (SCFA) levels were higher in RS-fed than in PS-fed pigs (P<0.001). Postprandial glucose and insulin responses were lower in RS-fed than in PS-fed pigs (P<0.001). Triglyceride levels were higher in RS-fed than in PS-fed pigs (P<0.01), and non-esterified fatty acid levels did not differ between diets (P=0.90). Glucagon-like peptide-1 (GLP-1) levels were lower in RS-fed than in PS-fed pigs (P<0.001), and peptide tyrosine tyrosine (PYY) levels did not differ between diets (P=0.90). Blood serotonin levels were lower (P<0.001), whereas monoamine oxidase activity (P<0.05) and tryptophan (P<0.01) levels were higher in RS-fed than in PS-fed pigs. Despite a lower ME intake, RS seemed to prolong satiety, based on behavioural observations. Possible underlying mechanisms for RS-induced satiety include

  3. Gastrin in fetal and neonatal pigs.

    PubMed

    Xu, R J; Cranwell, P D

    1991-01-01

    1. The concentration and molecular profile of gastrin were examined in plasma and tissue extracts of fetal and neonatal pigs from 93 days gestation up to 12 weeks of age and also in the fetal gastric contents. 2. Gastrin was present in the gastrointestinal tract and plasma of fetal pigs at 93 days gestation. The concentration in both plasma and antral extracts increased progressively up to birth and continued to rise postnatally, reaching a peak at about 3 weeks of age in plasma and 6 weeks in the antrum. 3. In blood the major molecular form of gastrin was G34 (up to 80%), while in the antrum the major form was G17 (66-91%). The percentage of G34 in the antrum was highest in later gestation (21%), and reached adult proportion by 8 weeks of age (4%). 4. A considerable amount of gastrin, chiefly G17, was detected in the fetal gastric contents. Synthetic human G17 was stable in fetal gastric contents when incubated at 37 degrees C for 60 min, although, when incubated with gastric contents from a sow, it disappeared within 5 min. 5. It is suggested that the presence of gastrin in fetal gastric contents may be important in stimulation of fetal gut development.

  4. Human brucellosis at a pig slaughterhouse.

    PubMed

    Escobar, Gabriela I; Jacob, Néstor R; López, Gustavo; Ayala, Sandra M; Whatmore, Adrian M; Lucero, Nidia E

    2013-12-01

    Seventeen workers in a pig slaughterhouse with signs and symptoms compatible with brucellosis were clinically examined at the outpatient service of different health institutions and studied by serological tests during the period 2005-2011. Eleven blood cultures were taken and six Brucella suis strains were isolated, three biovar 1 and three with atypical characteristics. In order to confirm that these cases had no common source, a variable number of tandem repeat (VNTR) analyses were performed on 5 of the 6 strains whose results showed substantial heterogeneity in the genotypes, thereby demonstrating that the immediate origin was not the same. Two hundred adult pigs admitted for slaughter at the plant were sampled by convenience and tested by buffered antigen plate test (BPAT), serum agglutination test (SAT) and 2-mercapto-ethanol test (MET). Seven of 62 males (11%) and 25/138 (18%) females tested positive. The study results contribute information on risk scenarios for packing plant workers and underscore the need to improve plant workers' education on appropriate containment measures and to actively screen animals for swine brucellosis.

  5. [Measurement and analysis of hematology and blood chemistry parameters in northern pig-tailed macaques (Macaca leonina)].

    PubMed

    Pang, Wei; Lü, Long-Bao; Wang, Yun; Li, Gui; Huang, Dong-Ti; Lei, Ai-Hua; Zhang, Gao-Hong; Zheng, Yong-Tang

    2013-04-01

    The pig-tailed macaque is an important non-human primate experimental animal model that has been widely used in the research of AIDS and other diseases. Pig-tailed macaques include Mentawai macaques (Macaca pagensis), Sunda pig-tailed macaques (M. nemestrina) and northern pig-tailed macaques (M. leonina). Northern pig-tailed macaques inhabit China and surrounding Southeast Asia countries. To our knowledge, no reports have been published regarding the hematology and blood chemistry parameters of northern pig-tailed macaques, which are important for the objective evaluation of experimental results. We measured and analyzed 18 hematology parameters and 13 blood chemistry parameters in juvenile (aged 2-4 years) and adult (aged 5-10 years) northern pig-tailed macaques. We found that red blood cells, hemoglobin and alkaline phosphatase values were lower in female macaques than male macaques in both juvenile and adult groups. White blood cells, lymphocyte, monocytes, platelet distribution width, cholesterol, aspartate aminotransferase and alkaline phosphatase values were higher in juvenile macaques than adult macaques, while creatinine and triglycerides values were lower in juvenile macaques. Mean corpuscular hemoglobin and creatinine values were positively correlated with weight in juvenile groups. In adult groups, mean corpuscular hemoglobin, percentage of granulocyte, hemoglobin and creatinine were also positively correlated with weight, and lymphocyte, percentage of lymphocyte, red cell distribution width, aspartate aminotransferase and cholesterol values were negatively correlated with weight. The results suggest that age, gender and weight of northern pig-tailed macaques affected their hematology and blood chemistry parameters. This hematological and blood chemistry study has great significance in biomedical research and animal models using northern pig-tailed macaque as an experimental animal.

  6. Lessons learned from cloning dogs.

    PubMed

    Kim, M J; Oh, H J; Kim, G A; Park, J E; Park, E J; Jang, G; Ra, J C; Kang, S K; Lee, B C

    2012-08-01

    The aim of this article is to review dog cloning research and to suggest its applications based on a discussion about the normality of cloned dogs. Somatic cell nuclear transfer was successfully used for production of viable cloned puppies despite limited understanding of in vitro dog embryo production. Cloned dogs have similar growth characteristics to those born from natural fertilization, with no evidence of serious adverse effects. The offspring of cloned dogs also have similar growth performance and health to those of naturally bred puppies. Therefore, cloning in domestic dogs can be applied as an assisted reproductive technique to conserve endangered species, to treat sterile canids or aged dogs, to improve reproductive performance of valuable individuals and to generate disease model animals.

  7. Therapeutic cloning and reproductive liberty.

    PubMed

    Sparrow, Robert

    2009-04-01

    Concern for "reproductive liberty" suggests that decisions about embryos should normally be made by the persons who would be the genetic parents of the child that would be brought into existence if the embryo were brought to term. Therapeutic cloning would involve creating and destroying an embryo, which, if brought to term, would be the offspring of the genetic parents of the person undergoing therapy. I argue that central arguments in debates about parenthood and genetics therefore suggest that therapeutic cloning would be prima facie unethical unless it occurred with the consent of the parents of the person being cloned. Alternatively, if therapeutic cloning is thought to be legitimate, this undermines the case for some uses of reproductive cloning by implying that the genetic relation it establishes between clones and DNA donors does not carry the same moral weight as it does in cases of normal reproduction.

  8. Guinea pig models of asthma.

    PubMed

    McGovern, Alice E; Mazzone, Stuart B

    2014-01-01

    Described in this unit are methods for establishing guinea pig models of asthma. Sufficient detail is provided to enable investigators to study bronchoconstriction, cough, airway hyperresponsiveness, inflammation, and remodeling. PMID:25446291

  9. GPS Tracking of Free-Ranging Pigs to Evaluate Ring Strategies for the Control of Cysticercosis/Taeniasis in Peru

    PubMed Central

    Pray, Ian W.; Swanson, Dallas J.; Ayvar, Viterbo; Muro, Claudio; Moyano, Luz M.; Gonzalez, Armando E.; Garcia, Hector H.; O’Neal, Seth E.

    2016-01-01

    Background Taenia solium, a parasitic cestode that affects humans and pigs, is the leading cause of preventable epilepsy in the developing world. T. solium eggs are released into the environment through the stool of humans infected with an adult intestinal tapeworm (a condition called taeniasis), and cause cysticercosis when ingested by pigs or other humans. A control strategy to intervene within high-risk foci in endemic communities has been proposed as an alternative to mass antihelminthic treatment. In this ring strategy, antihelminthic treatment is targeted to humans and pigs residing within a 100 meter radius of a pig heavily-infected with cysticercosis. Our aim was to describe the roaming ranges of pigs in this region, and to evaluate whether the 100 meter radius rings encompass areas where risk factors for T. solium transmission, such as open human defecation and dense pig activity, are concentrated. Methodology/Principal Findings In this study, we used Global Positioning System (GPS) devices to track pig roaming ranges in two rural villages of northern Peru. We selected 41 pigs from two villages to participate in a 48-hour tracking period. Additionally, we surveyed all households to record the locations of open human defecation areas. We found that pigs spent a median of 82.8% (IQR: 73.5, 94.4) of their time roaming within 100 meters of their homes. The size of home ranges varied significantly by pig age, and 93% of the total time spent interacting with open human defecation areas occurred within 100 meters of pig residences. Conclusions/Significance These results indicate that 100 meter radius rings around heavily-infected pigs adequately capture the average pig’s roaming area (i.e., home range) and represent an area where the great majority of exposure to human feces occurs. PMID:27035825

  10. [The mutation site analysis on CAPN1 gene of Wild boar, Min pig and Yorkshire].

    PubMed

    Yang, Xiu-Qin; Liu, Hui; Guo, Li-Juan; Xu, Yao; Liu, Di

    2007-05-01

    In order to further evaluate the relationship between the variations of CAPN1 gene and meat tenderness, the CAPN1 genomic sequences were cloned and sequenced, its CDS was analyzed with PCR-SSCP, and the genotype analyses covered 109 individuals from Wild boar, Min pig and Yorkshire. Fifteen of total 21 introns were cloned. Five pairs polymorphic primers for PCR-SSCP analysis were designed based on the CDS of CAPN1 from GenBank and the cloned introns. Eight SNPs, resulting from single point mutation G to C, C to T, T to C, G to A, G to A, G to A, C to T and C to T at the base position 161 in exon2, 60 in exon5, 96 in exon5, 119 in exon5, 270 in intron8, 83 in exon10, 126 in exon13 and 138 in exon13 respectively, were identified, and 3 of which are missense mutations resulting to amino acid substitutions of S/T, G/E, V/I at the amino acid position of 54, 192 and 363 respectively. chi2 analysis showed that the distribution of genotypes among Yorkshire, Min pig and Wild boar are extremely significant difference, while there are no significant difference be-tween Min pig and Wild boar except in the S1 primer. The polymorphic sites may be used as molecular markers for meat tenderness and pork quality.

  11. Human cloning and child welfare.

    PubMed Central

    Burley, J; Harris, J

    1999-01-01

    In this paper we discuss an objection to human cloning which appeals to the welfare of the child. This objection varies according to the sort of harm it is expected the clone will suffer. The three formulations of it that we will consider are: 1. Clones will be harmed by the fearful or prejudicial attitudes people may have about or towards them (H1); 2. Clones will be harmed by the demands and expectations of parents or genotype donors (H2); 3. Clones will be harmed by their own awareness of their origins, for example the knowledge that the genetic donor is a stranger (H3). We will show why these three versions of the child welfare objection do not necessarily supply compelling reasons to ban human reproductive cloning. The claim that we will develop and defend in the course of our discussion is that even if it is the case that a cloned child will suffer harms of the type H1-H3, it is none the less permissible to conceive by cloning so long as these cloning-induced welfare deficits are not such as to blight the existence of the resultant child, whoever this may be. PMID:10226914

  12. Therapeutic cloning: The ethical limits

    SciTech Connect

    Whittaker, Peter A. . E-mail: p.whittaker@lancaster.ac.uk

    2005-09-01

    A brief outline of stem cells, stem cell therapy and therapeutic cloning is given. The position of therapeutic cloning with regard to other embryonic manipulations - IVF-based reproduction, embryonic stem formation from IVF embryos and reproductive cloning - is indicated. The main ethically challenging stages in therapeutic cloning are considered to be the nuclear transfer process including the source of eggs for this and the destruction of an embryo to provide stem cells for therapeutic use. The extremely polarised nature of the debate regarding the status of an early human embryo is noted, and some potential alternative strategies for preparing immunocompatible pluripotent stem cells are indicated.

  13. Review: influenza virus in pigs.

    PubMed

    Crisci, Elisa; Mussá, Tufária; Fraile, Lorenzo; Montoya, Maria

    2013-10-01

    Influenza virus disease still remains one of the major threats to human health, involving a wide range of animal species and pigs play an important role in influenza ecology. Pigs were labeled as "mixing vessels" since they are susceptible to infection with avian, human and swine influenza viruses and genetic reassortment between these viruses can occur. After the H1N1 influenza pandemic of 2009 with a swine origin virus, the most recent research in "influenzology" is directed at improving knowledge of porcine influenza virus infection. This tendency is probably due to the fact that domestic pigs are closely related to humans and represent an excellent animal model to study various microbial infectious diseases. In spite of the role of the pig in influenza virus ecology, swine immune responses against influenza viruses are not fully understood. Considering these premises, the aim of this review is to focus on the in vitro studies performed with porcine cells and influenza virus and on the immune responses of pigs against human, avian and swine influenza viruses in vivo. The increased acceptance of pigs as suitable and valuable models in the scientific community may stimulate the development of new tools to assess porcine immune responses, paving the way for their consideration as the future "gold standard" large-animal model in immunology.

  14. Geographic Correlation between Tapeworm Carriers and Heavily Infected Cysticercotic Pigs

    PubMed Central

    O'Neal, Seth E.; Moyano, Luz M.; Ayvar, Viterbo; Gonzalvez, Guillermo; Diaz, Andre; Rodriguez, Silvia; Wilkins, Patricia P.; Tsang, Victor C. W.; Gilman, Robert H.; Garcia, Hector H.; Gonzalez, Armando E.

    2012-01-01

    Background Neurocysticercosis is a leading cause of preventable epilepsy in the developing world. Sustainable community-based interventions are urgently needed to control transmission of the causative parasite, Taenia solium. We examined the geospatial relationship between live pigs with visible cysticercotic cysts on their tongues and humans with adult intestinal tapeworm infection (taeniasis) in a rural village in northern Peru. The objective was to determine whether tongue-positive pigs could indicate high-risk geographic foci for taeniasis to guide targeted screening efforts. This approach could offer significant benefit compared to mass intervention. Methods We recorded geographic coordinates of all village houses, collected stool samples from all consenting villagers, and collected blood and examined tongues of all village pigs. Stool samples were processed by enzyme-linked immunosorbent assay (ELISA) for presence of Taenia sp. coproantigens indicative of active taeniasis; serum was processed by enzyme-linked immunoelectrotransfer blot for antibodies against T. solium cysticercosis (EITB LLGP) and T. solium taeniasis (EITB rES33). Findings Of 548 pigs, 256 (46.7%) were positive for antibodies against cysticercosis on EITB LLGP. Of 402 fecal samples, 6 (1.5%) were positive for the presence of Taenia sp. coproantigens. The proportion of coproantigen-positive individuals differed significantly between residents living within 100-meters of a tongue-positive pig (4/79, 5.1%) and residents living >100 meters from a tongue-positive pig (2/323, 0.6%) (p = 0.02). The prevalence of taeniasis was >8 times higher among residents living within 100 meters of a tongue-positive pig compared to residents living outside this range (adjusted PR 8.1, 95% CI 1.4–47.0). Conclusions Tongue-positive pigs in endemic communities can indicate geospatial foci in which the risk for taeniasis is increased. Targeted screening or presumptive treatment for taeniasis within these high

  15. Quality and safety of bovine clones and their products.

    PubMed

    Heyman, Y; Chavatte-Palmer, P; Fromentin, G; Berthelot, V; Jurie, C; Bas, P; Dubarry, M; Mialot, J P; Remy, D; Richard, C; Martignat, L; Vignon, X; Renard, J P

    2007-08-01

    cattle derived from somatic nuclear transfer than in cattle born from conventional reproduction. Our results confirm that the quality and safety of products (milk and meat) from adult and clinically healthy cloned cattle is globally similar to normal animals. However, from a strictly biological point of view, the slightly delayed maturation we observed in the muscle of clones together with some marginal differences identified in FA composition of both muscle and milk, point to the need for more refined analysis to totally exclude any risks from the consumption of those products.

  16. Self-Cloning CRISPR.

    PubMed

    Arbab, Mandana; Sherwood, Richard I

    2016-01-01

    CRISPR/Cas9-gene editing has emerged as a revolutionary technology to easily modify specific genomic loci by designing complementary sgRNA sequences and introducing these into cells along with Cas9. Self-cloning CRISPR/Cas9 (scCRISPR) uses a self-cleaving palindromic sgRNA plasmid (sgPal) that recombines with short PCR-amplified site-specific sgRNA sequences within the target cell by homologous recombination to circumvent the process of sgRNA plasmid construction. Through this mechanism, scCRISPR enables gene editing within 2 hr once sgRNA oligos are available, with high efficiency equivalent to conventional sgRNA targeting: >90% gene knockout in both mouse and human embryonic stem cells and cancer cell lines. Furthermore, using PCR-based addition of short homology arms, we achieve efficient site-specific knock-in of transgenes such as GFP without traditional plasmid cloning or genome-integrated selection cassette (2% to 4% knock-in rate). The methods in this paper describe the most rapid and efficient means of CRISPR gene editing. © 2016 by John Wiley & Sons, Inc. PMID:27532819

  17. Self-Cloning CRISPR.

    PubMed

    Arbab, Mandana; Sherwood, Richard I

    2016-01-01

    CRISPR/Cas9-gene editing has emerged as a revolutionary technology to easily modify specific genomic loci by designing complementary sgRNA sequences and introducing these into cells along with Cas9. Self-cloning CRISPR/Cas9 (scCRISPR) uses a self-cleaving palindromic sgRNA plasmid (sgPal) that recombines with short PCR-amplified site-specific sgRNA sequences within the target cell by homologous recombination to circumvent the process of sgRNA plasmid construction. Through this mechanism, scCRISPR enables gene editing within 2 hr once sgRNA oligos are available, with high efficiency equivalent to conventional sgRNA targeting: >90% gene knockout in both mouse and human embryonic stem cells and cancer cell lines. Furthermore, using PCR-based addition of short homology arms, we achieve efficient site-specific knock-in of transgenes such as GFP without traditional plasmid cloning or genome-integrated selection cassette (2% to 4% knock-in rate). The methods in this paper describe the most rapid and efficient means of CRISPR gene editing. © 2016 by John Wiley & Sons, Inc.

  18. 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.

  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. PMID:27558653

  20. Construction and characterization of a full-length cDNA infectious clone of emerging porcine Senecavirus A.

    PubMed

    Chen, Zhenhai; Yuan, Fangfeng; Li, Yanhua; Shang, Pengcheng; Schroeder, Robin; Lechtenberg, Kelly; Henningson, Jamie; Hause, Benjamin; Bai, Jianfa; Rowland, Raymond R R; Clavijo, Alfonso; Fang, Ying

    2016-10-01

    A full-length cDNA infectious clone, pKS15-01-Clone, was constructed from an emerging Senecavirus A (SVA; strain KS15-01). To explore the potential use as a viral backbone for expressing marker genes, the enhanced green fluorescent protein (EGFP)-tagged reporter virus (vKS15-01-EGFP) was generated using reverse genetics. Compared to the parental virus, the pKS15-01-Clone derived virus (vKS15-01-Clone) replicated efficiently in vitro and in vivo, and induced similar levels of neutralizing antibody and cytokine responses in infected animals. In contrast, the vKS15-01-EGFP virus showed impaired growth ability and induced lower level of immune response in infected animals. Lesions on the dorsal snout and coronary bands were observed in all pigs infected by parental virus KS15-01, but not in pigs infected with vKS15-01-Clone or vKS15-01-EGFP viruses. These results demonstrated that the infectious clone and EGFP reporter virus could be used as important tools in further elucidating the SVA pathogenesis and development of control measures.

  1. The pig genome project has plenty to squeal about.

    PubMed

    Fan, B; Gorbach, D M; Rothschild, M F

    2011-01-01

    Significant progress on pig genetics and genomics research has been witnessed in recent years due to the integration of advanced molecular biology techniques, bioinformatics and computational biology, and the collaborative efforts of researchers in the swine genomics community. Progress on expanding the linkage map has slowed down, but the efforts have created a higher-resolution physical map integrating the clone map and BAC end sequence. The number of QTL mapped is still growing and most of the updated QTL mapping results are available through PigQTLdb. Additionally, expression studies using high-throughput microarrays and other gene expression techniques have made significant advancements. The number of identified non-coding RNAs is rapidly increasing and their exact regulatory functions are being explored. A publishable draft (build 10) of the swine genome sequence was available for the pig genomics community by the end of December 2010. Build 9 of the porcine genome is currently available with Ensembl annotation; manual annotation is ongoing. These drafts provide useful tools for such endeavors as comparative genomics and SNP scans for fine QTL mapping. A recent community-wide effort to create a 60K porcine SNP chip has greatly facilitated whole-genome association analyses, haplotype block construction and linkage disequilibrium mapping, which can contribute to whole-genome selection. The future 'systems biology' that integrates and optimizes the information from all research levels can enhance the pig community's understanding of the full complexity of the porcine genome. These recent technological advances and where they may lead are reviewed.

  2. Rapid DNA extraction of pig ear tissues.

    PubMed

    Kunhareang, S; Zhou, H; Hickford, J G H

    2010-07-01

    A single-step DNA isolation procedure from pig tissues was developed and the product used directly for polymerase chain reaction (PCR) amplification, single-strand conformational polymorphism (SSCP) analysis and sequencing. The procedure consists of proteinase K digestion of 2-10mg of fresh tissue, at 55 degrees C for 1h, followed by application of the products of digestion to filter paper. A 1.2mm-diameter punch of that paper has sufficient DNA to act as a template for PCR amplification. The quality of the genomic DNA appeared to be high as the PCR amplicons produced sharp banding patterns on both agarose gel electrophoresis and on SSCP analysis, and they could be used for DNA sequencing following cloning. The dried genomic DNA on filter paper can be kept at room temperature. The procedure is considered effective as it is simple, fast and inexpensive. It would be useful for large-scale genotyping and could be used to obtain genomic DNA from various tissues.

  3. [The discrete horror of cloning].

    PubMed

    Guibourg, Ricardo A

    2009-01-01

    The author raises the topic of cloning after the decision of the Argentine government, which concerned for the "dignity of the human person", passed a decree of need and urgency, No. 200/97 (Annex), prohibiting cloning experiments with human beings. Therefore, considering that the topic is so terribly urgent and necessary, the author feels it is timely to consider it.

  4. CATO: The Clone Alignment Tool.

    PubMed

    Henstock, Peter V; LaPan, Peter

    2016-01-01

    High-throughput cloning efforts produce large numbers of sequences that need to be aligned, edited, compared with reference sequences, and organized as files and selected clones. Different pieces of software are typically required to perform each of these tasks. We have designed a single piece of software, CATO, the Clone Alignment Tool, that allows a user to align, evaluate, edit, and select clone sequences based on comparisons to reference sequences. The input and output are designed to be compatible with standard data formats, and thus suitable for integration into a clone processing pipeline. CATO provides both sequence alignment and visualizations to facilitate the analysis of cloning experiments. The alignment algorithm matches each of the relevant candidate sequences against each reference sequence. The visualization portion displays three levels of matching: 1) a top-level summary of the top candidate sequences aligned to each reference sequence, 2) a focused alignment view with the nucleotides of matched sequences displayed against one reference sequence, and 3) a pair-wise alignment of a single reference and candidate sequence pair. Users can select the minimum matching criteria for valid clones, edit or swap reference sequences, and export the results to a summary file as part of the high-throughput cloning workflow.

  5. Animal Cloning and Food Safety

    MedlinePlus

    ... from clones and their offspring out of the food chain until CVM could further evaluate the issue. back to top FDA Studies Cloning For more than five years, CVM ... evaluate the safety of food from these animals. The resulting report, called a ...

  6. CATO: The Clone Alignment Tool.

    PubMed

    Henstock, Peter V; LaPan, Peter

    2016-01-01

    High-throughput cloning efforts produce large numbers of sequences that need to be aligned, edited, compared with reference sequences, and organized as files and selected clones. Different pieces of software are typically required to perform each of these tasks. We have designed a single piece of software, CATO, the Clone Alignment Tool, that allows a user to align, evaluate, edit, and select clone sequences based on comparisons to reference sequences. The input and output are designed to be compatible with standard data formats, and thus suitable for integration into a clone processing pipeline. CATO provides both sequence alignment and visualizations to facilitate the analysis of cloning experiments. The alignment algorithm matches each of the relevant candidate sequences against each reference sequence. The visualization portion displays three levels of matching: 1) a top-level summary of the top candidate sequences aligned to each reference sequence, 2) a focused alignment view with the nucleotides of matched sequences displayed against one reference sequence, and 3) a pair-wise alignment of a single reference and candidate sequence pair. Users can select the minimum matching criteria for valid clones, edit or swap reference sequences, and export the results to a summary file as part of the high-throughput cloning workflow. PMID:27459605

  7. CATO: The Clone Alignment Tool

    PubMed Central

    Henstock, Peter V.; LaPan, Peter

    2016-01-01

    High-throughput cloning efforts produce large numbers of sequences that need to be aligned, edited, compared with reference sequences, and organized as files and selected clones. Different pieces of software are typically required to perform each of these tasks. We have designed a single piece of software, CATO, the Clone Alignment Tool, that allows a user to align, evaluate, edit, and select clone sequences based on comparisons to reference sequences. The input and output are designed to be compatible with standard data formats, and thus suitable for integration into a clone processing pipeline. CATO provides both sequence alignment and visualizations to facilitate the analysis of cloning experiments. The alignment algorithm matches each of the relevant candidate sequences against each reference sequence. The visualization portion displays three levels of matching: 1) a top-level summary of the top candidate sequences aligned to each reference sequence, 2) a focused alignment view with the nucleotides of matched sequences displayed against one reference sequence, and 3) a pair-wise alignment of a single reference and candidate sequence pair. Users can select the minimum matching criteria for valid clones, edit or swap reference sequences, and export the results to a summary file as part of the high-throughput cloning workflow. PMID:27459605

  8. [The discrete horror of cloning].

    PubMed

    Guibourg, Ricardo A

    2009-01-01

    The author raises the topic of cloning after the decision of the Argentine government, which concerned for the "dignity of the human person", passed a decree of need and urgency, No. 200/97 (Annex), prohibiting cloning experiments with human beings. Therefore, considering that the topic is so terribly urgent and necessary, the author feels it is timely to consider it. PMID:19860340

  9. [Scientific ethics of human cloning].

    PubMed

    Valenzuela, Carlos Y

    2005-01-01

    True cloning is fission, budding or other types of asexual reproduction. In humans it occurs in monozygote twinning. This type of cloning is ethically and religiously good. Human cloning can be performed by twinning (TWClo) or nuclear transfer (NTClo). Both methods need a zygote or a nuclear transferred cell, obtained in vitro (IVTec). They are under the IVTec ethics. IVTecs use humans (zygotes, embryos) as drugs or things; increase the risk of malformations; increase development and size of abnormalities and may cause long-term changes. Cloning for preserving extinct (or almost extinct) animals or humans when sexual reproduction is not possible is ethically valid. The previous selection of a phenotype in human cloning violates some ethical principles. NTClo for reproductive or therapeutic purposes is dangerous since it increases the risk for nucleotide or chromosome mutations, de-programming or re-programming errors, aging or malignancy of the embryo cells thus obtained.

  10. Therapeutic cloning: promises and issues

    PubMed Central

    Kfoury, Charlotte

    2007-01-01

    Advances in biotechnology necessitate both an understanding of scientific principles and ethical implications to be clinically applicable in medicine. In this regard, therapeutic cloning offers significant potential in regenerative medicine by circumventing immunorejection, and in the cure of genetic disorders when used in conjunction with gene therapy. Therapeutic cloning in the context of cell replacement therapy holds a huge potential for de novo organogenesis and the permanent treatment of Parkinson’s disease, Duchenne muscular dystrophy, and diabetes mellitus as shown by in vivo studies. Scientific roadblocks impeding advancement in therapeutic cloning are tumorigenicity, epigenetic reprogramming, mitochondrial heteroplasmy, interspecies pathogen transfer, low oocyte availability. Therapeutic cloning is also often tied to ethical considerations concerning the source, destruction and moral status of IVF embryos based on the argument of potential. Legislative and funding issues are also addressed. Future considerations would include a distinction between therapeutic and reproductive cloning in legislative formulations. PMID:18523539

  11. Animal cloning: problems and prospects.

    PubMed

    Wells, D N

    2005-04-01

    An efficient animal cloning technology would provide many new opportunities for livestock agriculture, human medicine, and animal conservation. Nuclear cloning involves the production of animals that are genetically identical to the donor cells used in a technique known as nuclear transfer (NT). However, at present it is an inefficient process: in cattle, only around 6% of the embryos transferred to the reproductive tracts of recipient cows result in healthy, longterm surviving clones. Of concern are the high losses throughout gestation, during birth and in the post-natal period through to adulthood. Many of the pregnancy losses relate to failure of the placenta to develop and function correctly. Placental dysfunction may also have an adverse influence on postnatal health. These anomalies are probably due to incorrect epigenetic reprogramming of the donor genome following NT, leading to inappropriate patterns of gene expression during the development of clones. Whilst some physiological tests on surviving clones suggest normality, other reports indicate a variety of post-natal clone-associated abnormalities. This variability in outcome may reflect species-specific and/or cloning methodological differences. Importantly, to date it appears that these clone-associated phenotypes are not transmitted to offspring following sexual reproduction. This indicates that they represent epigenetic errors, rather than genetic errors, which are corrected during gametogenesis. Whilst this needs confirmation at the molecular level, it provides initial confidence in the first application of NT in agriculture, namely, the production of small numbers of cloned sires from genetically elite bulls, for natural mating, to effectively disseminate genetic gain. In addition to the animal welfare concerns with the technology, the underlying health of the animals and the consequential effect on food safety are critical aspects that require investigation to gain regulatory and consumer

  12. Differential Analysis of the Nasal Microbiome of Pig Carriers or Non-Carriers of Staphylococcus aureus.

    PubMed

    Espinosa-Gongora, Carmen; Larsen, Niels; Schønning, Kristian; Fredholm, Merete; Guardabassi, Luca

    2016-01-01

    Staphylococcus aureus is presently regarded as an emerging zoonotic agent due to the spread of specific methicillin-resistant S. aureus (MRSA) clones in pig farms. Studying the microbiota can be useful for the identification of bacteria that antagonize such opportunistic veterinary and zoonotic pathogen in animal carriers. The aim of this study was to determine whether the nasal microbiome of pig S. aureus carriers differs from that of non-carriers. The V3-V5 region of the 16S rRNA gene was sequenced from nasal swabs of 44 S. aureus carriers and 56 non-carriers using the 454 GS FLX titanium system. Carriers and non-carriers were selected on the basis of quantitative longitudinal data on S. aureus carriage in 600 pigs sampled at 20 Danish herds included in two previous studies in Denmark. Raw sequences were analysed with the BION meta package and the resulting abundance matrix was analysed using the DESeq2 package in R to identify operational taxonomic units (OTUs) with differential abundance between S. aureus carriers and non-carriers. Twenty OTUs were significantly associated to non-carriers, including species with known probiotic potential and antimicrobial effect such as lactic acid-producing isolates described among Leuconostoc spp. and some members of the Lachnospiraceae family, which is known for butyrate production. Further 5 OTUs were significantly associated to carriage, including known pathogenic bacteria such as Pasteurella multocida and Klebsiella spp. Our results show that the nasal microbiome of pigs that are not colonized with S. aureus harbours several species/taxa that are significantly less abundant in pig carriers, suggesting that the nasal microbiota may play a role in the individual predisposition to S. aureus nasal carriage in pigs. Further research is warranted to isolate these bacteria and assess their possible antagonistic effect on S. aureus for the pursuit of new strategies to control MRSA in pig farming. PMID:27509169

  13. Characterization and phylogenetic analysis of the swine leukocyte antigen 3 gene from Korean native pigs.

    PubMed

    Chung, H Y; Choi, Y C; Park, H N

    2015-05-18

    We investigated the phylogenetic relationships between pig breeds, compared the genetic similarity between humans and pigs, and provided basic genetic information on Korean native pigs (KNPs), using genetic variants of the swine leukocyte antigen 3 (SLA-3) gene. Primers were based on sequences from GenBank (accession Nos. AF464010 and AF464009). Polymerase chain reaction analysis amplified approximately 1727 bp of segments, which contained 1086 bp of coding regions and 641 bp of the 3'- and 5'-untranslated regions. Bacterial artificial chromosome clones of miniature pigs were used for sequencing the SLA-3 genomic region, which was 3114 bp in total length, including the coding (1086 bp) and non-coding (2028 bp) regions. Sequence analysis detected 53 single nucleotide polymorphisms (SNPs), based on a minor allele frequency greater than 0.01, which is low compared with other pig breeds, and the results suggest that there is low genetic variability in KNPs. Comparative analysis revealed that humans possess approximately three times more genetic variation than do pigs. Approximately 71% of SNPs in exons 2 and 3 were detected in KNPs, and exon 5 in humans is a highly polymorphic region. Newly identified sequences of SLA-3 using KNPs were submitted to GenBank (accession No. DQ992512-18). Cluster analysis revealed that KNPs were grouped according to three major alleles: SLA-3*0502 (DQ992518), SLA-3*0302 (DQ992513 and DQ992516), and SLA-3*0303 (DQ992512, DQ992514, DQ992515, and DQ992517). Alignments revealed that humans have a relatively close genetic relationship with pigs and chimpanzees. The information provided by this study may be useful in KNP management.

  14. Differential Analysis of the Nasal Microbiome of Pig Carriers or Non-Carriers of Staphylococcus aureus

    PubMed Central

    Espinosa-Gongora, Carmen; Larsen, Niels; Schønning, Kristian; Fredholm, Merete; Guardabassi, Luca

    2016-01-01

    Staphylococcus aureus is presently regarded as an emerging zoonotic agent due to the spread of specific methicillin-resistant S. aureus (MRSA) clones in pig farms. Studying the microbiota can be useful for the identification of bacteria that antagonize such opportunistic veterinary and zoonotic pathogen in animal carriers. The aim of this study was to determine whether the nasal microbiome of pig S. aureus carriers differs from that of non-carriers. The V3-V5 region of the 16S rRNA gene was sequenced from nasal swabs of 44 S. aureus carriers and 56 non-carriers using the 454 GS FLX titanium system. Carriers and non-carriers were selected on the basis of quantitative longitudinal data on S. aureus carriage in 600 pigs sampled at 20 Danish herds included in two previous studies in Denmark. Raw sequences were analysed with the BION meta package and the resulting abundance matrix was analysed using the DESeq2 package in R to identify operational taxonomic units (OTUs) with differential abundance between S. aureus carriers and non-carriers. Twenty OTUs were significantly associated to non-carriers, including species with known probiotic potential and antimicrobial effect such as lactic acid-producing isolates described among Leuconostoc spp. and some members of the Lachnospiraceae family, which is known for butyrate production. Further 5 OTUs were significantly associated to carriage, including known pathogenic bacteria such as Pasteurella multocida and Klebsiella spp. Our results show that the nasal microbiome of pigs that are not colonized with S. aureus harbours several species/taxa that are significantly less abundant in pig carriers, suggesting that the nasal microbiota may play a role in the individual predisposition to S. aureus nasal carriage in pigs. Further research is warranted to isolate these bacteria and assess their possible antagonistic effect on S. aureus for the pursuit of new strategies to control MRSA in pig farming. PMID:27509169

  15. Experimental aerosolized guinea pig-adapted Zaire ebolavirus (variant: Mayinga) causes lethal pneumonia in guinea pigs.

    PubMed

    Twenhafel, N A; Shaia, C I; Bunton, T E; Shamblin, J D; Wollen, S E; Pitt, L M; Sizemore, D R; Ogg, M M; Johnston, S C

    2015-01-01

    Eight guinea pigs were aerosolized with guinea pig-adapted Zaire ebolavirus (variant: Mayinga) and developed lethal interstitial pneumonia that was distinct from lesions described in guinea pigs challenged subcutaneously, nonhuman primates challenged by the aerosol route, and natural infection in humans. Guinea pigs succumbed with significant pathologic changes primarily restricted to the lungs. Intracytoplasmic inclusion bodies were observed in many alveolar macrophages. Perivasculitis was noted within the lungs. These changes are unlike those of documented subcutaneously challenged guinea pigs and aerosolized filoviral infections in nonhuman primates and human cases. Similar to findings in subcutaneously challenged guinea pigs, there were only mild lesions in the liver and spleen. To our knowledge, this is the first report of aerosol challenge of guinea pigs with guinea pig-adapted Zaire ebolavirus (variant: Mayinga). Before choosing this model for use in aerosolized ebolavirus studies, scientists and pathologists should be aware that aerosolized guinea pig-adapted Zaire ebolavirus (variant: Mayinga) causes lethal pneumonia in guinea pigs.

  16. Molecular cloning and sequencing of zeta-crystallin/quinone reductase cDNA from human liver.

    PubMed

    Gonzalez, P; Rao, P V; Zigler, J S

    1993-03-31

    Zeta-crystallin is an enzyme-crystallin highly expressed in the lens of some hystricomorph rodents and camels. It has been shown to have a novel NADPH: quinone oxidoreductase activity and is present at enzymatic levels in a variety of tissues from various mammals. We report here the cDNA cloning of zeta-crystallin from a human liver library. One clone with the complete open reading frame was obtained. Ten nucleotides of the 5' and 796 of the 3' nontranslated regions are present in the clone including two possible polyadenylation signals. The deduced amino acid sequence is 328 residues long with a calculated molecular mass of 34910 daltons and isoelectric point of 8.73. It shows 84% identity with the guinea pig protein.

  17. Human cloning: can it be made safe?

    PubMed

    Rhind, Susan M; Taylor, Jane E; De Sousa, Paul A; King, Tim J; McGarry, Michelle; Wilmut, Ian

    2003-11-01

    There are continued claims of attempts to clone humans using nuclear transfer, despite the serious problems that have been encountered in cloning other mammals. It is known that epigenetic and genetic mechanisms are involved in clone failure, but we still do not know exactly how. Human reproductive cloning is unethical, but the production of cells from cloned embryos could offer many potential benefits. So, can human cloning be made safe?

  18. The recent history of somatic cloning in mammals.

    PubMed

    Brem, Gottfried; Kühholzer, Birgit

    2002-01-01

    The history of somatic cell nuclear transfer (NT) in mammals is full of exciting experiments and findings regarding the technique and outcome of NT, despite only covering a period of 6 years. The production of Dolly, for the first time demonstrating cloning from an adult somatic cell, had a great impact on subsequent studies. However, the more progress we make, the more obvious it becomes how little we know about the processes during NT, specifically how reprogramming events occur. Therefore, it is certainly challenging to continue investigating every step of somatic cell NT more intensively, starting from the donor cell, (type, cell cycle, synchronization, population doublings) and continuing until the cloned offspring are born and even further, to see how and if NT has an influence on health, viability, quantitative traits, and reproduction of cloned individuals.

  19. The recent history of somatic cloning in mammals.

    PubMed

    Brem, Gottfried; Kühholzer, Birgit

    2002-01-01

    The history of somatic cell nuclear transfer (NT) in mammals is full of exciting experiments and findings regarding the technique and outcome of NT, despite only covering a period of 6 years. The production of Dolly, for the first time demonstrating cloning from an adult somatic cell, had a great impact on subsequent studies. However, the more progress we make, the more obvious it becomes how little we know about the processes during NT, specifically how reprogramming events occur. Therefore, it is certainly challenging to continue investigating every step of somatic cell NT more intensively, starting from the donor cell, (type, cell cycle, synchronization, population doublings) and continuing until the cloned offspring are born and even further, to see how and if NT has an influence on health, viability, quantitative traits, and reproduction of cloned individuals. PMID:12006157

  20. Molecular basis of non-responsiveness to peroxisome proliferators: the guinea-pig PPARalpha is functional and mediates peroxisome proliferator-induced hypolipidaemia.

    PubMed Central

    Bell, A R; Savory, R; Horley, N J; Choudhury, A I; Dickins, M; Gray, T J; Salter, A M; Bell, D R

    1998-01-01

    The guinea pig does not undergo peroxisome proliferation in response to peroxisome proliferators, in contrast with other rodents. To understand the molecular basis of this phenotype, the peroxisome proliferator activated receptor alpha (PPARalpha) from guinea-pig liver was cloned; it encodes a protein of 467 amino acid residues that is similar to rodent and human PPARalpha. The guinea-pig PPARalpha showed a high substitution rate: maximum likelihood analysis was consistent with rodent monophyly, but could not exclude rodent polyphyly (P approximately 0.06). The guinea-pig PPARalpha cDNA was expressed in 293 cells and mediated the induction of the luciferase reporter gene by the peroxisome proliferator, Wy-14,643, dependent on the presence of a peroxisome proliferator response element. Moreover the PPARalpha RNA and protein were expressed in guinea-pig liver, although at lower levels than in a species which is responsive to peroxisome proliferators, the mouse. To determine whether the guinea-pig PPARalpha mediated any physiological effects, guinea pigs were exposed to two selective PPARalpha agonists, Wy-14, 643 and methylclofenapate; both compounds induced hypolipidaemia. Thus the guinea pig is a useful model for human responses to peroxisome proliferators. PMID:9620871

  1. 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

  2. Wildlife conservation and reproductive cloning.

    PubMed

    Holt, William V; Pickard, Amanda R; Prather, Randall S

    2004-03-01

    Reproductive cloning, or the production of offspring by nuclear transfer, is often regarded as having potential for conserving endangered species of wildlife. Currently, however, low success rates for reproductive cloning limit the practical application of this technique to experimental use and proof of principle investigations. In this review, we consider how cloning may contribute to wildlife conservation strategies. The cloning of endangered mammals presents practical problems, many of which stem from the paucity of knowledge about their basic reproductive biology. However, situations may arise where resources could be targeted at recovering lost or under-represented genetic lines; these could then contribute to the future fitness of the population. Approaches of this type would be preferable to the indiscriminate generation of large numbers of identical individuals. Applying cloning technology to non-mammalian vertebrates may be more practical than attempting to use conventional reproductive technologies. As the scientific background to cloning technology was pioneered using amphibians, it may be possible to breed imminently threatened amphibians, or even restore extinct amphibian species, by the use of cloning. In this respect species with external embryonic development may have an advantage over mammals as developmental abnormalities associated with inappropriate embryonic reprogramming would not be relevant.

  3. Guinea-pig reaginic antibody

    PubMed Central

    Margni, R. A.; Hajos, Silvia E.

    1973-01-01

    The physicochemical and biological properties of purified guinea-pig reaginic antibody were studied. It is a labile protein different to γ1. Its antibody activity is completely destroyed by heating at 56° for 6 hours and by treatment with mercaptoethanol. The capacity to give PCA is decreased by repeated freezing and thawing. It is a bivalent antibody, haemagglutinating, does not fix complement and is capable of sensitizing guinea-pig skin for PCA reaction after a latent period of a week but not after 3 hours. Reaginic antibody appears on day 7–8 after the first inoculation and the higher levels (PCA reaction) are obtained at the eleventh to thirteenth days. After the fifteenth to seventeenth days the PCA is negative. The reaginic antibody does not pass the placenta. Higher levels of reaginic antibody were obtained when the guinea-pigs were inoculated with the antigen in saline with simultaneous inoculation, intraperitoneally, of killed Bordetella pertussis, phase I. PMID:4354828

  4. Guinea-pig reaginic antibody

    PubMed Central

    Margni, R. A.; Hajos, Silvia E.

    1973-01-01

    The methods for isolation and purification of a guinea-pig serum protein with homocytotropic antibody activity and characteristics of IgE are described. By precipitation in the equivalence zone or immunoadsorption and chromatography on DEAE-cellulose, we isolated an homocytotropic antibody, that was not able to give a precipitin line when it was reacted directly with the antigen. It was capable of sensitizing guinea-pig skin for PCA after a latent period of 24–48 hours but not after 3 hours; it was sensitive to treatment with mercaptoethanol. It had antigenic determinants present in the other guinea-pig immunoglobulins and particular antigenic determinants. All these properties make us believe that this protein belongs to an immunoglobulin different from γ1 and similar to the reaginic antibody (IgE) described in other species. ImagesFIG. 3FIG. 4FIG. 5 PMID:4126261

  5. Anthelmintic effects of phytogenic feed additives in Ascaris suum inoculated pigs.

    PubMed

    van Krimpen, M M; Binnendijk, G P; Borgsteede, F H M; Gaasenbeek, C P H

    2010-03-25

    Two experiments were performed to determine the anthelmintic effect of some phytogenic feed additives on a mild infection of Ascaris suum in growing and finishing pigs. Usually, an infection of A. suum is controlled by using conventional synthetic drugs. Organic farmers, however, prefer a non-pharmaceutical approach to worm control. Therefore, phytotherapy could be an appropriate alternative. In the first experiment, a commercial available organic starter diet was supplemented with 3% of a herb mixture, adding 1% Thymus vulgaris, 1% Melissa officinalis and 1% Echinacea purpurea to the diet, or with 4% of a herb mixture, thereby adding the mentioned herbs plus 1% Camellia sinensis (black tea). A negative control group (no treatment) and a positive control group (treatment with conventional synthetic drug flubendazole) were included. In the second experiment, the anthelmintic properties against A. suum of three individual herbs, Carica papaya, Peumus boldus and Artemisia vulgaris, each in a dose of 1%, were tested. Pigs were infected with 1000 infective worm eggs each. Each experiment was performed with 32 individually housed growing pigs (8 replicates/treatment), which were monitored for 67 days. It was hypothesized that the herbs would block the cycles of the larvae, thereby preventing the development of adult worms. Therefore, phytogenic feed additives were not supplied during the whole experimental period, but only from the start until D39. Pigs were inoculated with infective worm eggs during five consecutive days (D17-D21). At D67 all pigs were dissected, whereafter livers were checked for the presence of white spots. Also numbers of worms in the small intestine were counted. In experiment 1, the numbers of worm-infected pigs were similar for both the herb supplemented (groups 3 and 4) and the unsupplemented (group 1) treatments (5-6 pigs of 8), while the treatment with flubendazole (group 2) resulted in 0 infected pigs. In experiment 2, herb addition (groups 2

  6. Therapeutic cloning in the mouse.

    PubMed

    Mombaerts, Peter

    2003-09-30

    Nuclear transfer technology can be applied to produce autologous differentiated cells for therapeutic purposes, a concept termed therapeutic cloning. Countless articles have been published on the ethics and politics of human therapeutic cloning, reflecting the high expectations from this new opportunity for rejuvenation of the aging or diseased body. Yet the research literature on therapeutic cloning, strictly speaking, is comprised of only four articles, all in the mouse. The efficiency of derivation of embryonic stem cell lines via nuclear transfer is remarkably consistent among these reports. However, the efficiency is so low that, in its present form, the concept is unlikely to become widespread in clinical practice.

  7. Cloning: revisiting an old debate.

    PubMed

    Verhey, Allen D

    1994-09-01

    The debate about cloning that took place 25 years ago, although directed toward a different sort of cloning, elucidates fundamental issues currently at stake in reproductive technologies and research. Paul Ramsey and Joseph Fletcher were participants in this early debate. The differences between Ramsey and Fletcher about the meaning and sufficiency of freedom, the understanding and weighing of good and evil, the connection between embodiment and personhood, the relationship of humans with nature, and the meaning of parenthood suggest both a broader agenda for the debate about cloning and a cautious move forward in the development of embryo-splitting.

  8. Coccidiosis in guinea-pigs.

    PubMed

    Ellis, P A; Wright, A E

    1961-07-01

    The attention of laboratory workers is drawn to the possibility of coccidiosis as a cause of death in guinea-pigs. The purchase of a number of guinea-pigs infected with this protozoon was followed by 12 deaths when these animals were injected with material for diagnostic purposes. No deaths occurred in the laboratory stock herd, as these were kept separate from the newcomers and were not infected. The life history of the parasite is described, together with the post-mortem findings in our series of animals.

  9. Human therapeutic cloning (NTSC): applying research from mammalian reproductive cloning.

    PubMed

    French, Andrew J; Wood, Samuel H; Trounson, Alan O

    2006-01-01

    Human therapeutic cloning or nuclear transfer stem cells (NTSC) to produce patient-specific stem cells, holds considerable promise in the field of regenerative medicine. The recent withdrawal of the only scientific publications claiming the successful generation of NTSC lines afford an opportunity to review the available research in mammalian reproductive somatic cell nuclear transfer (SCNT) with the goal of progressing human NTSC. The process of SCNT is prone to epigenetic abnormalities that contribute to very low success rates. Although there are high mortality rates in some species of cloned animals, most surviving clones have been shown to have normal phenotypic and physiological characteristics and to produce healthy offspring. This technology has been applied to an increasing number of mammals for utility in research, agriculture, conservation, and biomedicine. In contrast, attempts at SCNT to produce human embryonic stem cells (hESCs) have been disappointing. Only one group has published reliable evidence of success in deriving a cloned human blastocyst, using an undifferentiated hESC donor cell, and it failed to develop into a hESC line. When optimal conditions are present, it appears that in vitro development of cloned and parthenogenetic embryos, both of which may be utilized to produce hESCs, may be similar to in vitro fertilized embryos. The derivation of ESC lines from cloned embryos is substantially more efficient than the production of viable offspring. This review summarizes developments in mammalian reproductive cloning, cell-to-cell fusion alternatives, and strategies for oocyte procurement that may provide important clues facilitating progress in human therapeutic cloning leading to the successful application of cell-based therapies utilizing autologous hESC lines.

  10. Induction of systemic IFITM3 expression does not effectively control foot-and-mouth disease viral infection in transgenic pigs.

    PubMed

    Zhang, Huawei; Zheng, Haixue; Qian, Ping; Xu, Jinfang; Yang, Xi; Zhou, Rui; Chen, Huanchun; Li, Xiangmin

    2016-08-15

    Foot-and-mouth disease (FMD) is a highly contagious disease of cloven-hoofed animals, and can cause severe economic loss. Interferon-induced transmembrane (IFITM) proteins constitute a family of viral restriction factors that can inhibit the replication of several types of viruses. Our previous study showed that overexpression of swine IFITM3 (sIFITM3) impeded replication of the FMD virus (FMDV) in BHK-21 cells and mice. In this study, sIFITM3-transgenic (TG) pigs were produced by handmade cloning. Results showed that sIFITM3 was highly overexpressed in many organs of sIFITM3-TG pigs compared to wild-type pigs. After a virulent FMDV strain (O/ES/2001) was intramuscularly inoculated, the sIFITM3-TG pigs showed slightly higher susceptibility to FMDV infection than wild-type pigs. Both groups displayed comparable degrees of clinical symptoms throughout the 14-day observation period. Therefore, the induction of systemic sIFITM3 expression does not protect pigs against FMDV infection. Based on these observations, we propose that a combination of interferons and vaccines be used to control FMDV infections and subsequent FMD outbreaks. PMID:27374903

  11. SLA typing using the PCR-SSP method and establishment of the SLA homozygote line in pedigreed SNU miniature pigs.

    PubMed

    Yeom, Su-Cheong; Park, Chung-Gyu; Lee, Byeong-Chun; Lee, Wang-Jae

    2010-04-01

    Seoul National University (SNU) miniature pigs represent a closed colony with 24 founder pigs and a well preserved pedigree. Characterization using mRNA sequence analysis was conducted for 6 swine leukocyte antigen (SLA) loci in parental or founder pigs, and 17 defined alleles were detected. Based on these complete coding sequences, 17 sequence specific primers (SSPs) were designed for polymorphic sites. To validate the specificity of each allele SSP, the PCR-SSP was conducted with defined allele clones as templates. PCR-SSP was conducted with the hot start polymerase and touch-down PCR. The parental or found SNU miniature pigs showed overall SLA class I and II heterozygotes. Using the established PCR-SSP method, we conducted SLA typing for breeding stock including 2 pedigreed pigs and identified the novel SLA class II homozygote haplotye (DRA*0201, DRB1*0403, DQA*0102 and DQB1*0701) and 2 SLA homozygote pig lines: SLA class I Hp-3.0 and class II Hp-0.3, and SLA class I Hp-2.0 and class II Hp-0.2. We thought that our PCR-SSP SLA typing method could be applicable for new SLA homozygote line establishment by assignment and scheduled breeding.

  12. Methylotroph cloning vehicle

    DOEpatents

    Hanson, Richard S.; Allen, Larry N.

    1989-04-25

    A cloning vehicle comprising: a replication determinant effective for replicating the vehicle in a non-C.sub.1 -utilizing host and in a C.sub.1 -utilizing host; DNA effective to allow the vehicle to be mobilized from the non-C.sub.1 -utilizing host to the C.sub.1 -utilizing host; DNA providing resistance to two antibiotics to which the wild-type C.sub.1 -utilizing host is susceptible, each of the antibiotic resistance markers having a recognition site for a restriction endonuclease; a cos site; and a means for preventing replication in the C.sub.1 -utilizing host. The vehicle is used for complementation mapping as follows. DNA comprising a gene from the C.sub.1 -utilizing organism is inserted at the restriction nuclease recognition site, inactivating the antibiotic resistance marker at that site. The vehicle can then be used to form a cosmid structure to infect the non-C.sub.1 -utilizing (e.g., E. coli) host, and then conjugated with a selected C.sub.1 -utilizing mutant. Resistance to the other antibiotic by the mutant is a marker of the conjugation. Other phenotypical changes in the mutant, e.g., loss of an auxotrophic trait, is attributed to the C.sub.1 gene. The vector is also used to inactivate genes whose protein products catalyze side reactions that divert compounds from a biosynthetic pathway to a desired product, thereby producing an organism that makes the desired product in higher yields.

  13. Impermanence of bacterial clones

    PubMed Central

    Bobay, Louis-Marie; Traverse, Charles C.; Ochman, Howard

    2015-01-01

    Bacteria reproduce asexually and pass on a single genome copied from the parent, a reproductive mode that assures the clonal descent of progeny; however, a truly clonal bacterial species is extremely rare. The signal of clonality can be interrupted by gene uptake and exchange, initiating homologous recombination that results in the unique sequence of one clone being incorporated into another. Because recombination occurs sporadically and on local scales, these events are often difficult to recognize, even when considering large samples of completely sequenced genomes. Moreover, several processes can produce the appearance of clonality in populations that undergo frequent recombination. The rates and consequences of recombination have been studied in Escherichia coli for over 40 y, and, during this time, there have been several shifting views of its clonal status, population structure, and rates of gene exchange. We reexamine the studies and retrace the evolution of the methods that have assessed the extent of DNA flux, largely focusing on its impact on the E. coli genome. PMID:26195749

  14. 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.

  15. 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

  16. Late effects of radiation on the lumbar spinal cord of guinea pigs: Re-treatment tolerance

    SciTech Connect

    Mason, K.A. ); Withers, H.R.; Chiang, Chi-Shiun )

    1993-07-15

    Using a guinea pig model of lumbar myelopathy, various factors affecting the tolerance of spinal cord to irradiation were assessed: (a) extent of initial injury; (b) time interval between priming and test doses; and (c) animal age at the time of initial radiation treatment. A 3 cm section of lumbar spinal cord of guinea pigs was irradiated with fractionated doses of 4.5 Gy gamma rays given as 9 fractions per week. Guinea pigs were primed with 9 x 4.5 Gy in 7 days which is 60% of the ED[sub 50] for a continuous course of treatment. After 28 or 40 weeks, animal were retreated with 6-14 fractions of 4.5 Gy. Animals were observed for 2 years following the priming dose and both the incidence and latency of myelopathy recorded. Young adult guinea pigs (8 wk old) showed both a decreased radiation tolerance and latency compared to old individuals (40 wk old). At 28 or 40 wk after 9 x 4.5 Gy, only about 8% of the initial injury was remembered in young adult guinea pigs. The amount of residual injury was dependent on the initial damage as a proportion of the tolerance dose. The spinal cord shows a greater capacity for long-term recovery than generally appreciated and re-treatment doses clinically prescribed may be lower than necessary. 8 refs., 3 figs., 2 tabs.

  17. Resolution of murine chlamydial genital infection by the adoptive transfer of a biovar-specific, Th1 lymphocyte clone.

    PubMed

    Igietseme, J U; Ramsey, K H; Magee, D M; Williams, D M; Kincy, T J; Rank, R G

    1993-01-01

    MoPn-specific T-cell clones were isolated from a T-cell line that was capable of curing chlamydial genital infection by the Chlamydia trachomatis agent of mouse pneumonitis (MoPn) after adoptive transfer. Two clones (designated as 2.14-0 and 2.14-3) were characterized by flow cytometry techniques to be homogenous for L3T4, CD3, and alpha/beta T cell receptor (TcR) T-helper cell markers. The two clones were biovar specific, because they reacted to MoPn but not the Chlamydia psittaci agent of guinea pig inclusion conjunctivitis (GPIC) or C. trachomatis, serovar type E. Cytokine profile analysis, by a combination of bioassays, ELISA, and slot/Northern blotting for specific cytokine messenger RNAs, further revealed that cultures of antigen-stimulated clone 2.14-0 contained interleukin-2 (IL-2), tumor necrosis factor-alpha, and gamma interferon (a T helper 1 cell [Th1] profile). Clone 2.14-3 was also positive for gamma interferon, a level much lower than that of clone 2.14-0, and negative for IL-4 secretion, suggesting a Th1 profile as well. The ability of these clones to bring about the resolution of the chronic genital chlamydial infection of nude mice was tested by the adoptive transfer of 10(7) cells of each clone into the mice. By 4 weeks after cell transfer of clone 2.14-0, 81% of recipient nude mice (30 of 37) resolved the disease. In contrast, clone 2.14-3 or a control T-cell clone specific for a heterologous antigen were unable to resolve the infection in 20 recipients in each case, even after 100 days.(ABSTRACT TRUNCATED AT 250 WORDS)

  18. A Clone of Your Own.

    ERIC Educational Resources Information Center

    Bilodeau, Kirsten

    1997-01-01

    Describes an activity used at the Washington Park Arboretum that helps students understand cloning through plant propagation. Students also learn how to make a pot from recycled newspapers and how to make soil that is appropriate for the plants. (DDR)

  19. Cloning of a quantum measurement

    SciTech Connect

    Bisio, Alessandro; D'Ariano, Giacomo Mauro; Perinotti, Paolo; Sedlak, Michal

    2011-10-15

    We analyze quantum algorithms for cloning of a quantum measurement. Our aim is to mimic two uses of a device performing an unknown von Neumann measurement with a single use of the device. When the unknown device has to be used before the bipartite state to be measured is available we talk about 1{yields}2 learning of the measurement, otherwise the task is called 1{yields}2 cloning of a measurement. We perform the optimization for both learning and cloning for arbitrary dimension d of the Hilbert space. For 1{yields}2 cloning we also propose a simple quantum network that achieves the optimal fidelity. The optimal fidelity for 1{yields}2 learning just slightly outperforms the estimate and prepare strategy in which one first estimates the unknown measurement and depending on the result suitably prepares the duplicate.

  20. Human Cloning: Let's Discuss It.

    ERIC Educational Resources Information Center

    Taras, Loretta; Stavroulakis, Anthea M.; Ortiz, Mary T.

    1999-01-01

    Describes experiences with holding discussions on cloning at a variety of levels in undergraduate biology courses. Discusses teaching methods used and student reactions to the discussions. Contains 12 references. (WRM)

  1. Human cloning and 'posthuman' society.

    PubMed

    Blackford, Russell

    2005-01-01

    Since early 1997, when the creation of Dolly the sheep by somatic cell nuclear transfer was announced in Nature, numerous government reports, essays, articles and books have considered the ethical problems and policy issues surrounding human reproductive cloning. In this article, I consider what response a modern liberal society should give to the prospect of human cloning, if it became safe and practical. Some opponents of human cloning have argued that permitting it would place us on a slippery slope to a repugnant future society, comparable to that portrayed in Aldous Huxley's novel, Brave New World. I conclude that, leaving aside concerns about safety, none of the psychological or social considerations discussed in this article provides an adequate policy justification for invoking the state's coercive powers to prevent human cloning.

  2. Are cloned quantum states macroscopic?

    PubMed

    Fröwis, F; Dür, W

    2012-10-26

    We study quantum states produced by optimal phase covariant quantum cloners. We argue that cloned quantum superpositions are not macroscopic superpositions in the spirit of Schrödinger's cat, despite their large particle number. This is indicated by calculating several measures for macroscopic superpositions from the literature, as well as by investigating the distinguishability of the two superposed cloned states. The latter rapidly diminishes when considering imperfect detectors or noisy states and does not increase with the system size. In contrast, we find that cloned quantum states themselves are macroscopic, in the sense of both proposed measures and their usefulness in quantum metrology with an optimal scaling in system size. We investigate the applicability of cloned states for parameter estimation in the presence of different kinds of noise.

  3. Human cloning and 'posthuman' society.

    PubMed

    Blackford, Russell

    2005-01-01

    Since early 1997, when the creation of Dolly the sheep by somatic cell nuclear transfer was announced in Nature, numerous government reports, essays, articles and books have considered the ethical problems and policy issues surrounding human reproductive cloning. In this article, I consider what response a modern liberal society should give to the prospect of human cloning, if it became safe and practical. Some opponents of human cloning have argued that permitting it would place us on a slippery slope to a repugnant future society, comparable to that portrayed in Aldous Huxley's novel, Brave New World. I conclude that, leaving aside concerns about safety, none of the psychological or social considerations discussed in this article provides an adequate policy justification for invoking the state's coercive powers to prevent human cloning. PMID:16007753

  4. Postmortem findings in cloned and transgenic piglets dead before weaning.

    PubMed

    Schmidt, M; Winther, K D; Secher, J O; Callesen, H

    2015-10-01

    frequent in Yucatan piglets (26.9% vs. 2.1% in LW and 5.3% in Göttingen). In conclusion, these results show that pig cloning results in a considerable loss of piglets and that many of these can be related to various malformations that all are also seen in noncloned piglets. Because approximately half of the cloned piglets still survive, even with eventual unknown minor malformations, use of pigs as models for human diseases is still realistic. However, continued efforts are needed to further reduce the level of malformations. PMID:26166169

  5. Local cloning of entangled qubits

    SciTech Connect

    Choudhary, Sujit K.; Kunkri, Samir; Rahaman, Ramij; Roy, Anirban

    2007-11-15

    We discuss the exact cloning of orthogonal but entangled qubits under local operations and classical communication. The amount of entanglement necessary in a blank copy is obtained for various cases. Surprisingly, this amount is more than 1 ebit for certain sets of two nonmaximal but equally entangled states of two qubits. To clone any three Bell states, at least log{sub 2} 3 ebit is necessary.

  6. Cloning and expression of mouse legumain, a lysosomal endopeptidase.

    PubMed Central

    Chen, J M; Dando, P M; Stevens, R A; Fortunato, M; Barrett, A J

    1998-01-01

    Legumain, a recently discovered mammalian cysteine endopeptidase, was found in all mouse tissues examined, but was particularly abundant in kidney and placenta. The distribution in subcellular fractions of mouse and rat kidney showed a lysosomal localization, and activity was detectable only after the organelles were disrupted. Nevertheless, ratios of legumain activity to that of cathepsin B differed considerably between mouse tissues. cDNA encoding mouse legumain was cloned and sequenced, the deduced amino acid sequence proving to be 83% identical to that of the human protein [Chen, Dando, Rawlings, Brown, Young, Stevens, Hewitt, Watts and Barrett (1997) J. Biol. Chem. 272, 8090-8098]. Recombinant mouse legumain was expressed in human embryonic kidney 293 cells by use of a vector containing a cytomegalovirus promoter. The recombinant enzyme was partially purified and found to be an asparagine-specific endopeptidase closely similar to naturally occurring pig kidney legumain. PMID:9742219

  7. A Pilot Study of Uterine Artery Embolization with Tris-Acryl Gelatin Microspheres in Guinea Pigs

    SciTech Connect

    Zhuang Wenquan; Tan Guosheng; Guo Wenbo; Yang Jianyong

    2012-06-15

    Objective: This study was designed to establish guinea pigs as an animal model for uterine artery embolization (UAE) with tris-acryl gelatin microspheres (TAGM). Methods: Twenty-five female adult guinea pigs were randomly divided into two groups, including a uterine artery casting mould group (n = 10) and a UAE group (n = 15). Pelvic angiography and vascular casting mould were performed in the first group. The anatomical characters of the pelvic cavity in guinea pigs were described. In the second group, the technical feasibility of performing UAE with TAGM in guinea pigs was investigated. The histopathological slides of the uterus of guinea pigs after UAE were examined to inspect the outcomes of UAE. Results: The uterine artery springs from the internal iliac artery, ascends tortuously along the cervix, and gives off vertically 8-10 branches to the cervix uteri and uterine horns. The diameters of the trunk of the uterine artery and its first branch were 0.32 {+-} 0.027 mm and 0.14 {+-} 0.01 mm, respectively. For UAE animals, the dosages of 40-120 and 100-300 {mu}m TAGM were 0.033 {+-} 0.003 ml and 0.015 {+-} 0.002 ml, respectively. On histopathological slides, embosphere particles were found in the first branches of the uterine artery, the subserous arteries, and the intramural arteries. Inflammatory reactions in the uterus were common in guinea pigs after UAE. Local or dispersed areas of necrosis in uterus also were observed in a few guinea pigs. Conclusions: Guinea pigs are an appropriate and feasible model for UAE with TAGM.

  8. Artificial cloning of domestic animals.

    PubMed

    Keefer, Carol L

    2015-07-21

    Domestic animals can be cloned using techniques such as embryo splitting and nuclear transfer to produce genetically identical individuals. Although embryo splitting is limited to the production of only a few identical individuals, nuclear transfer of donor nuclei into recipient oocytes, whose own nuclear DNA has been removed, can result in large numbers of identical individuals. Moreover, clones can be produced using donor cells from sterile animals, such as steers and geldings, and, unlike their genetic source, these clones are fertile. In reality, due to low efficiencies and the high costs of cloning domestic species, only a limited number of identical individuals are generally produced, and these clones are primarily used as breed stock. In addition to providing a means of rescuing and propagating valuable genetics, somatic cell nuclear transfer (SCNT) research has contributed knowledge that has led to the direct reprogramming of cells (e.g., to induce pluripotent stem cells) and a better understanding of epigenetic regulation during embryonic development. In this review, I provide a broad overview of the historical development of cloning in domestic animals, of its application to the propagation of livestock and transgenic animal production, and of its scientific promise for advancing basic research.

  9. Artificial cloning of domestic animals.

    PubMed

    Keefer, Carol L

    2015-07-21

    Domestic animals can be cloned using techniques such as embryo splitting and nuclear transfer to produce genetically identical individuals. Although embryo splitting is limited to the production of only a few identical individuals, nuclear transfer of donor nuclei into recipient oocytes, whose own nuclear DNA has been removed, can result in large numbers of identical individuals. Moreover, clones can be produced using donor cells from sterile animals, such as steers and geldings, and, unlike their genetic source, these clones are fertile. In reality, due to low efficiencies and the high costs of cloning domestic species, only a limited number of identical individuals are generally produced, and these clones are primarily used as breed stock. In addition to providing a means of rescuing and propagating valuable genetics, somatic cell nuclear transfer (SCNT) research has contributed knowledge that has led to the direct reprogramming of cells (e.g., to induce pluripotent stem cells) and a better understanding of epigenetic regulation during embryonic development. In this review, I provide a broad overview of the historical development of cloning in domestic animals, of its application to the propagation of livestock and transgenic animal production, and of its scientific promise for advancing basic research. PMID:26195770

  10. Artificial cloning of domestic animals

    PubMed Central

    Keefer, Carol L.

    2015-01-01

    Domestic animals can be cloned using techniques such as embryo splitting and nuclear transfer to produce genetically identical individuals. Although embryo splitting is limited to the production of only a few identical individuals, nuclear transfer of donor nuclei into recipient oocytes, whose own nuclear DNA has been removed, can result in large numbers of identical individuals. Moreover, clones can be produced using donor cells from sterile animals, such as steers and geldings, and, unlike their genetic source, these clones are fertile. In reality, due to low efficiencies and the high costs of cloning domestic species, only a limited number of identical individuals are generally produced, and these clones are primarily used as breed stock. In addition to providing a means of rescuing and propagating valuable genetics, somatic cell nuclear transfer (SCNT) research has contributed knowledge that has led to the direct reprogramming of cells (e.g., to induce pluripotent stem cells) and a better understanding of epigenetic regulation during embryonic development. In this review, I provide a broad overview of the historical development of cloning in domestic animals, of its application to the propagation of livestock and transgenic animal production, and of its scientific promise for advancing basic research. PMID:26195770

  11. Cloning goes to the movies.

    PubMed

    Cormick, Craig

    2006-10-01

    Public attitude research conducted by Biotechnology Australia shows that one of the major sources of information on human reproductive cloning is movies. Traditionally, understanding of new and emerging technologies has come through the mass media but human cloning, being so widely addressed through the popular culture of movies, is more effectively defined by Hollywood than the news media or science media. But how well are the science and social issues of cloning portrayed in box office hits such as The Island, Multiplicity, Star Wars: Attack of the Clones and Jurassic Park? These movies have enormous reach and undoubted influence, and are therefore worth analyzing in some detail. This study looks at 33 movies made between 1971 and 2005 that address human reproductive cloning, and it categorizes the films based on their genre and potential influence. Yet rather than simply rating the quality of the science portrayed, the study compares the key messages in these movies with public attitudes towards cloning, to examine the correlations.

  12. Islamic perspectives on human cloning.

    PubMed

    Sadeghi, Mahmoud

    2007-01-01

    The present paper seeks to assess various views from Islamic jurists relating to human cloning, which is one of the controversial topics in the recent past. Taking Islamic jurisprudence principles, such as the rule of necessity for self preservation and respect for human beings, the rule of la darar wa la dirar ('the necessity to refrain from causing harm to oneself and others') and the rule of usr wa haraj, one may indicate that if human cloning could not be prohibited, as such, it could still be opposed because it gives way to various harmful consequences, which include family disorder, chaos in the clone's family relationships, physical and mental diseases for clones and suffering of egg donors and surrogate mothers. However with due attention to the fact that the reasons behind the prohibition of abortion only restrict the destruction of human embryos in their post-implantation stages, human cloning for biomedical research and exploitation of stem cells from cloned embryos at the blastocyst stage for therapeutic purposes would be acceptable.

  13. Cloning goes to the movies.

    PubMed

    Cormick, Craig

    2006-10-01

    Public attitude research conducted by Biotechnology Australia shows that one of the major sources of information on human reproductive cloning is movies. Traditionally, understanding of new and emerging technologies has come through the mass media but human cloning, being so widely addressed through the popular culture of movies, is more effectively defined by Hollywood than the news media or science media. But how well are the science and social issues of cloning portrayed in box office hits such as The Island, Multiplicity, Star Wars: Attack of the Clones and Jurassic Park? These movies have enormous reach and undoubted influence, and are therefore worth analyzing in some detail. This study looks at 33 movies made between 1971 and 2005 that address human reproductive cloning, and it categorizes the films based on their genre and potential influence. Yet rather than simply rating the quality of the science portrayed, the study compares the key messages in these movies with public attitudes towards cloning, to examine the correlations. PMID:17214211

  14. Cloning of a developmentally regulated tegument antigen of Schistosoma mansoni.

    PubMed

    Stein, L D; David, J R

    1986-09-01

    We have cloned a gene encoding a 22.6 kDa antigen from a Schistosoma mansoni cDNA library. Northern blots indicate that transcription of this antigen occurs in adults and sporocysts but not in cercariae, eggs or in newly-transformed schistosomula. Immunoprecipitation and Western blotting with specific antisera indicate that the antigen is not detectable in the newly transformed schistosomulum but appears within 24 h of schistosomulum transformation. Indirect immunofluorescence of adult worms shows this protein to be located in the tegument.

  15. Fermented liquid feed for pigs.

    PubMed

    Missotten, Joris A M; Michiels, Joris; Ovyn, Anneke; De Smet, Stefaan; Dierick, Noël A

    2010-12-01

    Since the announcement of the ban on the use of antibiotics as antimicrobial growth promoters in the feed of pigs in 2006 the investigation towards alternative feed additives has augmented considerably. Although fermented liquid feed is not an additive, but a feeding strategy, the experimental work examining its possible advantages also saw a rise. The use of fermented liquid feed (FLF) has two main advantages, namely that the simultaneous provision of feed and water may result in an alleviation of the transition from the sow milk to solid feed and may also reduce the time spent to find both sources of nutrients, and secondly, that offering FLF with a low pH may strengthen the potential of the stomach as a first line of defence against possible pathogenic infections. Because of these two advantages, FLF is often stated as an ideal feed for weaned piglets. The results obtained so far are rather variable, but in general they show a better body weight gain and worse feed/gain ratio for the piglets. However, for growing-finishing pigs on average a better feed/gain ratio is found compared to pigs fed dry feed. This better performance is mostly associated with less harmful microbiota and better gut morphology. This review provides an overview of the current knowledge of FLF for pigs,dealing with the FLF itself as well as its effect on the gastrointestinal tract and animal performance. PMID:21214019

  16. Fermented liquid feed for pigs.

    PubMed

    Missotten, Joris A M; Michiels, Joris; Ovyn, Anneke; De Smet, Stefaan; Dierick, Noël A

    2010-12-01

    Since the announcement of the ban on the use of antibiotics as antimicrobial growth promoters in the feed of pigs in 2006 the investigation towards alternative feed additives has augmented considerably. Although fermented liquid feed is not an additive, but a feeding strategy, the experimental work examining its possible advantages also saw a rise. The use of fermented liquid feed (FLF) has two main advantages, namely that the simultaneous provision of feed and water may result in an alleviation of the transition from the sow milk to solid feed and may also reduce the time spent to find both sources of nutrients, and secondly, that offering FLF with a low pH may strengthen the potential of the stomach as a first line of defence against possible pathogenic infections. Because of these two advantages, FLF is often stated as an ideal feed for weaned piglets. The results obtained so far are rather variable, but in general they show a better body weight gain and worse feed/gain ratio for the piglets. However, for growing-finishing pigs on average a better feed/gain ratio is found compared to pigs fed dry feed. This better performance is mostly associated with less harmful microbiota and better gut morphology. This review provides an overview of the current knowledge of FLF for pigs,dealing with the FLF itself as well as its effect on the gastrointestinal tract and animal performance.

  17. Methylotroph cloning vehicle

    DOEpatents

    Hanson, R.S.; Allen, L.N.

    1989-04-25

    A cloning vehicle comprising: a replication determinant effective for replicating the vehicle in a non-C[sub 1]-utilizing host and in a C[sub 1]-utilizing host; DNA effective to allow the vehicle to be mobilized from the non-C[sub 1]-utilizing host to the C[sub 1]-utilizing host; DNA providing resistance to two antibiotics to which the wild-type C[sub 1]-utilizing host is susceptible, each of the antibiotic resistance markers having a recognition site for a restriction endonuclease; a cos site; and a means for preventing replication in the C[sub 1]-utilizing host. The vehicle is used for complementation mapping as follows. DNA comprising a gene from the C[sub 1]-utilizing organism is inserted at the restriction nuclease recognition site, inactivating the antibiotic resistance marker at that site. The vehicle can then be used to form a cosmid structure to infect the non-C[sub 1]-utilizing (e.g., E. coli) host, and then conjugated with a selected C[sub 1]-utilizing mutant. Resistance to the other antibiotic by the mutant is a marker of the conjugation. Other phenotypical changes in the mutant, e.g., loss of an auxotrophic trait, is attributed to the C[sub 1] gene. The vector is also used to inactivate genes whose protein products catalyze side reactions that divert compounds from a biosynthetic pathway to a desired product, thereby producing an organism that makes the desired product in higher yields. 3 figs.

  18. Imperfect Cloning Operations in Algebraic Quantum Theory

    NASA Astrophysics Data System (ADS)

    Kitajima, Yuichiro

    2015-01-01

    No-cloning theorem says that there is no unitary operation that makes perfect clones of non-orthogonal quantum states. The objective of the present paper is to examine whether an imperfect cloning operation exists or not in a C*-algebraic framework. We define a universal -imperfect cloning operation which tolerates a finite loss of fidelity in the cloned state, and show that an individual system's algebra of observables is abelian if and only if there is a universal -imperfect cloning operation in the case where the loss of fidelity is less than . Therefore in this case no universal -imperfect cloning operation is possible in algebraic quantum theory.

  19. Apparent lack of beta 3-adrenoceptors and of insulin regulation of glucose transport in brown adipose tissue of guinea pigs.

    PubMed

    Himms-Hagen, J; Triandafillou, J; Begin-Heick, N; Ghorbani, M; Kates, A L

    1995-01-01

    Norepinephrine-induced thermogenesis was substantial in adipocytes from brown adipose tissue (BAT) of cold-acclimated guinea pigs but absent in adipocytes from BAT of warm-acclimated guinea pigs. There was no thermogenic response to any beta 3-adrenergic agonist (CL-316,243, ZD-7114, BRL-28410, CGP-12177). The receptor was characterized as a beta 1-adrenoceptor. Adrenergic agonists stimulated adenylate cyclase in membranes from BAT of both warm- and cold-acclimated guinea pigs also via a beta 1-adrenoceptor; beta 3-adrenergic agonists had no effect. Glucose transport by brown adipocytes from warm-acclimated guinea pigs was not stimulated by either norepinephrine or insulin. Cold acclimation induced the appearance of stimulation of glucose transport by norepinephrine in association with the appearance of a large capacity for thermogenesis, but there was little improvement in response to insulin. GLUT4 was present in membranes from BAT of both warm- and cold-acclimated guinea pigs. Insulin is known to have an antilipolytic effect on both BAT and white adipose tissue of guinea pigs. Thus there is a selective lack of insulin-regulated glucose transport that is not improved by cold acclimation. Guinea pigs may have a mutated component of the translocation mechanism for GLUT4. beta 3-Adrenoceptors appear to be absent in brown adipocytes of adult guinea pigs, as in white adipocytes of guinea pigs, yet are known to be present in the gut. Tissue-specific expression of beta 3-adrenergic receptors in guinea pigs may differ from that in rats, in which receptors are expressed in the adipose tissues and gut. PMID:7840345

  20. Local cloning of two product states

    SciTech Connect

    Ji Zhengfeng; Feng Yuan; Ying Mingsheng

    2005-09-15

    Local quantum operations and classical communication (LOCC) put considerable constraints on many quantum information processing tasks such as cloning and discrimination. Surprisingly, however, discrimination of any two pure states survives such constraints in some sense. We show that cloning is not that lucky; namely, probabilistic LOCC cloning of two product states is strictly less efficient than global cloning. We prove our result by giving explicitly the efficiency formula of local cloning of any two product states.

  1. Tracheal ultrastructure in kerosene treated guinea pigs. A preliminary report.

    PubMed

    Noa, N; Sanabria, J

    1984-01-01

    p6high correlation between the usage of kerosene and the appearance of asthmatic crises has been demonstrated. The ultrastructure of the upper respiratory tract of animals treated with kerosene has not been previously reported. Kerosene aerosol was administered for 15 minutes daily during 21 days to adult male guinea pigs with fragments of trachea being processed for ultrathin electron microscopical studies. Controls did not receive any treatment. Trachea of guinea pigs submitted to kerosene aerosols showed swelling, ruffling and breakdown of the ciliary membrane. The regularly arranged ciliary border was disturbed to a certain degree in some areas by the development of cytoplasmatic protrusions at the apical part of the ciliated cells. An eosinophilic infiltrate was observed deep inside the epithelium and into the lamina propria. Therefore, these ciliary alterations can be considered as one of the most important changes induced by kerosene in tracheal epithelial cells. The protrusions may represent a sign of cell alteration produced by kerosene aerosol inhalation in the guinea pig.

  2. Production of Transgenic Pigs with an Introduced Missense Mutation of the Bone Morphogenetic Protein Receptor Type IB Gene Related to Prolificacy

    PubMed Central

    Zhao, Xueyan; Yang, Qiang; Zhao, Kewei; Jiang, Chao; Ren, Dongren; Xu, Pan; He, Xiaofang; Liao, Rongrong; Jiang, Kai; Ma, Junwu; Xiao, Shijun; Ren, Jun; Xing, Yuyun

    2016-01-01

    In the last few decades, transgenic animal technology has witnessed an increasingly wide application in animal breeding. Reproductive traits are economically important to the pig industry. It has been shown that the bone morphogenetic protein receptor type IB (BMPR1B) A746G polymorphism is responsible for the fertility in sheep. However, this causal mutation exits exclusively in sheep and goat. In this study, we attempted to create transgenic pigs by introducing this mutation with the aim to improve reproductive traits in pigs. We successfully constructed a vector containing porcine BMPR1B coding sequence (CDS) with the mutant G allele of A746G mutation. In total, we obtained 24 cloned male piglets using handmade cloning (HMC) technique, and 12 individuals survived till maturation. A set of polymerase chain reactions indicated that 11 of 12 matured boars were transgene-positive individuals, and that the transgenic vector was most likely disrupted during cloning. Of 11 positive pigs, one (No. 11) lost a part of the terminator region but had the intact promoter and the CDS regions. cDNA sequencing showed that the introduced allele (746G) was expressed in multiple tissues of transgene-positive offspring of No.11. Western blot analysis revealed that BMPR1B protein expression in multiple tissues of transgene-positive F1 piglets was 0.5 to 2-fold higher than that in the transgene-negative siblings. The No. 11 boar showed normal litter size performance as normal pigs from the same breed. Transgene-positive F1 boars produced by No. 11 had higher semen volume, sperm concentration and total sperm per ejaculate than the negative siblings, although the differences did not reached statistical significance. Transgene-positive F1 sows had similar litter size performance to the negative siblings, and more data are needed to adequately assess the litter size performance. In conclusion, we obtained 24 cloned transgenic pigs with the modified porcine BMPR1B CDS using HMC. c

  3. Production of Transgenic Pigs with an Introduced Missense Mutation of the Bone Morphogenetic Protein Receptor Type IB Gene Related to Prolificacy.

    PubMed

    Zhao, Xueyan; Yang, Qiang; Zhao, Kewei; Jiang, Chao; Ren, Dongren; Xu, Pan; He, Xiaofang; Liao, Rongrong; Jiang, Kai; Ma, Junwu; Xiao, Shijun; Ren, Jun; Xing, Yuyun

    2016-07-01

    In the last few decades, transgenic animal technology has witnessed an increasingly wide application in animal breeding. Reproductive traits are economically important to the pig industry. It has been shown that the bone morphogenetic protein receptor type IB (BMPR1B) A746G polymorphism is responsible for the fertility in sheep. However, this causal mutation exits exclusively in sheep and goat. In this study, we attempted to create transgenic pigs by introducing this mutation with the aim to improve reproductive traits in pigs. We successfully constructed a vector containing porcine BMPR1B coding sequence (CDS) with the mutant G allele of A746G mutation. In total, we obtained 24 cloned male piglets using handmade cloning (HMC) technique, and 12 individuals survived till maturation. A set of polymerase chain reactions indicated that 11 of 12 matured boars were transgene-positive individuals, and that the transgenic vector was most likely disrupted during cloning. Of 11 positive pigs, one (No. 11) lost a part of the terminator region but had the intact promoter and the CDS regions. cDNA sequencing showed that the introduced allele (746G) was expressed in multiple tissues of transgene-positive offspring of No.11. Western blot analysis revealed that BMPR1B protein expression in multiple tissues of transgene-positive F1 piglets was 0.5 to 2-fold higher than that in the transgene-negative siblings. The No. 11 boar showed normal litter size performance as normal pigs from the same breed. Transgene-positive F1 boars produced by No. 11 had higher semen volume, sperm concentration and total sperm per ejaculate than the negative siblings, although the differences did not reached statistical significance. Transgene-positive F1 sows had similar litter size performance to the negative siblings, and more data are needed to adequately assess the litter size performance. In conclusion, we obtained 24 cloned transgenic pigs with the modified porcine BMPR1B CDS using HMC. c

  4. A Simple Model for Learning Improvement: Weigh Pig, Feed Pig, Weigh Pig. Occasional Paper #23

    ERIC Educational Resources Information Center

    Fulcher, Keston H.; Good, Megan R.; Coleman, Chris M.; Smith, Kristen L.

    2014-01-01

    Assessing learning does not by itself result in increased student accomplishment, much like a pig never fattened up because it was weighed. Indeed, recent research shows that while institutions are more regularly engaging in assessment, they have little to show in the way of stronger student performance. This paper clarifies how assessment results…

  5. Local cloning of entangled states

    SciTech Connect

    Gheorghiu, Vlad; Yu Li; Cohen, Scott M.

    2010-08-15

    We investigate the conditions under which a set S of pure bipartite quantum states on a DxD system can be locally cloned deterministically by separable operations, when at least one of the states is full Schmidt rank. We allow for the possibility of cloning using a resource state that is less than maximally entangled. Our results include that: (i) all states in S must be full Schmidt rank and equally entangled under the G-concurrence measure, and (ii) the set S can be extended to a larger clonable set generated by a finite group G of order |G|=N, the number of states in the larger set. It is then shown that any local cloning apparatus is capable of cloning a number of states that divides D exactly. We provide a complete solution for two central problems in local cloning, giving necessary and sufficient conditions for (i) when a set of maximally entangled states can be locally cloned, valid for all D; and (ii) local cloning of entangled qubit states with nonvanishing entanglement. In both of these cases, we show that a maximally entangled resource is necessary and sufficient, and the states must be related to each other by local unitary 'shift' operations. These shifts are determined by the group structure, so need not be simple cyclic permutations. Assuming this shifted form and partially entangled states, then in D=3 we show that a maximally entangled resource is again necessary and sufficient, while for higher-dimensional systems, we find that the resource state must be strictly more entangled than the states in S. All of our necessary conditions for separable operations are also necessary conditions for local operations and classical communication (LOCC), since the latter is a proper subset of the former. In fact, all our results hold for LOCC, as our sufficient conditions are demonstrated for LOCC, directly.

  6. Pigs taking wing with transposons and recombinases

    PubMed Central

    Clark, Karl J; Carlson, Daniel F; Fahrenkrug, Scott C

    2007-01-01

    Swine production has been an important part of our lives since the late Mesolithic or early Neolithic periods, and ranks number one in world meat production. Pig production also contributes to high-value-added medical markets in the form of pharmaceuticals, heart valves, and surgical materials. Genetic engineering, including the addition of exogenous genetic material or manipulation of the endogenous genome, holds great promise for changing pig phenotypes for agricultural and medical applications. Although the first transgenic pigs were described in 1985, poor survival of manipulated embryos; inefficiencies in the integration, transmission, and expression of transgenes; and expensive husbandry costs have impeded the widespread application of pig genetic engineering. Sequencing of the pig genome and advances in reproductive technologies have rejuvenated efforts to apply transgenesis to swine. Pigs provide a compelling new resource for the directed production of pharmaceutical proteins and the provision of cells, vascular grafts, and organs for xenotransplantation. Additionally, given remarkable similarities in the physiology and size of people and pigs, swine will increasingly provide large animal models of human disease where rodent models are insufficient. We review the challenges facing pig transgenesis and discuss the utility of transposases and recombinases for enhancing the success and sophistication of pig genetic engineering. 'The paradise of my fancy is one where pigs have wings.' (GK Chesterton). PMID:18047690

  7. Treating Cloned Embryos, But Not Donor Cells, with 5-aza-2’-deoxycytidine Enhances the Developmental Competence of Porcine Cloned Embryos

    PubMed Central

    HUAN, Yan Jun; ZHU, Jiang; XIE, Bing Teng; WANG, Jian Yu; LIU, Shi Chao; ZHOU, Yang; KONG, Qing Ran; HE, Hong Bin; LIU, Zhong Hua

    2013-01-01

    The efficiency of cloning by somatic cell nuclear transfer (SCNT) has remained low. In most cloned embryos, epigenetic reprogramming is incomplete, and usually the genome is hypermethylated. The DNA methylation inhibitor 5-aza-2’-deoxycytidine (5-aza-dC) could improve the developmental competence of cow, pig, cat and human SCNT embryos in previous studies. However, the parameters of 5-aza-dC treatment among species are different, and whether 5-aza-dC could enhance the developmental competence of porcine cloned embryos has still not been well studied. Therefore, in this study, we treated porcine fetal fibroblasts (PFF) that then were used as donor nuclei for nuclear transfer or fibroblast-derived reconstructed embryos with 5-aza-dC, and the concentration- and time-dependent effects of 5-aza-dC on porcine cloned embryos were investigated by assessing pseudo-pronucleus formation, developmental potential and pluripotent gene expression of these reconstructed embryos. Our results showed that 5-aza-dC significantly reduced the DNA methylation level in PFF (0 nM vs. 10 nM vs. 25 nM vs. 50 nM, 58.70% vs. 37.37% vs. 45.43% vs. 39.53%, P<0.05), but did not improve the blastocyst rate of cloned embryos derived from these cells. Treating cloned embryos with 25 nM 5-aza-dC for 24 h significantly enhanced the blastocyst rate compared with that of the untreated group. Furthermore, treating cloned embryos, but not donor cells, significantly promoted pseudo-pronucleus formation at 4 h post activation (51% for cloned embryos treated, 34% for donor cells treated and 36% for control, respectively, P<0.05) and enhanced the expression levels of pluripotent genes (Oct4, Nanog and Sox2) up to those of in vitro fertilized embryos during embryo development. In conclusion, treating cloned embryos, but not donor cells, with 5-aza-dC enhanced the developmental competence of porcine cloned embryos by promotion of pseudo-pronucleus formation and improvement of pluripotent gene expression. PMID

  8. Testing of an intrathoracic artificial lung in a pig model.

    PubMed

    Cook, K E; Makarewicz, A J; Backer, C L; Mockros, L F; Przybylo, H J; Crawford, S E; Hernandez, J M; Leonard, R J; Mavroudis, C

    1996-01-01

    A low input impedance, intrathoracic artificial lung is being developed for use in acute respiratory failure or as a bridge to transplantation. The device uses microporous, hollow fibers in a 0.74 void fraction, 1.83 m2 surface area bundle. The bundle is placed within a thermoformed polyethylene terephthalate glucose modified housing with a gross volume of 800 cm3. The blood inlet and outlet are 18 mm inner diameter vascular grafts. Between the inlet graft and the device is a 1 inch inner diameter, thin-walled, latex tubing compliance chamber. These devices were implanted in Yorkshire pigs via median sternotomy with an end to side anastomosis to the pulmonary artery and left atrium. The distal pulmonary artery was occluded to divert the right ventricular output to the device. Pigs 1 and 2 were supported fully for 24 hrs and then killed. Pig 3 was supported partially for 20 hrs and died from bleeding complications. The first implant, in a 55 kg male pig, transferred an average of 176 ml/min +/- 42.4 of O2 and 190 ml/min +/- 39.7 of CO2 with an average blood flow rate of 2.71/min +/- 0.46. The normalized average right ventricular output power, Pn, was 0.062 W/(L/min) +/- 0.0082, and the average device resistance, R, was 3.5 mmHg/(L/min) +/- 0.62. The second implant, in a 60 kg male pig, transferred an average of 204 ml/min +/- 22.5 of O2 and 242 ml/min +/- 17.2 of CO2 with an average blood flow rate of 3.7 L/min +/- 0.45, Pn of 0.064 W/(L/min) +/- 0.0067, and R of 4.3 mmHg/(L/min) +/- 0.89. The third implant, in an 89 kg male pig, transferred an average of 156 ml/min +/- 39.6 of O2 and 187 ml/min +/- 21.4 of CO2 with an average blood flow rate of 2.5 L/min +/- 0.49, Pn of 0.052 W/(l/min) +/- 0.0067, and R of 3.4 mmHg/(L/min) +/- 0.74. These experiments suggest that such an artificial lung can temporarily support the gas transfer requirements of adult humans without over-loading the right ventricle.

  9. Cloning and characterization of a new swine MHC (SLA) class II DQB allele.

    PubMed

    Hosokawa, T; Tanioka, Y; Tanigawa, M; Matsumoto, Y; Onodera, T; Matsumoto, Y

    1998-06-01

    Major histocompatibility complex (MHC) of pigs is known as swine leukocyte antigen (SLA). The cDNA encoding a new allele of SLA class II DQ beta-chain was successfully isolated from a CSK miniature pig (derived from Göttingen strain) and characterized by sequence analyses. SLA-DQB cDNA fragment encoding beta 2-domain was amplified by reverse transcriptase-polymerase chain reaction using the sequences preserved in a various vertebrates as primers. Using non-radioisotope technique with the PCR product as a probe, cDNA clone G01 was isolated from a spleen cDNA library, and nucleotide sequence of this clone was determined. This clone encompassed a whole SLA-DQ beta-chain coding region, containing a total length of 1161 nucleotides with an open reading frame (ORF) of 786 nucleotides, 5' untranslated region of 15 nucleotides, and 3' untranslated region of 360 nucleotides ending with a canonical polyadenylation signal, followed by a poly A tail. Sequence comparisons of the ORF of this clone with those of known SLA-DQB genes confirmed that this clone is a new allele (SLA-DQB*G01). Phylogenetic analysis of the nucleotide sequences of swine, human, and murine MHC class II genes indicated that SLA-DQB was more similar to HLA-DQB1 than H-2A beta. Comparison of the nucleotide and deduced amino acid sequences among SLA-DQB alleles showed that the SLA-DQ beta-chain polymorphism was found almost in beta 1-domain which contains the antigenic peptide binding sites.

  10. Biodegradation of Pig Manure by the Housefly, Musca domestica: A Viable Ecological Strategy for Pig Manure Management

    PubMed Central

    Čičková, Helena; Pastor, Berta; Kozánek, Milan; Martínez-Sánchez, Anabel; Rojo, Santos; Takáč, Peter

    2012-01-01

    The technology for biodegradation of pig manure by using houseflies in a pilot plant capable of processing 500–700 kg of pig manure per week is described. A single adult cage loaded with 25,000 pupae produced 177.7±32.0 ml of eggs in a 15-day egg-collection period. With an inoculation ratio of 0.4–1.0 ml eggs/kg of manure, the amount of eggs produced by a single cage can suffice for the biodegradation of 178–444 kg of manure. Larval development varied among four different types of pig manure (centrifuged slurry, fresh manure, manure with sawdust, manure without sawdust). Larval survival ranged from 46.9±2.1%, in manure without sawdust, to 76.8±11.9% in centrifuged slurry. Larval development took 6–11 days, depending on the manure type. Processing of 1 kg of wet manure produced 43.9–74.3 g of housefly pupae and the weight of the residue after biodegradation decreased to 0.18–0.65 kg, with marked differences among manure types. Recommendations for the operation of industrial-scale biodegradation facilities are presented and discussed. PMID:22431982

  11. Development of large intestinal attaching and effacing lesions in pigs in association with the feeding of a particular diet.

    PubMed Central

    Neef, N A; McOrist, S; Lysons, R J; Bland, A P; Miller, B G

    1994-01-01

    Hysterotomy-derived piglets were kept in gnotobiotic isolators and artificially colonized at 7 days of age with an adult bovine enteric microflora. At 3 weeks of age, the pigs were transferred to conventional experimental accommodation and weaned, either onto a solid diet that had been associated with field cases of typhlocolitis in pigs or onto a solid control diet. At necropsy at 5 weeks of age, groups of pigs fed the diet associated with field cases of typhlocolitis were found to have developed typhlocolitis. This was absent from the groups fed the control diet. The typhlocolitis was characterized by attaching and effacing lesions typical of those described following experimental inoculation of various species with enteropathogenic Escherichia coli. A nonverocytotoxic, eae probe-positive E. coli serotype O116 was isolated from pigs on the colitis-associated diet but not from any of the pigs on the control diet. Coliform bacteria attached to the colonic lesions reacted with polyclonal antiserum to E. coli O116 in an immunoperoxidase assay of histological sections of affected tissue. No reaction with this antiserum was observed in corresponding tissue sections taken from pigs on the control diet. No colon lesions were observed in germfree pigs fed either of the diets. It is postulated that proliferation and possibly expression of pathogenicity of the attaching and effacing E. coli responsible for the lesions are strongly influenced by diet. Images PMID:7927691

  12. Detecting mitochondrial signatures of selection in wild Tibetan pigs and domesticated pigs.

    PubMed

    Li, Mingzhou; Jin, Long; Ma, Jideng; Tian, Shilin; Li, Ruiqiang; Li, Xuewei

    2016-01-01

    Selection in genomic regions is prevalent in mammals; however, the effects of selection on the mitogenome are not clearly understood. We determined the complete mitochondrial DNA (mtDNA) sequences from six wild Tibetan pigs from the Tibetan plateau and four domestic pig breeds from the lowland of neighboring southwest China. Nucleotide diversity analysis using the sliding window method showed that the nucleotide diversity of wild Tibetan pigs in most regions of the mitogenome was higher than that of domestic pigs. The 12 s ribosomal RNA showed relatively lower nucleotide diversity in Tibetan pigs, suggesting purifying selection of these genes during high-altitude adaptation. More non-synonymous nucleotide substitutions in the ATP6 were found in wild Tibetan pigs, indicating adaptive selection in Tibetan pigs. The results suggested distinct impacts of natural selection and artificial selection upon the mitogenome, especially the mitochondrial signatures of adaptive evolution in wild Tibetan pigs under natural selection.

  13. [Cloning and law in Hungary].

    PubMed

    Julesz, Máté

    2015-03-01

    Reproductive human cloning is prohibited in Hungary, as in many other countries. Therapeutic human cloning is not prohibited, just like in many other countries. Stem cell therapy is also allowed. Article III, paragraph (3) of the Hungarian basic law (constitution) strictly forbids total human cloning. Article 1 of the Additional Protocol to the Oviedo Convention, on the Prohibition of Cloning Human Beings (1998) stipulates that any intervention seeking to create a human being genetically identical to another human being, whether living or dead, is prohibited. In Hungary, according to Article 174 of the Criminal Code, total human cloning constitutes a crime. Article 180, paragraph (3) of the Hungarian Act on Health declares that embryos shall not be brought about for research purposes; research shall be conducted only on embryos brought about for reproductive purposes when this is authorized by the persons entitled to decide upon its disposal, or when the embryo is damaged. Article 180, paragraph (5) of the Hungarian Act on Health stipulates that multiple individuals who genetically conform to one another shall not be brought about. According to Article 181, paragraph (1) of the Hungarian Act on Health, an embryo used for research shall be kept alive for not longer than 14 days, not counting the time it was frozen for storage and the time period of research.

  14. [Mystery and problems of cloning].

    PubMed

    Nikitin, V A

    2010-01-01

    The attention of investigators is attracted to the fact that, in spite of great efforts in mammalian cloning, advances that have been made in this area of research are not great, and cloned animals have developmental pathologies often incompatible with life and/or reproduction ability. It is yet not clear what technical or biological factors underlie this, and how they are connected or interact with each other, which is more realistic strategically. There is a great number of articles dealing with the influence of cloning with the nuclear transfer on genetic and epigenetic reprogramming of donor cells. At the same time we can see the practical absence of analytical investigations concerning the technology of cloning as such, its weak points, and possible sources of cellular trauma in the course of microsurgery of nuclear transfer or twinning. This article discusses step by step several nuclear transfer techniques and the methods of dividing early preimplanted embryos for twinning with the aim to reveal possible sources of cell damage during micromanipulation that may have negative influence on the development of cloned organisms. Several new author's technologies based on the study of cell biophysical characteristics are described, which allow one to avoid cellular trauma during manipulation and minimize the possibility of cell damage at any rate.

  15. Bioactivation and Regioselectivity of Pig Cytochrome P450 3A29 towards Aflatoxin B₁.

    PubMed

    Wu, Jun; Chen, Ruohong; Zhang, Caihui; Li, Kangbai; Xu, Weiying; Wang, Lijuan; Chen, Qingmei; Mu, Peiqiang; Jiang, Jun; Wen, Jikai; Deng, Yiqun

    2016-01-01

    Due to unavoidable contaminations in feedstuff, pigs are easily exposed to aflatoxin B₁ (AFB₁) and suffer from poisoning, thus the poisoned products potentially affect human health. Heretofore, the metabolic process of AFB₁ in pigs remains to be clarified, especially the principal cytochrome P450 oxidases responsible for its activation. In this study, we cloned CYP3A29 from pig liver and expressed it in Escherichia coli, and its activity has been confirmed with the typical P450 CO-reduced spectral characteristic and nifedipine-oxidizing activity. The reconstituted membrane incubation proved that the recombinant CYP3A29 was able to oxidize AFB₁ to form AFB₁-exo-8,9-epoxide in vitro. The structural basis for the regioselective epoxidation of AFB₁ by CYP3A29 was further addressed. The T309A mutation significantly decreased the production of AFBO, whereas F304A exhibited an enhanced activation towards AFB₁. In agreement with the mutagenesis study, the molecular docking simulation suggested that Thr309 played a significant role in stabilization of AFB₁ binding in the active center through a hydrogen bond. In addition, the bulk phenyl group of Phe304 potentially imposed steric hindrance on the binding of AFB₁. Our study demonstrates the bioactivation of pig CYP3A29 towards AFB₁ in vitro, and provides the insight for understanding regioselectivity of CYP3A29 to AFB₁. PMID:27626447

  16. Bioactivation and Regioselectivity of Pig Cytochrome P450 3A29 towards Aflatoxin B1

    PubMed Central

    Wu, Jun; Chen, Ruohong; Zhang, Caihui; Li, Kangbai; Xu, Weiying; Wang, Lijuan; Chen, Qingmei; Mu, Peiqiang; Jiang, Jun; Wen, Jikai; Deng, Yiqun

    2016-01-01

    Due to unavoidable contaminations in feedstuff, pigs are easily exposed to aflatoxin B1 (AFB1) and suffer from poisoning, thus the poisoned products potentially affect human health. Heretofore, the metabolic process of AFB1 in pigs remains to be clarified, especially the principal cytochrome P450 oxidases responsible for its activation. In this study, we cloned CYP3A29 from pig liver and expressed it in Escherichia coli, and its activity has been confirmed with the typical P450 CO-reduced spectral characteristic and nifedipine-oxidizing activity. The reconstituted membrane incubation proved that the recombinant CYP3A29 was able to oxidize AFB1 to form AFB1-exo-8,9-epoxide in vitro. The structural basis for the regioselective epoxidation of AFB1 by CYP3A29 was further addressed. The T309A mutation significantly decreased the production of AFBO, whereas F304A exhibited an enhanced activation towards AFB1. In agreement with the mutagenesis study, the molecular docking simulation suggested that Thr309 played a significant role in stabilization of AFB1 binding in the active center through a hydrogen bond. In addition, the bulk phenyl group of Phe304 potentially imposed steric hindrance on the binding of AFB1. Our study demonstrates the bioactivation of pig CYP3A29 towards AFB1 in vitro, and provides the insight for understanding regioselectivity of CYP3A29 to AFB1. PMID:27626447

  17. Primary structure and functional expression of a guinea pig kappa opioid (dynorphin) receptor.

    PubMed Central

    Xie, G X; Meng, F; Mansour, A; Thompson, R C; Hoversten, M T; Goldstein, A; Watson, S J; Akil, H

    1994-01-01

    A full-length cDNA encoding the guinea pig kappa opioid (dynorphin) receptor has been isolated. The deduced protein contains 380 aa and seven hydrophobic alpha-helices characteristic of the G protein-coupled receptors. This receptor is 90% identical to the mouse and rat kappa receptors, with the greatest level of divergence in the N-terminal region. When expressed in COS-7 cells, the receptor displays high affinity and stereospecificity toward dynorphin peptides and other kappa-selective opioid ligands such as U50, 488. It does not bind the mu- and delta-selective opioid ligands. The expressed receptor is functionally coupled to G protein(s) to inhibit adenylyl cyclase and Ca2+ channels. The guinea pig kappa receptor mRNA is expressed in many brain areas, including the cerebellum, a pattern that agrees well with autoradiographic maps of classical guinea pig kappa binding sites. Species differences in the pharmacology and mRNA distribution between the cloned guinea pig and rat kappa receptors may be worthy of further examination. Images PMID:8170987

  18. 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

  19. Ontogenic increase in PGE2 and PGF2 alpha receptor density in brain microvessels of pigs.

    PubMed Central

    Li, D. Y.; Varma, D. R.; Chemtob, S.

    1994-01-01

    1. The hypothesis that the relative vasoconstrictor ineffectiveness of prostaglandin E2 (PGE2) and PGF2 alpha on cerebral vessels of newborn pigs might be due to fewer receptors for these prostanoids was tested by comparing receptors for PGE2 (EP) and PGF2 alpha (FP) in cerebral microvessels from newborn and adult pigs. 2. Specific binding of [3H]-PGE2 and [3H]-PGF2 alpha to membranes prepared from brain microvessels showed that EP and FP receptor density (Bmax) in tissues from newborn animals was less than 50% of that determined in tissues from adults. By contrast, estimates of affinity (KD) were unchanged. 3. Specifically bound [3H]-PGE2 to brain microvessels from both the newborn and adult was displaced by AH 6809 (EP1-selective antagonist) by 80-90%, and only by approximately 30-35% by both 11-deoxy PGE1 (EP2/EP3 agonist) and M&B 28,767 (EP3 agonist); butaprost (EP2 agonist) was completely ineffective. 4. PGE2, 17-phenyl trinor PGE2 (EP1 agonist), PGF2 alpha and fenprostalene (PGF2 alpha analogue) caused significantly less increase in inositol 1,4,5-triphosphate (IP3) in brain microvessels from the newborn than in those from adult pigs. The stimulation of IP3 by PGE2 and 17-phenyl trinor PGE2 was almost completely inhibited by the EP1 antagonist, AH 6809. 5. PGE2, 11-deoxy PGE1 and M&B 28,767 produced small reduction of adenosine 3':5'-cyclic monophosphate (cyclic AMP) production in adult vessels but no effect in newborn tissues. 6. The lower density of EP and FP receptors in microvessels of newborn pigs compared to adults may explain the reduced ability of PGE2 and PGF2 alpha to stimulate production of IP3 in tissues from newborn animals.(ABSTRACT TRUNCATED AT 250 WORDS) PMID:8032662

  20. The topsy-turvy cloning law.

    PubMed

    Brassington, Iain; Oultram, Stuart

    2011-03-01

    In debates about human cloning, a distinction is frequently drawn between therapeutic and reproductive uses of the technology. Naturally enough, this distinction influences the way that the law is framed. The general consensus is that therapeutic cloning is less morally problematic than reproductive cloning--one can hold this position while holding that both are morally unacceptable--and the law frequently leaves the way open for some cloning for the sake of research into new therapeutic techniques while banning it for reproductive purposes. We claim that the position adopted by the law has things the wrong way around: if we accept a moral distinction between therapeutic and reproductive cloning, there are actually more reasons to be morally worried about therapeutic cloning than about reproductive cloning. If cloning is the proper object of legal scrutiny, then, we ought to make sure that we are scrutinising the right kind of clone.

  1. Decomposition Rate and Pattern in Hanging Pigs.

    PubMed

    Lynch-Aird, Jeanne; Moffatt, Colin; Simmons, Tal

    2015-09-01

    Accurate prediction of the postmortem interval requires an understanding of the decomposition process and the factors acting upon it. A controlled experiment, over 60 days at an outdoor site in the northwest of England, used 20 freshly killed pigs (Sus scrofa) as human analogues to study decomposition rate and pattern. Ten pigs were hung off the ground and ten placed on the surface. Observed differences in the decomposition pattern required a new decomposition scoring scale to be produced for the hanging pigs to enable comparisons with the surface pigs. The difference in the rate of decomposition between hanging and surface pigs was statistically significant (p=0.001). Hanging pigs reached advanced decomposition stages sooner, but lagged behind during the early stages. This delay is believed to result from lower variety and quantity of insects, due to restricted beetle access to the aerial carcass, and/or writhing maggots falling from the carcass.

  2. Clonal distribution of Streptococcus suis isolated from diseased pigs in the central region of Chile

    PubMed Central

    Morales, Bárbara; Ruiz, Álvaro; Lacouture, Sonia; Gottschalk, Marcelo

    2015-01-01

    The characteristics of 29 Chilean field strains of Streptococcus suis recovered between 2007 and 2011 from pigs with clinical signs at different farms were studied. Serotyping with use of the coagglutination test revealed that all but 1 strain belonged to serotype 6; the remaining strain was serotype 22. All the serotype-6 strains were suilysin (hemolysin)-negative; in addition, they were found to be genotypically homogeneous by enterobacterial repetitive intergenic consensus sequence-based polymerase chain reaction (ERIC-PCR) and sensitive to ampicillin, ceftiofur, penicillin, and trimethoprim/sulfamethoxazole. The results indicate that, in contrast to what is generally observed in other countries, a single clone of S. suis was isolated from diseased pigs in the central region of Chile. PMID:26424917

  3. Zeta-crystallin, a novel protein from the guinea pig lens is related to alcohol dehydrogenases.

    PubMed

    Rodokanaki, A; Holmes, R K; Borrás, T

    1989-05-30

    zeta-Crystallin is a major component of the water-soluble proteins of the guinea pig lens. We have constructed a lens cDNA library from one- to seven-day-old guinea pigs in the plasmid Bluescript KS+ and used the 16 amino acid (aa) sequence of a CNBr peptide to design an oligodeoxyribonucleotide probe. Analysis of two positive clones and direct sequence of the 5' end of the RNA resulted in the completion of a most probably full-length mRNA comprising 1842 nucleotides (nt). The ATG start codon occurs 83 nt downstream from the 5' end. The open reading frame, ending with a stop codon at nt position 1070, predicts a protein of 328 aa with a calculated Mr of 35,071. Comparison of the amino acid sequence with the National Biomedical Research Foundation protein data base reveals a significant similarity of zeta-crystallin with the enzyme of the alcohol dehydrogenase family.

  4. Assessing learning and memory in pigs.

    PubMed

    Gieling, Elise Titia; Nordquist, Rebecca Elizabeth; van der Staay, Franz Josef

    2011-03-01

    In recent years, there has been a surge of interest in (mini) pigs (Sus scrofa) as species for cognitive research. A major reason for this is their physiological and anatomical similarity with humans. For example, pigs possess a well-developed, large brain. Assessment of the learning and memory functions of pigs is not only relevant to human research but also to animal welfare, given the nature of current farming practices and the demands they make on animal health and behavior. In this article, we review studies of pig cognition, focusing on the underlying processes and mechanisms, with a view to identifying. Our goal is to aid the selection of appropriate cognitive tasks for research into pig cognition. To this end, we formulated several basic criteria for pig cognition tests and then applied these criteria and knowledge about pig-specific sensorimotor abilities and behavior to evaluate the merits, drawbacks, and limitations of the different types of tests used to date. While behavioral studies using (mini) pigs have shown that this species can perform learning and memory tasks, and much has been learned about pig cognition, results have not been replicated or proven replicable because of the lack of validated, translational behavioral paradigms that are specially suited to tap specific aspects of pig cognition. We identified several promising types of tasks for use in studies of pig cognition, such as versatile spatial free-choice type tasks that allow the simultaneous measurement of several behavioral domains. The use of appropriate tasks will facilitate the collection of reliable and valid data on pig cognition.

  5. Erysipelas in turkeys, sheep and pigs.

    PubMed

    2015-03-21

    Erysipelas diagnosed in turkeys, sheep and pigs. Parasitic gastroenteritis reported in cattle on several farms. Unusual presentation of Actinobacillus suis causing spinal abscesses in pigs on a breeder-finisher unit. First APHA diagnosis of oedema disease in pigs in East Anglia for many years. Infectious coryza confirmed in a hobby breeding flock. These are among matters discussed in the Animal and Plant Health Agency's (APHA's) disease surveillance report for November 2014.

  6. Cryptosporidium parvum pig genotype II diagnosed in pigs from the state of Rio de Janeiro, Brazil

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Pigs may represent a source of Cryptosporidium sp. infection to humans. The objective of this study was to identify the species present in pigs from the State of Rio de Janeiro, Brazil, and verify what risks pigs represent in transmission of human cryptosporidiosis, since there is no such informati...

  7. Association between microbiological and serological prevalence of human pathogenic Yersinia spp. in pigs and pig batches.

    PubMed

    Vanantwerpen, Gerty; Berkvens, Dirk; De Zutter, Lieven; Houf, Kurt

    2015-07-01

    Pigs are the main reservoir of human pathogenic Y. enterocolitica, and the microbiological and serological prevalence of this pathogen differs between pig farms. The infection status of pig batches at moment of slaughter is unknown while it is a possibility to classify batches. A relation between the presence of human pathogenic Yersinia spp. and the presence of antibodies could help to predict the infection of the pigs prior to slaughter. Pigs from 100 different batches were sampled. Tonsils and pieces of diaphragm were collected from 7047 pigs (on average 70 pigs per batch). The tonsils were analyzed using a direct plating method and the meat juice collected from the pieces of diaphragm was analyzed by Enzyme Linked ImmunoSorbent Assay. The microbiological and serological results were compared using a mixed-effects logistic regression at pig and batch level. Yersinia spp. were found in 2031 (28.8%) pigs, antibodies were present in 4692 (66.6%) pigs. According to the logistic regression, there was no relation at pig level between the presence of Yersinia spp. in tonsils and the presence of antibodies. Contrarily, at batch level, a mean activity value of 37 Optical Density (OD)% indicated a Yersinia spp. positive farm and the microbiological prevalence in pig batches could be estimated before shipment to the slaughterhouse. This offers the opportunity to classify batches based on their potential risk to contaminate carcasses with human pathogenic Yersinia spp.

  8. Diversity of forensic rove beetles (Coleoptera, Staphylinidae) associated with decaying pig carcass in a forest biotope.

    PubMed

    Dekeirsschieter, Jessica; Frederick, Christine; Verheggen, Francois J; Drugmand, Didier; Haubruge, Eric

    2013-07-01

    Most forensic studies are focused on Diptera pattern colonization while neglecting Coleoptera succession. So far, little information is available on the postmortem colonization by beetles and the decomposition process they initiate under temperate biogeoclimatic countries. These beetles have, however, been referred to as being part of the entomofaunal colonization of a dead body. Forensic entomologists need increased databases detailing the distribution, ecology, and phenology of necrophagous insects, including staphylinids (Coleoptera, Staphylinidae). While pig carcasses are commonly used in forensic entomology studies to surrogate human decomposition and to investigate the entomofaunal succession, very few works have been conducted in Europe on large carcasses. Our work reports the monitoring of the presence of adult rove beetles (Coleoptera, Staphylinidae) on decaying pig carcasses in a forest biotope during four seasons (spring, summer, fall, and winter). A total of 23 genera comprising 60 species of rove beetles were collected from pig carcasses.

  9. Naturally occurring Parelaphostrongylus tenuis-associated choriomeningitis in a guinea pig with neurologic signs.

    PubMed

    Southard, T; Bender, H; Wade, S E; Grunenwald, C; Gerhold, R W

    2013-05-01

    An adult male guinea pig (Cavia porcellus) with a 1-month history of hind limb paresis, torticollis, and seizures was euthanized and submitted for necropsy. Gross examination was unremarkable, but histologic examination revealed multifocal eosinophilic and lymphoplasmacytic choriomeningitis and cross sections of nematode parasites within the leptomeninges of the midbrain and diencephalon. Morphologic features of the nematode were consistent with a metastrongyle, and the parasite was identified as Parelaphostrongylus tenuis by polymerase chain reaction testing and nucleotide sequencing. Further questioning of the owner revealed that the guinea pig was fed grass from a yard often grazed by white-tailed deer (Odocoileus virginianus). To the authors' knowledge, this is the first report of a naturally occurring P. tenuis infection in a guinea pig.

  10. Worm recovery and precipitin antibody response in guinea pigs and rats infected with Clonorchis sinensis.

    PubMed

    Su, K E; Wang, F Y; Chi, P Y

    1998-12-01

    Guinea pigs (Hartley strain) and rats (Wistar strain) were each fed 200 and 100 Clonorchis sinensis metacercariae, respectively. Five animals from each species were sacrificed weekly between 1-8 weeks postinfection (WPI) and then at 12, 16, 20 and 30 WPI for collection of worms, bile and sera. The overall worm recovery rates for guinea pigs and rats were 18.7% and 12.4%, respectively. Only one of the five rats examined at 20 WPI still harbored one worm, while all were worm-free at 30 WPI. By a double diffusion test, no antibodies were detected against C. sinensis adult antigens in the bile juice. Serum antibodies were detected in at least 95% of the infected guinea pigs between 4-30 WPI and rats between 3-16 WPI. Precipitin antibodies seemed to be correlated with the presence of live worms in rats that had been infected for more than 12 weeks.

  11. Diversity of forensic rove beetles (Coleoptera, Staphylinidae) associated with decaying pig carcass in a forest biotope.

    PubMed

    Dekeirsschieter, Jessica; Frederick, Christine; Verheggen, Francois J; Drugmand, Didier; Haubruge, Eric

    2013-07-01

    Most forensic studies are focused on Diptera pattern colonization while neglecting Coleoptera succession. So far, little information is available on the postmortem colonization by beetles and the decomposition process they initiate under temperate biogeoclimatic countries. These beetles have, however, been referred to as being part of the entomofaunal colonization of a dead body. Forensic entomologists need increased databases detailing the distribution, ecology, and phenology of necrophagous insects, including staphylinids (Coleoptera, Staphylinidae). While pig carcasses are commonly used in forensic entomology studies to surrogate human decomposition and to investigate the entomofaunal succession, very few works have been conducted in Europe on large carcasses. Our work reports the monitoring of the presence of adult rove beetles (Coleoptera, Staphylinidae) on decaying pig carcasses in a forest biotope during four seasons (spring, summer, fall, and winter). A total of 23 genera comprising 60 species of rove beetles were collected from pig carcasses. PMID:23550535

  12. Hygiene and Welfare Evaluation of Pigs Slaughtered in Agritourisms

    PubMed Central

    Piras, Francesca; Agus, Vanessa; Porcheddu, Gabriella; Fois, Giuseppe; Consolati, Simonetta Gianna

    2015-01-01

    The slaughtering procedures at agritourism farms must be carried out in accordance with the general and hygiene requirements of Regulations (EC) No 852 and 853/2004. In addition, regional laws define minimum requirements allowing some flexibility. Piglets and finishing pigs are the most frequently slaughtered animal in Sardinian agritourism farms. The aim of the present survey was to evaluate: the general and hygiene requirements of outbuilding slaughterhouses in agritourisms; the animal welfare indicators; the microbial contamination of piglets and finishing pigs carcasses. Six agritourisms outbuilding slaughterhouses – EU-approved – were investigated. General and hygiene requirements of outbuilding slaughterhouses and animal welfare indicators of 68 piglets and 5 finishing pigs were evaluated by mean of a checklist. The following parameters were determined on 45 piglets and 5 finishing pigs carcasses: i) pH 1 and 24 h after slaughter, and ii) carcass surface microbial contamination by non destructive method (sponge) on the following sampling sites: ham; back (adults); belly; jowl (adults). Aerobic colony count (ACC; ISO 4833:2003), Enterobacteriaceae (EB; ISO 21528-2:2004), Salmonella spp. (ISO 6579:2002), Listeria monocytogenes (ISO 11290-1:1996 and 11290-2:1998) were also tested. All the plants except one have two separate rooms, for clean and dirty zones, stunning and bleeding operations being frequently carried out on open air. The piglet scalding was carried out in hot water bowls, and hair removal by singeing. Animal welfare signs revealed the following aspects: handling: hoisting prior to stunning, vocalizations (41%); stunning: not individual access to box, repeated shocks (4%), mean voltage 135.6 V, mean current for head-only electrical stunning 0.78 A; indicators of not effective stunning: palpebral reflex (24.2%), corneal reflex (12.8%), vocalizations (15.4%); bleeding: conscious and sensitive animal shackling (53.8%). Results of carcass evaluation

  13. A cross-sectional study of hepatitis E virus infection in healthy people directly exposed and unexposed to pigs in a rural community in northern Thailand.

    PubMed

    Hinjoy, S; Nelson, K E; Gibbons, R V; Jarman, R G; Mongkolsirichaikul, D; Smithsuwan, P; Fernandez, S; Labrique, A B; Patchanee, P

    2013-12-01

    A cross-sectional study of the association between occupational pig exposure and hepatitis E virus (HEV) infection in adult pig farmers and the general population who were not directly exposed to pigs was conducted in Nan Province, Thailand, from November 2010 to April 2011. All participants were interviewed to provide information on their job history, eating habits and other potential confounders. The prevalence of anti-HEV immunoglobulin G antibodies (IgG) among 513 subjects was 23.0%. Hand washing with water and soap was associated with a lower seroprevalence of HEV infection, whereas living in an area with frequent flooding (OR 1.64, 95% CI: 1.00-2.68) and consuming internal pig organs more than twice per week (OR 3.23, 95%CI: 1.15-9.01) were both associated with a higher seroprevalence of anti-HEV IgG. There was no association between HEV seroprevalence and frequent, direct occupational pig contact.

  14. The economic and environmental value of genetic improvements in fattening pigs: An integrated dynamic model approach.

    PubMed

    Niemi, J K; Sevón-Aimonen, M-L; Stygar, A H; Partanen, K

    2015-08-01

    The selection of animals for improved performance affects the profitability of pig fattening and has environmental consequences. The goal of this paper was to examine how changes in genetic and market parameters impact the biophysical (feeding patterns, timing of slaughter, nitrogen excretion) and economic (return per pig space unit) results describing pig fattening in a Finnish farm. The analysis can be viewed as focusing on terminal line breeding goals. An integrated model using recursive stochastic dynamic programming and a biological pig growth model was used to estimate biophysical results and economic values. Combining these models allowed us to provide more accurate estimates for the value of genetic improvement and, thus, provide better feedback to animal breeding programs than the traditional approach, which is based on fixed management patterns. Besides the benchmark scenario, the results were simulated for 5 other scenarios. In each scenario, genotype was improved regarding daily growth potential, carcass lean meat content, or the parameters of the Gompertz growth curve (maturing rate [], adult weight of protein [α], and adult weight of lipid mass []). The change in each parameter was equal to approximately 1 SD genetic improvement (ceteris paribus). Increasing , , daily growth potential, or carcass lean meat content increased the return on pig space unit by €12.60, €7.60, €4.10, or €2.90 per year, respectively, whereas an increase in decreased the return by €3.10. The genetic improvement in and resulted in the highest decrease in nitrogen excretion calculated in total or per kilogram of carcass gain but only under the optimal feeding pattern. Simulated changes in the Gompertz growth function parameters imply greater changes in ADG and lean meat content than changes in scenarios focusing on improving ADG and lean meat content directly. The economic value of genetic improvements as well as the quantity of nitrogen excreted during the fattening

  15. The economic and environmental value of genetic improvements in fattening pigs: An integrated dynamic model approach.

    PubMed

    Niemi, J K; Sevón-Aimonen, M-L; Stygar, A H; Partanen, K

    2015-08-01

    The selection of animals for improved performance affects the profitability of pig fattening and has environmental consequences. The goal of this paper was to examine how changes in genetic and market parameters impact the biophysical (feeding patterns, timing of slaughter, nitrogen excretion) and economic (return per pig space unit) results describing pig fattening in a Finnish farm. The analysis can be viewed as focusing on terminal line breeding goals. An integrated model using recursive stochastic dynamic programming and a biological pig growth model was used to estimate biophysical results and economic values. Combining these models allowed us to provide more accurate estimates for the value of genetic improvement and, thus, provide better feedback to animal breeding programs than the traditional approach, which is based on fixed management patterns. Besides the benchmark scenario, the results were simulated for 5 other scenarios. In each scenario, genotype was improved regarding daily growth potential, carcass lean meat content, or the parameters of the Gompertz growth curve (maturing rate [], adult weight of protein [α], and adult weight of lipid mass []). The change in each parameter was equal to approximately 1 SD genetic improvement (ceteris paribus). Increasing , , daily growth potential, or carcass lean meat content increased the return on pig space unit by €12.60, €7.60, €4.10, or €2.90 per year, respectively, whereas an increase in decreased the return by €3.10. The genetic improvement in and resulted in the highest decrease in nitrogen excretion calculated in total or per kilogram of carcass gain but only under the optimal feeding pattern. Simulated changes in the Gompertz growth function parameters imply greater changes in ADG and lean meat content than changes in scenarios focusing on improving ADG and lean meat content directly. The economic value of genetic improvements as well as the quantity of nitrogen excreted during the fattening

  16. Current status of pig liver xenotransplantation.

    PubMed

    Ekser, Burcin; Markmann, James F; Tector, A Joseph

    2015-11-01

    The shortage of organs from deceased human donors is a major problem limiting the number of organs transplanted each year and results in the death of thousands of patients on the waiting list. Pigs are currently the preferred species for clinical organ xenotransplantation. Progress in genetically-engineered (GE) pig liver xenotransplantation increased graft and recipient survival from hours with unmodified pig livers to up to 9 days with normal to near-normal liver function. Deletion of genes such as GGTA1 (Gal-knockout pigs) or adding genes such as human complement regulatory proteins (hCD55, hCD46 expressing pigs) enabled hyperacute rejection to be overcome. Although survival up to 9 days was recorded, extended pig graft survival was not achieved due to lethal thrombocytopenia. The current status of GE pig liver xenotransplantation with world experience, potential factors causing thrombocytopenia, new targets on pig endothelial cells, and novel GE pigs with more genes deletion to avoid remaining antibody response, such as beta1,4-N-acetyl galactosaminyl transferase 2 (β4GalNT2), are discussed.

  17. Gastric zygomycosis (mucormycosis) in 4 suckling pigs.

    PubMed

    Sanford, S E

    1985-02-15

    Acute gastric zygomycosis (mucormycosis) was diagnosed in four 6- to 7-day-old pigs with large venous infarcts in the gastric fundus. Two pigs were from one farm where several dams had developed fever at parturition and most of their litters had died. The other 2 pigs, from separate farms, had diarrhea that was unresponsive to broad-spectrum antibiotic therapy. Histologically there was severe hemorrhagic, ulcerative gastritis associated with numerous transmurally invading, mucoraceous fungi. The discussion includes speculation on the pathogenesis of this lesion in neonatal pigs.

  18. 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.

  19. Health status of cloned cattle at different ages.

    PubMed

    Chavatte-Palmer, P; Remy, D; Cordonnier, N; Richard, C; Issenman, H; Laigre, P; Heyman, Y; Mialot, J-P

    2004-01-01

    The procedure of somatic cloning is associated with important losses during pregnancy and in the perinatal period, reducing the overall efficacy to less than 5% in most cases. A mean of 30% of the cloned calves die before reaching 6 months of age with a wide range of pathologies, including, for the most common, respiratory failure, abnormal kidney development, liver steatosis. Heart and liver weight in relation to body weight are also increased. Surviving animals, although mostly clinically normal, differ from controls obtained by artificial insemination (AI) within the first 1-2 months, to become undistinguishable from them thereafter. Hemoglobin concentrations, for instance, are lower, and leptin concentrations are elevated. In response to the lack of prospective studies addressing the health of adult clones, a long-term, 3-4-year study is currently being conducted to assess the health of mature bovine clones at INRA. Preliminary results over 1 year of study do not show any statistical difference between groups for hematological parameters. PMID:15268782

  20. 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

  1. Prenatal ontogeny of lymphocyte subpopulations in pigs.

    PubMed Central

    Sinkora, M; Sinkora, J; Reháková, Z; Splíchal, I; Yang, H; Parkhouse, R M; Trebichavsk, I

    1998-01-01

    Although porcine lymphocytes have been classified into numerous subpopulations in postnatal animals, little is known about the ontogeny of these complex cell subsets. Using double- and triple-colour flow cytometry (FCM), we investigated the surface phenotype of fetal lymphoid cells in the thymus, cord blood, spleen and mesenteric lymph nodes at different stages of gestation. It was found that the major lymphocyte subpopulations started to appear at the beginning of the second third of the gestation period, with B cells being the earliest lymphocyte subpopulation to appear in the periphery. The T-cell receptor (TCR) gamma delta+ cells were the earliest detectable T-cell subset, developing first in the thymus and subsequently arriving in the periphery. Later in ontogeny, however, the number of TCRalpha beta+ lymphocytes rapidly increased, becoming the predominant T cells both in the thymus and in the periphery. Cells with the phenotype of adult natural killer cells were also identified in pig fetuses, though their nature and functional roles remain to be investigated. In addition, CD2 was expressed on most B cells whilst very few CD4+ TCRalpha beta+ cells or CD2+ TCRgamma delta+ cells expressed CD8, suggesting that the expression of CD2 and CD8 may reflect the functional status of the cells in postnatal animals. Taken together, this study has provided a systematic analysis of fetal porcine lymphocyte subpopulations and may provide the base for studies to establish the physiological roles of these lymphocyte subsets. PMID:9893051

  2. Adult Books for Young Adults.

    ERIC Educational Resources Information Center

    Carter, Betty

    1997-01-01

    Considers the differences between young adult and adult books and maintains that teachers must be familiar with young adults' tastes for both. Suggests that traffic between these publishing divisions is a two-way street, with young adults reading adult books and adults reading young adult books. (TB)

  3. Characterization of smallholder pig breeding practices within a rural commune of North Central Vietnam.

    PubMed

    Ieda, Nahoko; Van Bui, Quang; Nguyen, Nga Thi Duong; Lapar, Lucy; Marshall, Karen

    2015-08-01

    This case study focused on a pig production system in a rural area of North Central Vietnam, with a focus on describing household pig breeding practices and estimating herd demographic parameters, particularly on reproduction. One hundred five households undertaking small-scale piglet production were surveyed, with information gathered on 3268 individual pigs. Pig keeping contributed variably to the overall household livelihood portfolio, with female household members as the main decision makers, contributors to labor, and beneficiaries of income from the pig enterprise. All households kept between one and four young or adult sows, with 69% of these sows of a local breed type (predominantly Mong Cai), 28% a cross between a local sow and an exotic sow (predominantly Large White), and the remainder (3%) as exotic sows. Eighty-eight percent of the piglets produced were cross-bred, while 12% were local breed. No adult males were kept by the surveyed households, reflecting the common use of artificial insemination for mating purposes. The most common breeding system practiced-the keeping of Mong Cai females and production of cross-bred piglets-capitalizes on the small body size and high fecundity of the sows and the fast growth rate and leanness of the cross-bred piglets. The survey tool used, which was based on farmer recall of events over the preceding 12-month period, appeared to give reasonable results although some recall bias could be detected. This case study will serve as an entry point to planned broader scale characterization and development of pig breeding systems in North Central Vietnam.

  4. Ascaris suum infections in pigs born and raised on contaminated paddocks.

    PubMed

    Mejer, H; Roepstorff, A

    2006-09-01

    The transmission of Ascaris suum was studied in outdoor reared pigs. From May to June 2001, 6 farrowing paddocks were naturally contaminated with A. suum using experimentally infected seeder pigs. Early July, 1 sow farrowed on each paddock. One piglet per litter was slaughtered every second week starting at week 3 post-partum (p.p.) for registration of liver white spots and recovery of A. suum from the lungs and the small intestine. The last pigs were slaughtered at week 19 p.p. Faeces was examined for parasite eggs and blood was analysed for A. suum-specific antibodies. Weaning took place at week 7 p.p. by removing the sow. Paddock infection levels were estimated by regular examination of soil samples and in late June and late November using parasite naïve tracer pigs. Paddock contamination was high but eggs developed slowly resulting in a low initial transmission to the experimental pigs. By week 5 p.p. transmission had increased and the numbers of infective eggs in the soil increased during the study. The results indicate a continuous uptake of infective eggs, but visceral larval migration was reduced with time, probably due to the development of a pre-hepatic barrier. Nevertheless, a rather large population of adult worms remained in the pigs throughout the study, and it may primarily have been eggs ingested in the early infection phase that gave rise to the patent infections. It is suggested that neonatal exposure may result in increased persistence and size of adult worm burden and that the higher 'life-time worm burden' may be of significant economic importance. PMID:16740179

  5. Movements of wild pigs in Louisiana and Mississippi, 2011-13

    USGS Publications Warehouse

    Hartley, Stephen B.; Goatcher, Buddy L.; Sapkota, Sijan

    2015-01-01

    The prolific breeding capability, behavioral adaptation, and adverse environmental impacts of invasive wild pigs (Sus scrofa) have increased efforts towards managing their populations and understanding their movements. Currently, little is known about wild pig populations and movements in Louisiana and Mississippi. From 2011 to 2013, the U.S. Geological Survey investigated spatial and temporal movements of wild pigs in both marsh and nonmarsh physiographic regions. Twenty-one Global Positioning System satellite telemetry tracking collars were installed on adult wild pigs captured with trained dogs and released. Coordinates of their locations were recorded hourly. We collected 16,674 hourly data points including date, time, air temperature, and position during a 3-year study. Solar and lunar attributes, such as sun and moon phases and azimuth angles, were not related significantly to the movements among wild pigs. Movements were significantly correlated negatively with air temperature. Differences in movements between seasons and years were observed. On average, movements of boars were significantly greater than those of sows. Average home range, determined by using a minimum convex polygon as a proxy, was 911 hectares for boars, whereas average home range for sows was 116 hectares. Wild pigs in marsh habitat traveled lesser distances relative to those from more arid, nonmarsh habitats. Overall, results of this study indicate that wild pigs in Louisiana and Mississippi have small home ranges. These small home ranges suggest that natural movements have not been a major factor in the recent broad-scale range expansion observed in this species in the United States.

  6. Foot and mouth disease virus transmission among vaccinated pigs after exposure to virus shedding pigs.

    PubMed

    Orsel, K; de Jong, M C M; Bouma, A; Stegeman, J A; Dekker, A

    2007-08-21

    The aim of this study was to design a transmission experiment that enabled quantification of the effectiveness of vaccination against foot and mouth disease (FMD) virus in groups of pigs. Previous experiments showed that intradermal injection of pigs with FMD virus 14 days after vaccination was not suitable to start an infection chain, as inoculated vaccinated pigs resisted challenge. Therefore, we carried out two experiments in which we used direct contact to a non-vaccinated pig as route of infection. In the first experiment only the vaccine effect on susceptibility was quantified by exposing pigs, either vaccinated 14 days before or not vaccinated, each to a non-vaccinated seeder pig inoculated with FMD virus O/NET/2001. Since no significant differences were observed between contact infections in vaccinated or non-vaccinated pigs, we performed a second experiment in which both susceptibility and infectivity were subject to vaccination. We quantified virus transmission in homogenous groups of vaccinated or non-vaccinated pigs in which the infection chain was started by exposure to a third group of non-vaccinated infected pigs. Transmission occurred to all contact-exposed pigs in the non-vaccinated groups and to 9 out of 10 contact-exposed pigs in the vaccinated groups. The rate of transmission (beta) was significantly reduced in the vaccine group. Yet, the estimated reproduction ratio in both groups was still above 1. In conclusion, by adjusting our transmission study design and challenge method, we were able to quantify transmission of FMDV among vaccinated pigs. According to this study a single vaccination was not sufficient to stop pig to pig virus transmission. With these results major outbreaks may still be expected, even in groups of vaccinated pigs. PMID:17658199

  7. Study of insect succession and rate of decomposition on a partially burned pig carcass in an oil palm plantation in Malaysia.

    PubMed

    Heo, Chong Chin; Mohamad, Abdullah Marwi; Ahmad, Firdaus Mohd Salleh; Jeffery, John; Kurahashi, Hiromu; Omar, Baharudin

    2008-12-01

    Insects found associated with corpse can be used as one of the indicators in estimating postmortem interval (PMI). The objective of this study was to compare the stages of decomposition and faunal succession between a partially burnt pig (Sus scrofa Linnaeus) and natural pig (as control). The burning simulated a real crime whereby the victim was burnt by murderer. Two young pigs weighed approximately 10 kg were used in this study. Both pigs died from pneumonia and immediately placed in an oil palm plantation near a pig farm in Tanjung Sepat, Selangor, Malaysia. One pig was partially burnt by 1-liter petrol while the other served as control. Both carcasses were visited twice per day for the first week and once thereafter. Adult flies and larvae on the carcasses were collected and later processed in a forensic entomology laboratory. Results showed that there was no significant difference between the rate of decomposition and sequence of faunal succession on both pig carcasses. Both carcasses were completely decomposed to remain stage after nine days. The species of flies visiting the pig carcasses consisted of blow flies (Chrysomya megacephala, Chrysomya rufifacies, Hemipyrellia ligurriens), flesh fly (Sarcophagidae.), muscid fly (Ophyra spinigera), soldier fly (Hermetia illucens), coffin fly (Phoridae) and scavenger fly (Sepsidae). The only difference noted was in the number of adult flies, whereby more flies were seen in the control carcass. Faunal succession on both pig carcasses was in the following sequence: Calliphoridae, Sarcophagidae, Muscidae, Phoridae and lastly Stratiomyidae. However, there was overlap in the appearance of members of these families. Blowflies continued to oviposit on both carcasses. Hence postmortem interval (PMI) can still be estimated from the partially burnt pig carcass. PMID:19287358

  8. Study of insect succession and rate of decomposition on a partially burned pig carcass in an oil palm plantation in Malaysia.

    PubMed

    Heo, Chong Chin; Mohamad, Abdullah Marwi; Ahmad, Firdaus Mohd Salleh; Jeffery, John; Kurahashi, Hiromu; Omar, Baharudin

    2008-12-01

    Insects found associated with corpse can be used as one of the indicators in estimating postmortem interval (PMI). The objective of this study was to compare the stages of decomposition and faunal succession between a partially burnt pig (Sus scrofa Linnaeus) and natural pig (as control). The burning simulated a real crime whereby the victim was burnt by murderer. Two young pigs weighed approximately 10 kg were used in this study. Both pigs died from pneumonia and immediately placed in an oil palm plantation near a pig farm in Tanjung Sepat, Selangor, Malaysia. One pig was partially burnt by 1-liter petrol while the other served as control. Both carcasses were visited twice per day for the first week and once thereafter. Adult flies and larvae on the carcasses were collected and later processed in a forensic entomology laboratory. Results showed that there was no significant difference between the rate of decomposition and sequence of faunal succession on both pig carcasses. Both carcasses were completely decomposed to remain stage after nine days. The species of flies visiting the pig carcasses consisted of blow flies (Chrysomya megacephala, Chrysomya rufifacies, Hemipyrellia ligurriens), flesh fly (Sarcophagidae.), muscid fly (Ophyra spinigera), soldier fly (Hermetia illucens), coffin fly (Phoridae) and scavenger fly (Sepsidae). The only difference noted was in the number of adult flies, whereby more flies were seen in the control carcass. Faunal succession on both pig carcasses was in the following sequence: Calliphoridae, Sarcophagidae, Muscidae, Phoridae and lastly Stratiomyidae. However, there was overlap in the appearance of members of these families. Blowflies continued to oviposit on both carcasses. Hence postmortem interval (PMI) can still be estimated from the partially burnt pig carcass.

  9. Choosing appropriate space allowances for slaughter pigs transported by road: a review.

    PubMed

    Warriss, P D

    1998-04-25

    In the United Kingdom pigs can spend up to 11 hours in transit to slaughter but on average travel for two to three hours. In the past, international journeys have lasted up to 40 hours and have been over 900 miles long. There is evidence that pigs, like calves and sheep, but unlike adult cattle, prefer to lie down if provided with suitable conditions, particularly bedding, on the vehicle. They will, however, sometimes stand during short journeys, possibly when excessive vibration or uncomfortable flooring, particularly a lack of sufficient bedding, cause discomfort. Current UK legislation and EU Directive 95/29/EC specify that, in general, pigs must have sufficient space to lie down during transit. Measurements of the space needed for sternal recumbency, and direct observations of pigs at different stocking densities, suggest that the minimum space required is equivalent to about 250 kg/m2 for normal slaughter pigs of 90 to 100 kg liveweight. This figure may not be appropriate for very small or very large pigs. In the UK at present, more than half of all slaughter pigs are transported at densities greater than that prescribed (235 kg/m2) in the EU Directive. At stocking densities above about 250 kg/m2 there may not be enough room available for all the pigs to lie down, leading to continual disturbance of recumbent animals by those seeking a place to rest. A stocking density of 322 kg/m2 leads to clear evidence of physical stress. During long journeys (> or = 25 hours) meat quality is reduced by high stocking densities, implying muscle glycogen depletion and possibly fatigue. Higher stocking densities are also associated with higher mortality. There is evidence of wide variations in air temperature inside transporters, particularly for international journeys. Although there are small variations within vehicles, the temperature of the air inside is closely related to the outside temperature. It has been recommended that the temperature within the vehicle should not

  10. Long-term high intake of whole proteins results in renal damage in pigs.

    PubMed

    Jia, Yong; Hwang, Sun Young; House, James D; Ogborn, Malcolm R; Weiler, Hope A; O, Karmin; Aukema, Harold M

    2010-09-01

    Despite evidence of potential antiobesity effects of high-protein (HP) diets, the impact of consuming diets with protein levels at the upper limit of the acceptable macronutrient distribution range (AMDR) on kidney health is unknown. To test whether HP diets affect renal health, whole plant and animal proteins in proportions that mimicked human diets were given to pigs, because their kidneys have a similar anatomy and function to those of humans. Adult female pigs received either normal-protein (NP) or HP (15 or 35% of energy from protein, respectively) isocaloric diets for either 4 or 8 mo. The higher protein in the HP diet was achieved by increasing egg and dairy proteins. Although there were initial differences in body weight and composition, after 8 mo these were similar in pigs consuming the NP and HP diets. The HP compared with NP diet, however, resulted in enlarged kidneys at both 4 and 8 mo. Renal and glomerular volumes were 60-70% higher by the end of the study. These enlarged kidneys had greater evidence of histological damage, with 55% more fibrosis and 30% more glomerulosclerosis. Renal monocyte chemoattractant protein-1 levels also were 22% higher in pigs given the HP diet. Plasma homocysteine levels were higher in the HP pigs at 4 mo and continued to be elevated by 35% at 8 mo of feeding. These findings suggest that long-term intakes of protein at the upper limit of the AMDR from whole protein sources may compromise renal health.

  11. Experimental inoculation of foals and pigs with an enterotoxigenic E. coli isolated from a foal.

    PubMed

    Holland, R E; Grimes, S D; Walker, R D; Wilson, R A

    1996-10-01

    Hemolytic E. coli strain 807-13, O149:NM:K88(STb+, LT+), was isolated from the feces of a neonatal diarrheic foal. E. coli 807-13 was examined for adhesion to brush border membranes (BBM) from foals, adult horses and pigs, and its pathogenicity was assessed in neonatal foals and pigs. E. coli 807-13 did not adhere to equine BBM but adhered to pig BBM. It did not cause diarrhea nor did it colonize the intestinal epithelium of 3 colostrum-deprived and 3 suckled foals challenged at 24 h of age. Acute ulcerative gastritis and acute suppurative gastritis were observed in 2 colostrum-deprived challenged foals, and acute neutrophilic enteritis was observed in 1 colostrum-deprived and in 1 suckled challenged foal. No similar histopathologic lesions were detected in the control foals. Both gnotobiotic and suckled pigs developed diarrhea after challenge exposure to E. coli 807-13 and the intestinal epithelium of the pigs was colonized. Histopathologic evidence of gastritis and enteritis among the foals indicated some complicity of E. coli 807-13 in foal enteric disease.

  12. Tryptophan requirement of pigs in the weight category 10 to 20 kilograms.

    PubMed

    Han, Y; Chung, T K; Baker, D H

    1993-01-01

    Experiments were conducted to determine the tryptophan (Trp) requirement of 10- to 20-kg pigs. A Trp-deficient experimental diet (.11% total Trp, 18% CP, 3,320 kcal of ME/kg) was composed of corn, feather meal, corn gluten meal, soybean meal, and dried whey. True digestibility of Trp in the experimental diet was 70% (.077% digestible Trp) as established in a digestibility assay that involved cecectomized, adult cockerels (Exp. 1). An initial pig study (Exp. 2) verified that the experimental basal diet, when fortified with sufficient L-Trp, was capable of producing growth rate and feed efficiency similar to that of pigs fed a conventional corn-soybean meal-dried whey diet (18% CP, 3,320 kcal of ME/kg). In Exp. 3, crossbred pigs with an average BW of 10.9 kg were fed the Trp-deficient basal diet supplemented with 0, .015, .030, .045, .060, and .075% L-Trp. In Exp. 4, crossbred pigs that averaged 9.5 kg were fed the basal diet fortified with .030, .045, .060, and .075% L-Trp. By examining the data from Exp. 3 and 4 together, the digestible Trp requirement for maximum daily weight gain was estimated to be .14% of the diet. Assuming an 88% true digestibility of Trp in commercial diets based on corn and soybean meal (calculated from published data), the total Trp required in practice would be .16% (.89% of the dietary protein).

  13. Clone Poems and the Microcomputer.

    ERIC Educational Resources Information Center

    Irizarry, Estelle

    1989-01-01

    Describes how students can use the computer to study and create clone poems (altering original Spanish-language poems by substituting words and expressions), and how students can gain a deeper appreciation of the original poem's poetic structure and semantics. (CB)

  14. Healthy ageing of cloned sheep

    PubMed Central

    Sinclair, K. D.; Corr, S. A.; Gutierrez, C. G.; Fisher, P. A.; Lee, J.-H.; Rathbone, A. J.; Choi, I.; Campbell, K. H. S.; Gardner, D. S.

    2016-01-01

    The health of cloned animals generated by somatic-cell nuclear transfer (SCNT) has been of concern since its inception; however, there are no detailed assessments of late-onset, non-communicable diseases. Here we report that SCNT has no obvious detrimental long-term health effects in a cohort of 13 cloned sheep. We perform musculoskeletal assessments, metabolic tests and blood pressure measurements in 13 aged (7–9 years old) cloned sheep, including four derived from the cell line that gave rise to Dolly. We also perform radiological examinations of all main joints, including the knees, the joint most affected by osteoarthritis in Dolly, and compare all health parameters to groups of 5-and 6-year-old sheep, and published reference ranges. Despite their advanced age, these clones are euglycaemic, insulin sensitive and normotensive. Importantly, we observe no clinical signs of degenerative joint disease apart from mild, or in one case moderate, osteoarthritis in some animals. Our study is the first to assess the long-term health outcomes of SCNT in large animals. PMID:27459299

  15. Healthy ageing of cloned sheep.

    PubMed

    Sinclair, K D; Corr, S A; Gutierrez, C G; Fisher, P A; Lee, J-H; Rathbone, A J; Choi, I; Campbell, K H S; Gardner, D S

    2016-01-01

    The health of cloned animals generated by somatic-cell nuclear transfer (SCNT) has been of concern since its inception; however, there are no detailed assessments of late-onset, non-communicable diseases. Here we report that SCNT has no obvious detrimental long-term health effects in a cohort of 13 cloned sheep. We perform musculoskeletal assessments, metabolic tests and blood pressure measurements in 13 aged (7-9 years old) cloned sheep, including four derived from the cell line that gave rise to Dolly. We also perform radiological examinations of all main joints, including the knees, the joint most affected by osteoarthritis in Dolly, and compare all health parameters to groups of 5-and 6-year-old sheep, and published reference ranges. Despite their advanced age, these clones are euglycaemic, insulin sensitive and normotensive. Importantly, we observe no clinical signs of degenerative joint disease apart from mild, or in one case moderate, osteoarthritis in some animals. Our study is the first to assess the long-term health outcomes of SCNT in large animals.

  16. Healthy ageing of cloned sheep.

    PubMed

    Sinclair, K D; Corr, S A; Gutierrez, C G; Fisher, P A; Lee, J-H; Rathbone, A J; Choi, I; Campbell, K H S; Gardner, D S

    2016-01-01

    The health of cloned animals generated by somatic-cell nuclear transfer (SCNT) has been of concern since its inception; however, there are no detailed assessments of late-onset, non-communicable diseases. Here we report that SCNT has no obvious detrimental long-term health effects in a cohort of 13 cloned sheep. We perform musculoskeletal assessments, metabolic tests and blood pressure measurements in 13 aged (7-9 years old) cloned sheep, including four derived from the cell line that gave rise to Dolly. We also perform radiological examinations of all main joints, including the knees, the joint most affected by osteoarthritis in Dolly, and compare all health parameters to groups of 5-and 6-year-old sheep, and published reference ranges. Despite their advanced age, these clones are euglycaemic, insulin sensitive and normotensive. Importantly, we observe no clinical signs of degenerative joint disease apart from mild, or in one case moderate, osteoarthritis in some animals. Our study is the first to assess the long-term health outcomes of SCNT in large animals. PMID:27459299

  17. Guinea pig-adapted foot-and-mouth disease virus with altered receptor recognition can productively infect a natural host.

    PubMed

    Núñez, José I; Molina, Nicolas; Baranowski, Eric; Domingo, Esteban; Clark, Stuart; Burman, Alison; Berryman, Stephen; Jackson, Terry; Sobrino, Francisco

    2007-08-01

    We report that adaptation to infect the guinea pig did not modify the capacity of foot-and-mouth disease virus (FMDV) to kill suckling mice and to cause an acute and transmissible disease in the pig, an important natural host for this pathogen. Adaptive amino acid replacements (I(248)-->T in 2C, Q(44)-->R in 3A, and L(147)-->P in VP1), selected upon serial passages of a type C FMDV isolated from swine (biological clone C-S8c1) in the guinea pig, were maintained after virus multiplication in swine and suckling mice. However, the adaptive replacement L(147)-->P, next to the integrin-binding RGD motif at the GH loop in VP1, abolished growth of the virus in different established cell lines and modified its antigenicity. In contrast, primary bovine thyroid cell cultures could be productively infected by viruses with replacement L(147)-->P, and this infection was inhibited by antibodies to alphavbeta6 and by an FMDV-derived RGD-containing peptide, suggesting that integrin alphavbeta6 may be used as a receptor for these mutants in the animal (porcine, guinea pig, and suckling mice) host. Substitution T(248)-->N in 2C was not detectable in C-S8c1 but was present in a low proportion of the guinea pig-adapted virus. This substitution became rapidly dominant in the viral population after the reintroduction of the guinea pig-adapted virus into pigs. These observations illustrate how the appearance of minority variant viruses in an unnatural host can result in the dominance of these viruses on reinfection of the original host species. PMID:17522230

  18. Human reproductive cloning: a conflict of liberties.

    PubMed

    Havstad, Joyce C

    2010-02-01

    Proponents of human reproductive cloning do not dispute that cloning may lead to violations of clones' right to self-determination, or that these violations could cause psychological harms. But they proceed with their endorsement of human reproductive cloning by dismissing these psychological harms, mainly in two ways. The first tactic is to point out that to commit the genetic fallacy is indeed a mistake; the second is to invoke Parfit's non-identity problem. The argument of this paper is that neither approach succeeds in removing our moral responsibility to consider and to prevent psychological harms to cloned individuals. In fact, the same commitment to personal liberty that generates the right to reproduce by means of cloning also creates the need to limit that right appropriately. Discussion of human reproductive cloning ought to involve a careful and balanced consideration of both the relevant aspects of personal liberty - the parents' right to reproductive freedom and the cloned child's right to self-determination.

  19. Energy values of nine Populus clones

    SciTech Connect

    Strong, T.F.

    1980-01-01

    This paper compares calorific values for components of nine Populus clones. The components include stem wood, stem bark, and branches. Also compared are calorific values for clones of balsam poplar and black cottonwood parentages.

  20. Race quickens for the first human clone.

    PubMed

    Gross, M

    2001-04-01

    The dazzling creation of Dolly, the cloned sheep, led many states to legislate against the possibility of using similar technology to create human clones. But for some, this prize is proving too tempting to ignore. Michael Gross reports. PMID:11413008

  1. Method and a horizontal pipeline pig launching mechanism for sequentially launching pipeline pigs

    SciTech Connect

    Davis, G.W.

    1992-08-18

    This patent describes a method for timed automatic sequential launching of serially oriented pipeline pigs from a pig launching system having a tubular pig storage and launching magazine into a gas transmission pipeline. It comprises providing a source of hydraulic fluid medium; locating a free piston within the tubular pig storage and launching magazine for motive contact with the last of the serially oriented pipeline pigs; employing gas pressure from the gas transmission pipeline for pressurizing the hydraulic fluid medium from the source; introducing the pressurized hydraulic fluid medium into the hydraulic chamber; controllably releasing the restraining of the first of the serially oriented pipeline pigs from the tubular pig storage and launching magazine.

  2. Sp8 expression in putative neural progenitor cells in guinea pig and human cerebrum.

    PubMed

    Zhang, Xue-Mei; Cai, Yan; Wang, Fang; Wu, Jun; Mo, Lin; Zhang, Feng; Patrylo, Peter R; Pan, Aihua; Ma, Chao; Fu, Jin; Yan, Xiao-Xin

    2016-09-01

    Neural stem/progenitor cells have been characterized at neurogenic sites in adult mammalian brain with various molecular markers. Here it has been demonstrated that Sp8, a transcription factor typically expressed among mature GABAergic interneurons, also labels putative neural precursors in adult guinea pig and human cerebrum. In guinea pigs, Sp8 immunoreactive (Sp8+) cells were localized largely in the superficial layers of the cortex including layer I, as well as the subventricular zone (SVZ) and subgranular zone (SGZ). Sp8+ cells at the SGZ showed little colocalization with mature and immature neuronal markers, but co-expressed neural stem cell markers including Sox2. Some layer I Sp8+ cells also co-expressed Sox2. The amount of Sp8+ cells in the dentate gyrus was maintained 2 weeks after X-ray irradiation, while that of doublecortin (DCX+) cells was greatly reduced. Mild ischemic insult caused a transient increase of Sp8+ cells in the SGZ and layer I, with the subgranular Sp8+ cells exhibited an increased colabeling for the mitotic marker Ki67 and pulse-chased bromodeoxyuridine (BrdU). Sp8+ cells in the dentate gyrus showed an age-related decline in guinea pigs, in parallel with the loss of DCX+ cells in the same region. In adult humans, Sp8+ cells exhibited comparable morphological features as seen in guinea pigs, with those at the SGZ and some in cortical layer I co-expressed Sox2. Together, these results suggested that Sp8 may label putative neural progenitors in guinea pig and human cerebrum, with the labeled cells in the SGZ appeared largely not mitotically active under normal conditions. © 2015 Wiley Periodicals, Inc. Develop Neurobiol 76: 939-955, 2016.

  3. Probabilistic cloning of three symmetric states

    SciTech Connect

    Jimenez, O.; Bergou, J.; Delgado, A.

    2010-12-15

    We study the probabilistic cloning of three symmetric states. These states are defined by a single complex quantity, the inner product among them. We show that three different probabilistic cloning machines are necessary to optimally clone all possible families of three symmetric states. We also show that the optimal cloning probability of generating M copies out of one original can be cast as the quotient between the success probability of unambiguously discriminating one and M copies of symmetric states.

  4. Phase-covariant quantum cloning of qudits

    SciTech Connect

    Fan Heng; Imai, Hiroshi; Matsumoto, Keiji; Wang, Xiang-Bin

    2003-02-01

    We study the phase-covariant quantum cloning machine for qudits, i.e., the input states in a d-level quantum system have complex coefficients with arbitrary phase but constant module. A cloning unitary transformation is proposed. After optimizing the fidelity between input state and single qudit reduced density operator of output state, we obtain the optimal fidelity for 1 to 2 phase-covariant quantum cloning of qudits and the corresponding cloning transformation.

  5. [Cloning: reproductive medicine or breeding program?].

    PubMed

    Zülicke, F

    1998-01-01

    The presentation of clone-sheep Dolly in February 1997 which was the result of a long and costly research process by Ian Wilmut's team at Roslin Institute near Edinburgh brought world-wide headlines and a continuous debate. But neither cloning and cloning experiments nor the debates about it and the possible application on humans are as new as it is shown in the media. The following article gives some facts and arguments to the field of cloning.

  6. Ongoing research on mammalian cloning and embryo stem cell technologies: bioethics of their potential medical applications.

    PubMed

    Revel, M

    2000-07-01

    Reproduction by cloning has been achieved by transfer into enucleated oocytes of nuclei from embryonic cells and more recently from cells of adult animals. The efficiency at which embryos produced by such nuclear transfers will develop into healthy newborns is very low but has succeeded in producing some cloned bovines, ovines and mice. Since the first report of sheep cloning from an adult cell in 1997, the potential applications of reproductive cloning in human medicine have been envisaged amidst a flurry of moral debates. Although the technology is still far from being ready for any human use, it has been condemned up front. It has also led to irrational fantasies and fears, based mainly on the misconception that genetic identity means identical twin personalities. Scientific research is ongoing to refine the cloning technology for applications in the production of genetically homogeneous farm animals with useful nutritional or therapeutic genetic traits. A new area of research is non-reproductive therapeutic cloning for the purpose of producing autologous embryonic cells and tissues for transplantation.

  7. Cloning and Variation of Ground State Intestinal Stem Cells

    PubMed Central

    Wang, Xia; Yamamoto, Yusuke; Wilson, Lane H.; Zhang, Ting; Howitt, Brooke; Farrow, Melissa A.; Kern, Florian; Ning, Gang; Hong, Yue; Khor, Chiea Chuen; Chevalier, Benoit; Bertrand, Denis; Wu, Lingyan; Nagarajan, Niranjan; Sylvester, Francisco A.; Hyams, Jeffrey S.; Devers, Thomas; Bronson, Roderick; Lacy, D. Borden; Ho, Khek Yu; Crum, Christopher P.; McKeon, Frank; Xian, Wa

    2016-01-01

    Summary Stem cells of the gastrointestinal tract, pancreas, liver, and other columnar epithelia collectively resist cloning in their elemental states. Here we demonstrate the cloning and propagation of highly clonogenic, “ground state” stem cells of the human intestine and colon. We show that derived stem cell pedigrees sustain limited copy number and sequence variation despite extensive serial passaging and display exquisitely precise, cell-autonomous commitment to epithelial differentiation consistent with their origins along the intestinal tract. This developmentally patterned and epigenetically maintained commitment of stem cells likely enforces the functional specificity of the adult intestinal tract. Using clonally-derived colonic epithelia, we show that toxins A or B of the enteric pathogen C. difficile recapitulate the salient features of pseudomembranous colitis. The stability of the epigenetic commitment programs of these stem cells, coupled with their unlimited replicative expansion and maintained clonogenicity, suggests certain advantages for their use in disease modeling and regenerative medicine. PMID:26040716

  8. Economical phase-covariant cloning of qudits

    SciTech Connect

    Buscemi, Francesco; D'Ariano, Giacomo Mauro; Macchiavello, Chiara

    2005-04-01

    We derive the optimal N{yields}M phase-covariant quantum cloning for equatorial states in dimension d with M=kd+N, k integer. The cloning maps are optimal for both global and single-qudit fidelity. The map is achieved by an 'economical' cloning machine, which works without ancilla.

  9. Local cloning of arbitrarily entangled multipartite states

    SciTech Connect

    Kay, Alastair; Ericsson, Marie

    2006-01-15

    We examine the perfect cloning of nonlocal, orthogonal states using only local operations and classical communication. We provide a complete characterisation of the states that can be cloned under these restrictions, and their relation to distinguishability. We also consider the case of catalytic cloning, which we show provides no enhancement to the set of clonable states.

  10. Blastocystis tropism in the pig intestine

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Blastocystis subtype 5, a subtype known to infect humans, was detected by molecular methods in the feces of 36 naturally infected market age pigs. At necropsy, 6 heavily infected pigs were selected to determine the tropism of the infection within the gastrointestinal tract. Because so little is know...

  11. Sweating Like a Pig: Physics or Irony?

    ERIC Educational Resources Information Center

    Bohren, Craig F.

    2016-01-01

    In his interesting and informative book "Is That a Fact?," Joe Schwarcz avers that pigs do not sweat and the saying "sweating like a pig" originates in iron smelting. Oblong pieces of hot iron, with a fancied resemblance to a sow with piglets, cool in sand to the dew point of the surrounding air, and hence water condenses on…

  12. Archaea in the intestinal tract of pigs

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Knowledge of Archaea in the intestinal tract of pigs is limited. In order to investigate archaeal community structure, samples were taken from the cecum and proximal colon of finishing pigs (24) fed diets with either corn or solvent extracted corn germ meal (CGM). Corn germ meal feeding began in w...

  13. Genetically modified pig models for neurodegenerative disorders.

    PubMed

    Holm, Ida E; Alstrup, Aage Kristian Olsen; Luo, Yonglun

    2016-01-01

    Increasing incidence of neurodegenerative disorders such as Alzheimer's disease and Parkinson's disease has become one of the most challenging health issues in ageing humans. One approach to combat this is to generate genetically modified animal models of neurodegenerative disorders for studying pathogenesis, prognosis, diagnosis, treatment, and prevention. Owing to the genetic, anatomic, physiologic, pathologic, and neurologic similarities between pigs and humans, genetically modified pig models of neurodegenerative disorders have been attractive large animal models to bridge the gap of preclinical investigations between rodents and humans. In this review, we provide a neuroanatomical overview in pigs and summarize and discuss the generation of genetically modified pig models of neurodegenerative disorders including Alzheimer's diseases, Huntington's disease, Parkinson's disease, amyotrophic lateral sclerosis, spinal muscular atrophy, and ataxia-telangiectasia. We also highlight how non-invasive bioimaging technologies such as positron emission tomography (PET), computer tomography (CT), and magnetic resonance imaging (MRI), and behavioural testing have been applied to characterize neurodegenerative pig models. We further propose a multiplex genome editing and preterm recloning (MAP) approach by using the rapid growth of the ground-breaking precision genome editing technology CRISPR/Cas9 and somatic cell nuclear transfer (SCNT). With this approach, we hope to shorten the temporal requirement in generating multiple transgenic pigs, increase the survival rate of founder pigs, and generate genetically modified pigs that will more closely resemble the disease-causing mutations and recapitulate pathological features of human conditions. PMID:26446984

  14. Guinea Pigs: Versatile Animals for the Classroom

    ERIC Educational Resources Information Center

    Barman, Charles R.

    1977-01-01

    Guinea pigs are presented as versatile classroom animals. Suggestions for animal behavior and genetics studies are given. Also included is information concerning sex determination and the breeding of guinea pigs, and hints on keeping these animals in the classroom. References and illustrations complete the article. (MA)

  15. Size-sex variation in survival rates and abundance of pig frogs, Rana grylio, in northern Florida wetlands

    USGS Publications Warehouse

    Wood, K.V.; Nichols, J.D.; Percival, H.F.; Hines, J.E.

    1998-01-01

    During 1991-1993, we conducted capture-recapture studies on pig frogs, Rana grylio, in seven study locations in northcentral Florida. Resulting data were used to test hypotheses about variation in survival probability over different size-sex classes of pig frogs. We developed multistate capture-recapture models for the resulting data and used them to estimate survival rates and frog abundance. Tests provided strong evidence of survival differences among size-sex classes, with adult females showing the highest survival probabilities. Adult males and juvenile frogs had lower survival rates that were similar to each other. Adult females were more abundant than adult males in most locations at most sampling occasions. We recommended probabilistic capture-recapture models in general, and multistate models in particular, for robust estimation of demographic parameters in amphibian populations.

  16. Characterization of Salmonella enterica Serovar Typhimurium Isolates from Pigs and Pig Environment-Related Sources and Evidence of New Circulating Monophasic Strains in Spain.

    PubMed

    Andrés-Barranco, Sara; Vico, Juan Pablo; Marín, Clara María; Herrera-León, Silvia; Mainar-Jaime, Raú Carlos

    2016-03-01

    A total of 117 Salmonella enterica serovar Typhimurium and 59 monophasic Salmonella Typhimurium (S. enterica serovar 4,[5],12:i:-) strains isolated between 2008 and 2012 from pig, wild bird, rodent, and farm environment samples from the northeast of Spain were characterized by phage typing, antibiotic susceptibility testing, and multiple-locus variable-number tandem repeat analysis in order to evaluate their phenotypic and genetic relatedness. In Salmonella 4,[5],12:i:-, the most prevalent phage types were U311 (40.7%) and DT195 (22%), which did not correspond with the so-called Spanish clone and generally showed a different resistance pattern (ASSuT). Antibiotic resistance was found in 85.8% of the isolates, with 94.1% of them displaying multidrug resistance. Multiple-locus variable-number tandem repeat analysis identified 92 different profiles, six of them shared by both serovars. The minimum spanning tree showed one major cluster that included 95% of the Salmonella 4,[5],12:i:- isolates, which came from different animal sources, geographic locations, and time periods, suggesting high clonality among those Salmonella strains and the ability to spread among pig farms. Overall, isolates of Salmonella 4,[5],12:i:- were more similar to European strains than to the well-characterized Spanish clone. The spread of these new strains of Salmonella 4,[5],12:i:- would likely have been favored by the important pig trade between this Spanish region and other European countries. The overall high prevalence of multidrug resistance observed in these new strains should be noted.

  17. Isolation and characterization of Guinea pig properidin.

    PubMed

    Nicholson, A; Austen, K F

    1977-01-01

    Guinea pig properdin was purified to homogeneity by employing as an assay during isolation its capacity to augment the hemolytic activity of a heterologous human C3b-dependent C3 convertase, C3B. The purified protein elicited a monospecific antibody response in a rabbit. The antiserum, by immunodiffusion, gave a reaction of identity between a protein in whole guinea pig serum and the immunogen. A solid phase immunoadsorbent prepared with the antiserum removed properdin function from the purified protein. The purified guinea pig protein exhibited the classical properdin function of reconstituting a human RP for zymosan-induced C3 inactivation. The guinea pig properdin also agglutinated red cell intermediates bearing either guinea pig or human C3b and retarded the decay of homologous C3 convertase, C3B. These activities are the same as those observed for purified human properdin and validate the amplification function of properdin on terminal component activation in a second species.

  18. Endocrine tumours in the guinea pig.

    PubMed

    Künzel, Frank; Mayer, Jörg

    2015-12-01

    Functional endocrine tumours have long been thought to be rare in guinea pigs, although conditions such as hyperthyroidism and hyperadrenocorticism have been documented with increasing frequency so the prevalence of hormonal disorders may have been underestimated. Both the clinical signs and diagnosis of hyperthyroidism in guinea pigs appear to be very similar to those described in feline hyperthyroidism, and methimazole has been proven to be a practical therapy option. Hyperadrenocorticism has been confirmed in several guinea pigs with an adrenocorticotropic hormone stimulation test using saliva as a non-invasive sample matrix; trilostane has been successfully used to treat a guinea pig with hyperadrenocorticism. Insulinomas have only rarely been documented in guinea pigs and one animal was effectively treated with diazoxide. PMID:26542368

  19. Endocrine tumours in the guinea pig.

    PubMed

    Künzel, Frank; Mayer, Jörg

    2015-12-01

    Functional endocrine tumours have long been thought to be rare in guinea pigs, although conditions such as hyperthyroidism and hyperadrenocorticism have been documented with increasing frequency so the prevalence of hormonal disorders may have been underestimated. Both the clinical signs and diagnosis of hyperthyroidism in guinea pigs appear to be very similar to those described in feline hyperthyroidism, and methimazole has been proven to be a practical therapy option. Hyperadrenocorticism has been confirmed in several guinea pigs with an adrenocorticotropic hormone stimulation test using saliva as a non-invasive sample matrix; trilostane has been successfully used to treat a guinea pig with hyperadrenocorticism. Insulinomas have only rarely been documented in guinea pigs and one animal was effectively treated with diazoxide.

  20. Cloning humans, cloning literature: genetics and the imagination deficit.

    PubMed

    Van Dijck, J

    1999-01-01

    After the birth of Dolly, media stories on cloning were replete with references to well-known science fiction plots. This essay criticizes the 'imagination deficit' of scientists and journalists, first by problematizing the uncritical adoption of attentuated science fiction plots in the media coverage of Dolly, and second, by proposing to look at more expansive science fiction novels that carefully examine issues such as uniqueness and identity in relation to the new genetics.

  1. Guinea Pig Ciliary Muscle Development

    PubMed Central

    Pucker, Andrew D.; Carpenter, Ashley R.; McHugh, Kirk M.; Mutti, Donald O.

    2014-01-01

    Purpose The purpose of this study was to develop a method for quantifying guinea pig ciliary muscle volume (CMV) and to determine its relationship to age and ocular biometric measurements. Methods Six albino guinea pigs eyes were collected at each of five ages (n=30 eyes). Retinoscopy and photography were used to document refractive error, eye size, and eye shape. Serial sections through the excised eyes were made and then labeled with an α-smooth muscle actin antibody. The CM was then visualized with an Olympus BX51 microscope, reconstructed with Stereo Investigator (MBF Bioscience) and analyzed using Neurolucida Explorer (MBF Bioscience). Full (using all sections) and partial (using a subset of sections) reconstruction methods were used to determine CMV. Results There was no significant difference between the full and partial volume determination methods (P = 0.86). The mean CMV of the 1, 10, 20, 30, and 90-day old eyes was 0.40 ± 0.16 mm3, 0.48 ± 0.13 mm3, 0.67 ± 0.15 mm3, 0.86 ± 0.35 mm3, and 1.09 ± 0.63 mm3, respectively. CMV was significantly correlated with log age (P = 0.001), ocular length (P = 0.003), limbal circumference (P = 0.01), and equatorial diameter (P = 0.003). It was not correlated with refractive error (P = 0.73) or eye shape (P = 0.60). Multivariate regression determined that biometric variables were not significantly associated with CMV after adjustment for age. Conclusions Three-dimensional reconstruction was an effective means of determining CMV. These data provide evidence that CM growth occurs with age in tandem with eye size in normal albino guinea pigs. Additional work is needed to determine the relationship between CMV and abnormal ocular growth. PMID:24901488

  2. Stress susceptibility in pigs supplemented with ractopamine.

    PubMed

    Athayde, N B; Dalla Costa, O A; Roça, R O; Guidoni, A L; Ludtke, C B; Oba, E; Takahira, R K; Lima, G J M M

    2013-09-01

    Ractopamine is a β-adrenergic agonist used as an energy repartitioning agent in the diets of finishing pigs. Most ractopamine studies are limited to evaluations of growth performance and meat quality, and there is little information on the effects of this additive on the behavior and welfare of pigs. Therefore, the objective of this study was to evaluate various indicators of stress caused by feeding diets containing ractopamine. One hundred seventy barrows and 170 gilts weighing 107.3 kg were allocated to 30 pens with 10 to 12 barrows or gilts per pen. Pigs were offered 1 of the 3 dietary treatments (0, 5, or 10 mg ractopamine/kg) for 28 d with 5 barrow pens and 5 gilt pens per treatment. Pigs were evaluated for behavior 3 d per week 1 wk before the initiation of the experiment and throughout the experiment. Each pig was classified into 1 of the 13 activities (drinking water, lying alone, lying in clusters, standing, nosing pig, sitting, feeding, biting pig, walking, exploring, running away, playing, and mounting pen mates) and also grouped into 1 of the 3 categories (calm, moving, and feeding themselves) based on those activities. At the end of the experiment, 3 pigs from each pen were slaughtered, and blood samples were collected during exsanguination to determine physiological indicators of stress (cortisol, lactate, and creatine-kinase enzymes). The incidence of skin and carcass lesions was determined at shoulder, loin, and ham. Ractopamine had no effect (P > 0.05) on pig behavior, total number of skin and carcass lesions, or blood concentrations of cortisol or lactate. However, there was an increase (P < 0.05) of creatine kinase concentrations in pigs receiving ractopamine-supplemented feed. This finding is consistent with the concept that ractopamine may cause muscular disorders, and this warrants further investigation.

  3. Predators induce cloning in echinoderm larvae.

    PubMed

    Vaughn, Dawn; Strathmann, Richard R

    2008-03-14

    Asexual propagation (cloning) is a widespread reproductive strategy of plants and animals. Although larval cloning is well documented in echinoderms, identified stimuli for cloning are limited to those associated with conditions favorable for growth and reproduction. Our research shows that larvae of the sand dollar Dendraster excentricus also clone in response to cues from predators. Predator-induced clones were smaller than uncloned larvae, suggesting an advantage against visual predators. Our results offer another ecological context for asexual reproduction: rapid size reduction as a defense.

  4. Probabilistic cloning of three nonorthogonal states

    NASA Astrophysics Data System (ADS)

    Zhang, Wen; Rui, Pinshu; Yang, Qun; Zhao, Yan; Zhang, Ziyun

    2015-04-01

    We study the probabilistic cloning of three nonorthogonal states with equal success probabilities. For simplicity, we assume that the three states belong to a special set. Analytical form of the maximal success probability for probabilistic cloning is calculated. With the maximal success probability, we deduce the explicit form of probabilistic quantum cloning machine. In the case of cloning, we get the unambiguous form of the unitary operation. It is demonstrated that the upper bound for probabilistic quantum cloning machine in (Qiu in J Phys A 35:6931, 2002) can be reached only if the three states are equidistant.

  5. Optimal quantum cloning via spin networks

    SciTech Connect

    Chen Qing; Cheng Jianhua; Wang Kelin; Du Jiangfeng

    2006-09-15

    In this paper we demonstrate that optimal 1{yields}M phase-covariant cloning quantum cloning is available via free dynamical evolution of spin networks. By properly designing the network and the couplings between spins, we show that optimal 1{yields}M phase-covariant cloning can be achieved if the initial state is prepared as a specific symmetric state. Especially, when M is an odd number, the optimal phase-covariant cloning can be achieved without ancillas. Moreover, we demonstrate that the same framework is capable for optimal 1{yields}2 universal cloning.

  6. No-cloning theorem on quantum logics

    SciTech Connect

    Miyadera, Takayuki; Imai, Hideki

    2009-10-15

    This paper discusses the no-cloning theorem in a logicoalgebraic approach. In this approach, an orthoalgebra is considered as a general structure for propositions in a physical theory. We proved that an orthoalgebra admits cloning operation if and only if it is a Boolean algebra. That is, only classical theory admits the cloning of states. If unsharp propositions are to be included in the theory, then a notion of effect algebra is considered. We proved that an atomic Archimedean effect algebra admitting cloning operation is a Boolean algebra. This paper also presents a partial result, indicating a relation between the cloning on effect algebras and hidden variables.

  7. Therapeutic and reproductive cloning: a critique.

    PubMed

    Bowring, Finn

    2004-01-01

    This article is a critical examination of the science and ethics of human cloning. It summarises the key scientific milestones in the development of nuclear transplantation, explains the importance of cloning to research into the medical potential of embryonic stem cells, and discusses the well-worn distinction between 'therapeutic' and 'reproductive' cloning. Suggesting that this distinction will be impossible to police, it goes on to consider the ethics of full human cloning. It is concluded that it represents an unacceptable form of parental despotism, and that the genetic engineering and cloning of future human beings will fracture the foundations of modern humanism.

  8. Brain Mass and Encephalization Quotients in the Domestic Industrial Pig (Sus scrofa).

    PubMed

    Minervini, Serena; Accogli, Gianluca; Pirone, Andrea; Graïc, Jean-Marie; Cozzi, Bruno; Desantis, Salvatore

    2016-01-01

    In the present study we examined the brain of fetal, newborn, and adult pigs raised for meat production. The fresh and formalin-fixed weights of the brain have been recorded and used, together with body weight, to calculate the Encephalization Quotient (EQ). The weight of the cerebellum has been used to calculate the Cerebellar Quotient (CQ). The results have been discussed together with analogue data obtained in other terrestrial Cetartiodactyla (including the domestic bovine, sheep, goat, and camel), domesticated Carnivora, Proboscidata, and Primates. Our study, based on a relatively large experimental series, corrects former observations present in the literature based on smaller samples, and emphasizes that the domestic pig has a small brain relative to its body size (EQ = 0.38 for adults), possibly due to factors linked to the necessity of meat production and improved body weight. Comparison with other terrestrial Cetartiodactyla indicates a similar trend for all domesticated species. PMID:27351807

  9. Brain Mass and Encephalization Quotients in the Domestic Industrial Pig (Sus scrofa)

    PubMed Central

    Minervini, Serena; Accogli, Gianluca; Pirone, Andrea; Graïc, Jean-Marie; Cozzi, Bruno; Desantis, Salvatore

    2016-01-01

    In the present study we examined the brain of fetal, newborn, and adult pigs raised for meat production. The fresh and formalin-fixed weights of the brain have been recorded and used, together with body weight, to calculate the Encephalization Quotient (EQ). The weight of the cerebellum has been used to calculate the Cerebellar Quotient (CQ). The results have been discussed together with analogue data obtained in other terrestrial Cetartiodactyla (including the domestic bovine, sheep, goat, and camel), domesticated Carnivora, Proboscidata, and Primates. Our study, based on a relatively large experimental series, corrects former observations present in the literature based on smaller samples, and emphasizes that the domestic pig has a small brain relative to its body size (EQ = 0.38 for adults), possibly due to factors linked to the necessity of meat production and improved body weight. Comparison with other terrestrial Cetartiodactyla indicates a similar trend for all domesticated species. PMID:27351807

  10. 9 CFR 113.38 - Guinea pig safety test.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... 9 Animals and Animal Products 1 2011-01-01 2011-01-01 false Guinea pig safety test. 113.38 Section... Standard Procedures § 113.38 Guinea pig safety test. The guinea pig safety test provided in this section... be injected either intramuscularly or subcutaneously into each of two guinea pigs and the...

  11. Cultural and Economic Motivation of Pig Raising Practices in Bangladesh.

    PubMed

    Nahar, Nazmun; Uddin, Main; Gurley, Emily S; Jahangir Hossain, M; Sultana, Rebeca; Luby, Stephen P

    2015-12-01

    The interactions that pig raisers in Bangladesh have with their pigs could increase the risk of zoonotic disease transmission. Since raising pigs is a cultural taboo to Muslims, we aimed at understanding the motivation for raising pigs and resulting practices that could pose the risk of transmitting disease from pigs to humans in Bangladesh, a predominantly Muslim country. These understandings could help identify acceptable strategies to reduce the risk of disease transmission from pigs to people. To achieve this objective, we conducted 34 in-depth interviews among pig herders and backyard pig raisers in eight districts of Bangladesh. Informants explained that pig raising is an old tradition, embedded in cultural and religious beliefs and practices, the primary livelihood of pig herders, and a supplemental income of backyard pig raisers. To secure additional income, pig raisers sell feces, liver, bile, and other pig parts often used as traditional medicine. Pig raisers have limited economic ability to change the current practices that may put them at risk of exposure to diseases from their pigs. An intervention that improves their financial situation and reduces the risk of zoonotic disease may be of interest to pig raisers.

  12. 9 CFR 113.38 - Guinea pig safety test.

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... 9 Animals and Animal Products 1 2012-01-01 2012-01-01 false Guinea pig safety test. 113.38 Section... Standard Procedures § 113.38 Guinea pig safety test. The guinea pig safety test provided in this section... be injected either intramuscularly or subcutaneously into each of two guinea pigs and the...

  13. 9 CFR 113.38 - Guinea pig safety test.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... 9 Animals and Animal Products 1 2014-01-01 2014-01-01 false Guinea pig safety test. 113.38 Section... Standard Procedures § 113.38 Guinea pig safety test. The guinea pig safety test provided in this section... be injected either intramuscularly or subcutaneously into each of two guinea pigs and the...

  14. An Improved System for Generation of Diploid Cloned Porcine Embryos Using Induced Pluripotent Stem Cells Synchronized to Metaphase.

    PubMed

    Kim, Eunhye; Zheng, Zhong; Jeon, Yubyeol; Jin, Yong-Xun; Hwang, Seon-Ung; Cai, Lian; Lee, Chang-Kyu; Kim, Nam-Hyung; Hyun, Sang-Hwan

    2016-01-01

    Pigs provide outstanding models of human genetic diseases due to their striking similarities with human anatomy, physiology and genetics. Although transgenic pigs have been produced using genetically modified somatic cells and nuclear transfer (SCNT), the cloning efficiency was extremely low. Here, we report an improved method to produce diploid cloned embryos from porcine induced pluripotent stem cells (piPSCs), which were synchronized to the G2/M stage using a double blocking method with aphidicolin and nocodazole. The efficiency of this synchronization method on our piPSC lines was first tested. Then, we modified our traditional SCNT protocol to find a workable protocol. In particular, the removal of a 6DMAP treatment post-activation enhanced the extrusion rate of pseudo-second-polar bodies (p2PB) (81.3% vs. 15.8%, based on peak time, 4hpa). Moreover, an immediate activation method yielded significantly more blastocysts than delayed activation (31.3% vs. 16.0%, based on fused embryos). The immunofluorescent results confirmed the effect of the 6DMAP treatment removal, showing remarkable p2PB extrusion during a series of nuclear transfer procedures. The reconstructed embryos from metaphase piPSCs with our modified protocol demonstrated normal morphology at 2-cell, 4-cell and blastocyst stages and a high rate of normal karyotype. This study demonstrated a new and efficient way to produce viable cloned embryos from piPSCs when synchronized to the G2/M phase of the cell cycle, which may lead to opportunities to produce cloned pigs from piPSCs more efficiently. PMID:27472781

  15. An Improved System for Generation of Diploid Cloned Porcine Embryos Using Induced Pluripotent Stem Cells Synchronized to Metaphase

    PubMed Central

    Jeon, Yubyeol; Jin, Yong-Xun; Hwang, Seon-Ung; Cai, Lian; Lee, Chang-Kyu; Kim, Nam-Hyung; Hyun, Sang-Hwan

    2016-01-01

    Pigs provide outstanding models of human genetic diseases due to their striking similarities with human anatomy, physiology and genetics. Although transgenic pigs have been produced using genetically modified somatic cells and nuclear transfer (SCNT), the cloning efficiency was extremely low. Here, we report an improved method to produce diploid cloned embryos from porcine induced pluripotent stem cells (piPSCs), which were synchronized to the G2/M stage using a double blocking method with aphidicolin and nocodazole. The efficiency of this synchronization method on our piPSC lines was first tested. Then, we modified our traditional SCNT protocol to find a workable protocol. In particular, the removal of a 6DMAP treatment post-activation enhanced the extrusion rate of pseudo-second-polar bodies (p2PB) (81.3% vs. 15.8%, based on peak time, 4hpa). Moreover, an immediate activation method yielded significantly more blastocysts than delayed activation (31.3% vs. 16.0%, based on fused embryos). The immunofluorescent results confirmed the effect of the 6DMAP treatment removal, showing remarkable p2PB extrusion during a series of nuclear transfer procedures. The reconstructed embryos from metaphase piPSCs with our modified protocol demonstrated normal morphology at 2-cell, 4-cell and blastocyst stages and a high rate of normal karyotype. This study demonstrated a new and efficient way to produce viable cloned embryos from piPSCs when synchronized to the G2/M phase of the cell cycle, which may lead to opportunities to produce cloned pigs from piPSCs more efficiently. PMID:27472781

  16. Cloning Expeditions: Risky but Rewarding

    PubMed Central

    2013-01-01

    In the 1980s, a good part of my laboratory was using the then-new recombinant DNA techniques to clone and characterize many important cell surface membrane proteins: GLUT1 (the red cell glucose transporter) and then GLUT2 and GLUT4, the red cell anion exchange protein (Band 3), asialoglycoprotein receptor subunits, sucrase-isomaltase, the erythropoietin receptor, and two of the subunits of the transforming growth factor β (TGF-β) receptor. These cloned genes opened many new fields of basic research, including membrane insertion and trafficking of transmembrane proteins, signal transduction by many members of the cytokine and TGF-β families of receptors, and the cellular physiology of glucose and anion transport. They also led to many insights into the molecular biology of several cancers, hematopoietic disorders, and diabetes. This work was done by an exceptional group of postdocs and students who took exceptionally large risks in developing and using novel cloning technologies. Unsurprisingly, all have gone on to become leaders in the fields of molecular cell biology and molecular medicine. PMID:24061478

  17. Cloning expeditions: risky but rewarding.

    PubMed

    Lodish, Harvey

    2013-12-01

    In the 1980s, a good part of my laboratory was using the then-new recombinant DNA techniques to clone and characterize many important cell surface membrane proteins: GLUT1 (the red cell glucose transporter) and then GLUT2 and GLUT4, the red cell anion exchange protein (Band 3), asialoglycoprotein receptor subunits, sucrase-isomaltase, the erythropoietin receptor, and two of the subunits of the transforming growth factor β (TGF-β) receptor. These cloned genes opened many new fields of basic research, including membrane insertion and trafficking of transmembrane proteins, signal transduction by many members of the cytokine and TGF-β families of receptors, and the cellular physiology of glucose and anion transport. They also led to many insights into the molecular biology of several cancers, hematopoietic disorders, and diabetes. This work was done by an exceptional group of postdocs and students who took exceptionally large risks in developing and using novel cloning technologies. Unsurprisingly, all have gone on to become leaders in the fields of molecular cell biology and molecular medicine.

  18. Behavioral recovery induced by applied electric fields after spinal cord hemisection in guinea pig

    SciTech Connect

    Borgens, R.B.; Blight, A.R.; McGinnis, M.E.

    1987-10-16

    Applied electric fields were used to promote axonal regeneration in spinal cords of adult guinea pigs. A propriospinal intersegmental reflex (the cutaneous trunci muscle reflex) was used to test lateral tract function after hemisection of the thoracic spinal cord. An electrical field (200 microvolts per millimeter, cathode rostral) applied across the lesion led to functional recovery of the cutaneous trunci muscle reflex in 25 percent of experimental animals, whereas the functional deficit remained in control animals, which were implanted with inactive stimulators.

  19. Mutation analysis with random DNA identifiers (MARDI) catalogs Pig-a mutations in heterogeneous pools of CD48-deficient T cells derived from DMBA-treated rats.

    PubMed

    Revollo, Javier R; Crabtree, Nathaniel M; Pearce, Mason G; Pacheco-Martinez, M Monserrat; Dobrovolsky, Vasily N

    2016-03-01

    Identification of mutations induced by xenotoxins is a common task in the field of genetic toxicology. Mutations are often detected by clonally expanding potential mutant cells and genotyping each viable clone by Sanger sequencing. Such a "clone-by-clone" approach requires significant time and effort, and sometimes is even impossible to implement. Alternative techniques for efficient mutation identification would greatly benefit both basic and regulatory genetic toxicology research. Here, we report the development of Mutation Analysis with Random DNA Identifiers (MARDI), a novel high-fidelity Next Generation Sequencing (NGS) approach that circumvents clonal expansion and directly catalogs mutations in pools of mutant cells. MARDI uses oligonucleotides carrying Random DNA Identifiers (RDIs) to tag progenitor DNA molecules before PCR amplification, enabling clustering of descendant DNA molecules and eliminating NGS- and PCR-induced sequencing artifacts. When applied to the Pig-a cDNA analysis of heterogeneous pools of CD48-deficient T cells derived from DMBA-treated rats, MARDI detected nearly all Pig-a mutations that were previously identified by conventional clone-by-clone analysis and discovered many additional ones consistent with DMBA exposure: mostly A to T transversions, with the mutated A located on the non-transcribed DNA strand. PMID:26683280

  20. Sarcoptic mange infestation in pigs: an overview.

    PubMed

    Laha, R

    2015-12-01

    Sarcoptic mange infestation in pigs is caused by Sarcoptes scabiei var. suis. It is the most common mange infestation of pigs. The parasite is distributed worldwide. Pig owners are generally concerned about the internal parasitic infections and ignored the external parasitic infestations. But the external parasitic infestation with S. scabiei var. suis has economic significance as it causes morbidity, mortality, decreased fertility and feed conversion ratio in pigs. Keeping in view of importance of S. scabies var. suis infestation in pigs, this communication discussed about the present and past research works done on S. scabies var. suis infestation in pigs, particularly its prevalence, life cycle, pathological lesions, clinical symptoms, haematobiochemical changes, diagnosis, treatment and control, to have an idea about this infestation at a glance. It has been concluded that the research work done on sarcoptic mange infestation in pigs in India is less in comparison to other countries. It may be due to its consideration as a neglected parasite or due to it's under report. Organization of awareness programs for the farmers by extension personalities or other authorities might be able to save the farmers from economic losses due to this infestation. PMID:26688620

  1. Continuous odour measurement from fattening pig units

    NASA Astrophysics Data System (ADS)

    Romain, Anne-Claude; Nicolas, Jacques; Cobut, Pierre; Delva, Julien; Nicks, Baudouin; Philippe, François-Xavier

    2013-10-01

    A study in experimental slatted-system fattening pig units was conducted with the aim of estimating the odour emission factor (in ou s.pig-1), which can subsequently be used in dispersion models to assess the odour annoyance zone. Dynamic olfactometry measurements carried out at different development stages of pigs showed a logical trend of the mean assessed odour emission factor with the pig mass. However, the variation within the same mass class was much larger than variation between classes. Possible causes of such variation were identified as the evolution of ventilation rate during the day and the circadian rhythm of pig. To be able to monitor continuously the daily variation of the odour, an electronic nose was used with suitable regression model calibrated against olfactometric measurements. After appropriate validation check, the electronic nose proved to be convenient, as a complementary tool to dynamic olfactometry, to record the daily variation of the odour emission factor in the pig barn. It was demonstrated that, in the controlled conditions of the experimental pens, the daily variation of the odour emission rate could be mainly attributed to the sole influence of the circadian rhythm of pig. As a consequence, determining a representative odour emission factor in a real case cannot be based on a snapshot odour sampling.

  2. Chronic prenatal ethanol exposure alters glucocorticoid signalling in the hippocampus of the postnatal Guinea pig.

    PubMed

    Iqbal, U; Brien, J F; Banjanin, S; Andrews, M H; Matthews, S G; Reynolds, J N

    2005-09-01

    The present study tested the hypothesis that chronic prenatal ethanol exposure causes long-lasting changes in glucocorticoid signalling in postnatal offspring. Pregnant guinea pigs were treated with ethanol (4 g/kg maternal body weight/day), isocaloric-sucrose/pair-feeding or water throughout gestation, and maternal saliva cortisol concentration was determined 2 h after treatment at different stages of gestation. Electrically-stimulated release of glutamate and GABA, in the presence or absence of dexamethasone, as well as glucocorticoid and mineralocorticoid receptor mRNA expression, was determined in the hippocampus and prefrontal cortex of adult offspring of treated pregnant guinea pigs. Maternal saliva cortisol concentration increased throughout pregnancy, which was associated with increased foetal plasma and amniotic fluid cortisol concentration. Ethanol administration to pregnant guinea pigs increased maternal saliva cortisol concentration during early and mid-gestation. In late gestation, ethanol administration did not increase saliva cortisol concentration above that induced by pregnancy. Chronic prenatal ethanol exposure had no effect on stimulated glutamate or GABA release, but selectively prevented dexamethasone-mediated suppression of stimulated glutamate release, and decreased expression of mineralocorticoid, but not glucocorticoid, receptor mRNA in the hippocampus of adult offspring. These data indicate that maternal ethanol administration leads to excessively increased maternal cortisol concentration that can impact negatively the developing foetal brain, leading to persistent postnatal deficits in glucocorticoid regulation of glutamate signalling in the adult hippocampus. PMID:16101899

  3. Immunity to Escherichia coli in pigs: Serum Gamma Globulin Levels, Indirect Hemagglutinating Antibody Titres and Bactericidal Activity Against E. coli in pigs up to five Weeks of Age

    PubMed Central

    Wilson, M. R.; Svendsen, J.

    1972-01-01

    Serum gamma globulin levels, indirect hemagglutinating antibody titres and bactericidal activity against the 0149:K91;K88ac:H10 Serotype of Escherichia coli were determined in pigs up to five weeks of age from vaccinated and non-vaccinated sows. Gamma globulin levels at two days of age were approximately twice adult levels, by three weeks of age they were one quarter of adult levels and remained so until five weeks of age. Indirect hemagglutinating antibody activity was highest at two days of age, fell until three weeks of age and then rose. Little or no indirect hemagglutinating antibody activity was detected in sera taken at two days of age from pigs from non-vaccinated sows. Only three of 26 two day old pigs had demonstrable bactericidal activity; by three weeks of age 16 of 26 had bactericidal activity. Serum from piglets of vaccinated sows had no more bactericidal activity than did sera from non-vaccinated sows. PMID:4110608

  4. Pigging ends freezeups in caustic piping system

    SciTech Connect

    Gros, R.; Gaines, A.

    1985-03-01

    Convent Chemical Corporation in Convent, LA produces and ships bulk quantities of chlorine and caustic soda (NaOH). The caustic soda is available in various grades, including a 50% aqueous solution that freezes at 56/sup 0/F. An extensive network of chemical-resistant polypropylene-lined steel pipe (without heat tracing) is used to transfer the caustic soda from the production area to storage tanks and to the loading facilities for tank trucks, rail tank cars and barges. A sudden drop in ambient temperature can cause freezeup of the caustic transfer pipes which may result in downtime of as much as a week. Convent plant engineers designed a pigging system for the outdoor caustic transfer lines in the tank farm and to the loading stations. The patented design pig, (internal pipeline cleaner) consists of a flexible, bullet-shaped cylinder of chemical-resistant polyurethane foam with strips of urethane rubber on the surface, or with a solid coating of the tough material. Fluid or gas pressure on the sealed concave base propels the flexible pig through the pipe, valves, elbows, and other fittings, and material ahead of the pig is discharged into an appropriate receiver. The pigging system has eliminated the caustic freezing and plugging problems since it was installed in the summer of 1981. The flexible pig, propelled by 80 psi air, is used to clear the pipelines whenever caustic is transferred during the winter months. The air-propelled pig is designed and sized to easily pass through restrictions in the piping system, such as reduced port plug valves, but was once stuck when it reached a section of pipe that had collapsed. A pig containing the Cobalt 60 nuclear element was inserted into the line to quickly locate the stuck pig with the Geiger counter. The faulty section of pipe was replaced with a new polypropylene lined spool piece.

  5. Growth failure after recurrent fever in young guinea pigs.

    PubMed

    Madu, S C; Faurie, A; Pettifor, J M; Laburn, H P

    2007-03-16

    Infection causes fever and suppression of appetite, a combination of effects which threatens normal growth in infected children. We have used an animal model to study the effects on growth of recurrent simulated Gram-positive bacterial infection. After weaning, 10 guinea pig pups underwent surgery under general anaesthesia for the implantation of temperature-sensitive radiotelemeters and thereafter were assigned to receive intramuscular injections of either 50 microg/kg muramyl dipeptide (MDP), or sterile saline. During a 30-day period corresponding to their rapid growth phase, the pups were given eight injections. MDP resulted in fevers of about 1.5 degrees C on each occasion, but no significant change in body temperature occurred after saline injections. Food intake was suppressed during each febrile episode such that 24-h intake was significantly lower on days of injections of MDP, compared to days between MDP injections in the same animals, and compared to that of animals injected with saline. The rate of weight gain of the MDP-injected guinea pigs was significantly lower than that of the control group and failed even to achieve a rate similar to the saline-injected group in their more adult-like growth phase. Plasma zinc concentration was significantly lower in MDP-compared to saline-injected animals sampled 8 days after the last injection. Our results show that recurrent fever during the growth phase of young guinea pigs results in irreversible growth failure, and that reduced food intake on days when the animals were febrile was at least partly responsible for this reduced rate of growth.

  6. PCR-based cloning and immunocytological titration of infectious porcine endogenous retrovirus subgroup A and B.

    PubMed

    Bartosch, Birke; Weiss, Robin A; Takeuchi, Yasuhiro

    2002-09-01

    Two pig endogenous retroviruses (PERV), PERV-A and -B, productively infect human cells and are therefore considered to constitute a potential risk in pig-to-human xenotransplantation. A PCR-based cloning technique to isolate infectious PERV proviruses was established. Overlapping 3' half and 5' halves of PERV proviral genomes were amplified using DNA extracted from human 293 cells infected with PERV-A or -B. These clones were fused at a unique restriction site in the overlapping region and tested for their infectivity. Representative constructs possessed the same infectious properties as their parent isolates. We also developed a polyclonal anti-PERV serum by using recombinant PERV capsid protein derived from one of the infectious constructs as immunogen and established an immunocytological method for detection and titration of PERV infection. This detection method proved to be more sensitive than the current method of choice (transfer of MLV-lacZ vectors) for infectivity assessment of PERV. These findings should be considered for future characterization of PERV isolates.

  7. Passive immunization of guinea pigs with llama single-domain antibody fragments against foot-and-mouth disease.

    PubMed

    Harmsen, M M; van Solt, C B; Fijten, H P D; van Keulen, L; Rosalia, R A; Weerdmeester, K; Cornelissen, A H M; De Bruin, M G M; Eblé, P L; Dekker, A

    2007-03-10

    Foot-and-mouth disease (FMD) is a highly contagious disease that occasionally causes outbreaks in Europe. There is a need for therapies that provide rapid protection against FMD in outbreak situations. We aim to provide such rapid protection by passive immunization with llama single-domain antibody fragments (VHHs). Twenty-four VHHs binding serotype O FMDV in vitro were isolated from immunized llamas by phage display and expressed in bakers yeast for further characterization. They recognized four functionally independent antigenic sites. Six strongly FMDV neutralizing VHHs bound to a peptide representing the GH-loop of viral protein 1 known to be involved in binding to the cellular receptor of FMDV. Clone M8, recognizing this antigenic site, and clone M23, recognizing another antigenic site, showed synergistic in vitro virus neutralization. Three FMDV specific VHHs were PEGylated in order to decrease their rapid blood clearance and thus enable in vivo guinea pig protection experiments. Passive immunization with individual VHHs showed no protection, but a mixture of M8 and M23 showed partial transient protection. The protection afforded by these VHHs was however low as compared to the complete protection afforded by convalescent guinea pig serum. In contrast, these VHHs showed far more efficient in vitro FMDV neutralization than convalescent guinea pig serum. This lack of correlation between in vitro neutralization and in vivo protection lends further credence to the notion that opsonophagocytosis of FMDV is important for protection in vivo.

  8. 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

  9. Passive immunization of guinea pigs with llama single-domain antibody fragments against foot-and-mouth disease.

    PubMed

    Harmsen, M M; van Solt, C B; Fijten, H P D; van Keulen, L; Rosalia, R A; Weerdmeester, K; Cornelissen, A H M; De Bruin, M G M; Eblé, P L; Dekker, A

    2007-03-10

    Foot-and-mouth disease (FMD) is a highly contagious disease that occasionally causes outbreaks in Europe. There is a need for therapies that provide rapid protection against FMD in outbreak situations. We aim to provide such rapid protection by passive immunization with llama single-domain antibody fragments (VHHs). Twenty-four VHHs binding serotype O FMDV in vitro were isolated from immunized llamas by phage display and expressed in bakers yeast for further characterization. They recognized four functionally independent antigenic sites. Six strongly FMDV neutralizing VHHs bound to a peptide representing the GH-loop of viral protein 1 known to be involved in binding to the cellular receptor of FMDV. Clone M8, recognizing this antigenic site, and clone M23, recognizing another antigenic site, showed synergistic in vitro virus neutralization. Three FMDV specific VHHs were PEGylated in order to decrease their rapid blood clearance and thus enable in vivo guinea pig protection experiments. Passive immunization with individual VHHs showed no protection, but a mixture of M8 and M23 showed partial transient protection. The protection afforded by these VHHs was however low as compared to the complete protection afforded by convalescent guinea pig serum. In contrast, these VHHs showed far more efficient in vitro FMDV neutralization than convalescent guinea pig serum. This lack of correlation between in vitro neutralization and in vivo protection lends further credence to the notion that opsonophagocytosis of FMDV is important for protection in vivo. PMID:17127019

  10. A guinea-pig hereditary cataract contains a splice-site deletion in a crystallin gene.

    PubMed

    Rodriguez, I R; Gonzalez, P; Zigler, J S; Borrás, T

    1992-10-13

    A congenital cataract present in guinea pigs provided a unique opportunity to study a hereditary lens disease at the molecular level. zeta-Crystallin, one of the most abundant guinea pig lens proteins, was found to be altered in the lens of cataractous animals. Several zeta-crystallin cDNA clones were isolated from a cataractous lens library and found to contain a 102-bp deletion towards the 3' end of the coding region. This deletion does not interfere with the reading frame but results in a protein 34 amino acids shorter. Sequence analysis of a normal genomic zeta-crystallin clone revealed that the missing 102-bp fragment corresponds to an entire exon (exon 7). PCR analysis of the genomic DNA isolated from cataractous animals showed that exon 7, though missing from the mRNA, is intact in the cataractous genome. Further sequence analysis of the zeta-crystallin gene disclosed a dinucleotide deletion of the universal AG at the acceptor splice-site of intron 6 of the mutant gene. The presence of this mutation results in the skipping of exon 7 during the mRNA processing which in turn results in the altered zeta-crystallin protein. This is the first time a genomic mutation in an enzyme/crystallin gene has been directly linked to a congenital cataract.

  11. 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.

  12. 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

  13. Effect of terramycin in balantidiosis of pigs.

    PubMed

    Mwamba, T; Pandey, V S

    1977-01-01

    In an industrial pig farm in Lubumashi, Zaïre, Balantidium coli produced severe clinical and fatal disease. Terramycin at a dose rate of 15 mg/kg body weight administered twice daily with concentrates gave clinical recovery in all the pigs treated. The complete parasitological recovery was obtained in 14 out of 20 animals. In the remaining 6 there was significant reduction in the number of B. coli. Keeping in view the large spectrum of activity of terramycin in various infections as well as B. coli, terramycin can be useful in the treatment of balantidiosis of pigs. PMID:596795

  14. Agro-economic impact of cattle cloning.

    PubMed

    Faber, D C; Ferre, L B; Metzger, J; Robl, J M; Kasinathan, P

    2004-01-01

    The purpose of this paper is to review the economic and social implications of cloned cattle, their products, and their offspring as related to production agriculture. Cloning technology in cattle has several applications outside of traditional production agriculture. These applications can include bio-medical applications, such as the production of pharmaceuticals in the blood or milk of transgenic cattle. Cloning may also be useful in the production of research models. These models may or may not include genetic modifications. Uses in agriculture include many applications of the technology. These include making genetic copies of elite seed stock and prize winning show cattle. Other purposes may range from "insurance" to making copies of cattle that have sentimental value, similar to cloning of pets. Increased selection opportunities available with cloning may provide for improvement in genetic gain. The ultimate goal of cloning has often been envisioned as a system for producing quantity and uniformity of the perfect dairy cow. However, only if heritability were 100%, would clone mates have complete uniformity. Changes in the environment may have significant impact on the productivity and longevity of the resulting clones. Changes in consumer preferences and economic input costs may all change the definition of the perfect cow. The cost of producing such animals via cloning must be economically feasible to meet the intended applications. Present inefficiencies limit cloning opportunities to highly valued animals. Improvements are necessary to move the applications toward commercial application. Cloning has additional obstacles to conquer. Social and regulatory acceptance of cloning is paramount to its utilization in production agriculture. Regulatory acceptance will need to address the animal, its products, and its offspring. In summary, cloning is another tool in the animal biotechnology toolbox, which includes artificial insemination, sexing of semen, embryo

  15. Agro-economic impact of cattle cloning.

    PubMed

    Faber, D C; Ferre, L B; Metzger, J; Robl, J M; Kasinathan, P

    2004-01-01

    The purpose of this paper is to review the economic and social implications of cloned cattle, their products, and their offspring as related to production agriculture. Cloning technology in cattle has several applications outside of traditional production agriculture. These applications can include bio-medical applications, such as the production of pharmaceuticals in the blood or milk of transgenic cattle. Cloning may also be useful in the production of research models. These models may or may not include genetic modifications. Uses in agriculture include many applications of the technology. These include making genetic copies of elite seed stock and prize winning show cattle. Other purposes may range from "insurance" to making copies of cattle that have sentimental value, similar to cloning of pets. Increased selection opportunities available with cloning may provide for improvement in genetic gain. The ultimate goal of cloning has often been envisioned as a system for producing quantity and uniformity of the perfect dairy cow. However, only if heritability were 100%, would clone mates have complete uniformity. Changes in the environment may have significant impact on the productivity and longevity of the resulting clones. Changes in consumer preferences and economic input costs may all change the definition of the perfect cow. The cost of producing such animals via cloning must be economically feasible to meet the intended applications. Present inefficiencies limit cloning opportunities to highly valued animals. Improvements are necessary to move the applications toward commercial application. Cloning has additional obstacles to conquer. Social and regulatory acceptance of cloning is paramount to its utilization in production agriculture. Regulatory acceptance will need to address the animal, its products, and its offspring. In summary, cloning is another tool in the animal biotechnology toolbox, which includes artificial insemination, sexing of semen, embryo

  16. Cloning and analysis of differentially expressed ESTs in swine muscle tissue.

    PubMed

    Li, Chongsheng; Chen, Yaosheng; Wang, Chong; Li, Jiaqi

    2006-08-01

    The obvious difference in muscle growth and meat quality traits exists between Chinese indigenous pig and exotic pigs. In order to study the reason of these phenotypic differences and search the potential gene related to growth and meat quality traits, silver-stained mRNA differential display technique was used to detect the difference with mRNA of loin-eye muscle tissue from maturity pigs of Lantang in Guangdong Province and Large Yorkshire. One of the newly discovered expressed sequence tag (ESTsp3) was analyzed by using bioinformatic technique. The results showed: (1) nearly 2000 cDNA fragments were detected with 30 primer pairs, and 6 differentially expressed ESTs in the loin-eye muscle tissues from the two breeds were isolated and obtained. The differential fragments were cloned and sequenced. The all sequences were recorded in the GenBank. (2) The 786 bp fragment of ESTsp3 was obtained with in silico elongation system, the ORF analysis revealed that it existed as an 83 aa complete open reading frame, and the elongation sequences were verified by RT-PCR. The analysis of in silico expression profile showed that ESTsp3 is expressed in various growth stages and in most tissues and organs, such as soft tissue, skin, skeletal muscle and kidney, but with variant expression quantity.

  17. The high-level accumulation of n-3 polyunsaturated fatty acids in transgenic pigs harboring the n-3 fatty acid desaturase gene from Caenorhabditis briggsae.

    PubMed

    Zhou, Yanrong; Lin, Yanli; Wu, Xiaojie; Feng, Chong; Long, Chuan; Xiong, Fuyin; Wang, Ning; Pan, Dengke; Chen, Hongxing

    2014-02-01

    Livestock meat is generally low in n-3 polyunsaturated fatty acids (PUFAs), which are beneficial to human health. An alternative approach to increasing the levels of n-3 PUFAs in meat is to generate transgenic livestock animals. In this study, we describe the generation of cloned pigs that express the cbr-fat-1 gene from Caenorhabditis briggsae, encoding an n-3 fatty acid desaturase. Analysis of fatty acids demonstrated that the cbr-fat-1 transgenic pigs produced high levels of n-3 fatty acids from n-6 analogs; consequently, a significantly reduced ratio of n-6/n-3 fatty acids was observed. We demonstrated that the n-3 desaturase gene from C. briggsae was functionally expressed, and had a significant effect on the fatty acid composition of the transgenic pigs, which may allow the production of pork enriched in n-3 PUFAs.

  18. Analysis of the temperature sensitivity of Japanese rubella vaccine strain TO-336.vac and its effect on immunogenicity in the guinea pig.

    PubMed

    Okamoto, Kiyoko; Ami, Yasushi; Suzaki, Yuriko; Otsuki, Noriyuki; Sakata, Masafumi; Takeda, Makoto; Mori, Yoshio

    2016-04-01

    The marker of Japanese domestic rubella vaccines is their lack of immunogenicity in guinea pigs. This has long been thought to be related to the temperature sensitivity of the viruses, but supporting evidence has not been described. In this study, we generated infectious clones of TO-336.vac, a Japanese domestic vaccine, TO-336.GMK5, the parental virus of TO-336.vac, and their mutants, and determined the molecular bases of their temperature sensitivity and immunogenicity in guinea pigs. The results revealed that Ser(1159) in the non-structural protein-coding region was responsible for the temperature sensitivity of TO-336.vac dominantly, while the structural protein-coding region affected the temperature sensitivity subordinately. The findings further suggested that the temperature sensitivity of TO-336.vac affected the antibody induction in guinea pigs after subcutaneous inoculation.

  19. 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.

  20. CDX2 increases SLC7A7 expression and proliferation of pig intestinal epithelial cells

    PubMed Central

    Li, Xiang-guang; Xu, Gao-feng; Zhai, Zhen-ya; Gao, Chun-qi; Yan, Hui-chao; Xi, Qian-yun; Guan, Wu-tai; Wang, Song-bo; Wang, Xiu-qi

    2016-01-01

    Nutrient absorption mediated by nutrient transporters expressed in the intestinal epithelium supplies substrates to support intestinal processes, including epithelial cell proliferation. We evaluated the role of Caudal type homeobox 2 (CDX2), an intestine-specific transcription factor, in the proliferation of pig intestinal epithelial cells (IPEC-1) and searched for novel intestinal nutrient transporter genes activated by CDX2. Our cloned pig CDX2 cDNA contains a “homeobox” DNA binding motif, suggesting it is a transcriptional activator. CDX2 overexpression in IPEC-1 cells increased cell proliferation, the percentage of cells in S/G2 phase, and the abundance of transcripts of the cell cycle-related genes Cyclin A2; Cyclin B; Cyclin D2; proliferating cell nuclear antigen; and cell cycle cyclin-dependent kinases 1, 2 and 4, as well as the predicted CDX2 target genes SLC1A1, SLC5A1 and SLC7A7. In addition, luciferase reporter and chromatin immunoprecipitation assays revealed that CDX2 binds directly to the SLC7A7 promoter. This is the first report of CDX2 function in pig intestinal epithelial cells and identifies SLC7A7 as a novel CDX2 target gene. Our findings show that nutrient transporters are activated during CDX2-induced proliferation of normal intestinal epithelial cells. PMID:27121315

  1. Comparison of market hog characteristics of pigs selected by feeder pig frame size or current USDA feeder pig grade standards.

    PubMed

    Siemens, A L; Lipsey, R J; Hedrick, H B; Williams, F L; Yokley, S W; Siemens, M G

    1990-08-01

    Two feeder pig grading systems were tested. Forty-five barrows were selected using current USDA Feeder Pig Grade Standards (U.S. No. 1, No. 2 and No. 3). Additionally, 45 barrows were selected using three frame sizes (large, medium and small). Pigs were slaughtered at 100, 113.5 of 127 kg live weight. Trimmed four lean cuts were separated into soft tissue, skin and bone. The skinless belly and soft tissue from the four lean cuts were ground separately and analyzed chemically. Data from each grading system were analyzed separately in a 3 X 3 factorial plan. Pigs selected using current USDA grade standards differed (P less than .05) for last rib backfat, 10th rib fat depth, longissimus muscle area, percentage of trimmed four lean cuts and USDA carcass grade. In the frame size system, pigs with large frame size had less last rib backfat, less 10th rib fat depth, longer carcasses, higher percentage of four lean cuts and superior USDA carcass grades than pigs with small frame size did (P less than .05). The Bradley and Schumann test of sensitivity showed that selection by frame size was more sensitive than current USDA grade standards for discriminating feeder pig foreleg length, body depth and ham width. In addition, selection by frame size was more sensitive than current USDA grade standards for discriminating carcass length and carcass radius length. No increase in sensitivity (P greater than .10) was noted for carcass composition or growth traits over the current USDA Feeder Pig Grade Standards.

  2. Cloning

    MedlinePlus

    ... mammals. These twins are produced when a fertilized egg splits, creating two or more embryos that carry ... of the donor animal's somatic cell into an egg cell, or oocyte, that has had its own ...

  3. Cloning and expression of new microRNAs from zebrafish

    PubMed Central

    Kloosterman, Wigard P.; Steiner, Florian A.; Berezikov, Eugene; de Bruijn, Ewart; van de Belt, Jose; Verheul, Mark; Cuppen, Edwin; Plasterk, Ronald H.A.

    2006-01-01

    MicroRNAs (miRNAs) play an important role in development and regulate the expression of many animal genes by post-transcriptional gene silencing. Here we describe the cloning and expression of new miRNAs from zebrafish. By high-throughput sequencing of small-RNA cDNA libraries from 5-day-old zebrafish larvae and adult zebrafish brain we found 139 known miRNAs and 66 new miRNAs. For 65 known miRNAs and for 11 new miRNAs we also cloned the miRNA star sequence. We analyzed the temporal and spatial expression patterns for 35 new miRNAs and for 32 known miRNAs in the zebrafish by whole mount in situ hybridization and northern blotting. Overall, 23 of the 35 new miRNAs and 30 of the 32 known miRNAs could be detected. We found that most miRNAs were expressed during later stages of development. Some were expressed ubiquitously, but many of the miRNAs were expressed in a tissue-specific manner. Most newly discovered miRNAs have low expression levels and are less conserved in other vertebrate species. Our cloning and expression analysis indicates that most abundant and conserved miRNAs in zebrafish are now known. PMID:16698962

  4. Detectability of Plasmodium falciparum clones

    PubMed Central

    2010-01-01

    Background In areas of high transmission people often harbour multiple clones of Plasmodium falciparum, but even PCR-based diagnostic methods can only detect a fraction (the detectability, q) of all clones present in a host. Accurate measurements of detectability are desirable since it affects estimates of multiplicity of infection, prevalence, and frequency of breakthrough infections in clinical drug trials. Detectability can be estimated by typing repeated samples from the same host but it has been unclear what should be the time interval between the samples and how the data should be analysed. Methods A longitudinal molecular study was conducted in the Kassena-Nankana district in northern Ghana. From each of the 80 participants, four finger prick samples were collected over a period of 8 days, and tested for presence of different Merozoite Surface Protein (msp) 2 genotypes. Implications for estimating q were derived from these data by comparing the fit of statistical models of serial dependence and over-dispersion. Results The distribution of the frequencies of detection for msp2 genotypes was close to binomial if the time span between consecutive blood samples was at least 7 days. For shorter intervals the probabilities of detection were positively correlated, i.e. the shorter the interval between two blood collections, the more likely the diagnostic results matched for a particular genotype. Estimates of q were rather insensitive to the statistical model fitted. Conclusions A simple algorithm based on analysing blood samples collected 7 days apart is justified for generating robust estimates of detectability. The finding of positive correlation of detection probabilities for short time intervals argues against imperfect detection being directly linked to the 48-hour periodicity of P. falciparum. The results suggest that the detectability of a given parasite clone changes over time, at an unknown rate, but fast enough to regard blood samples taken one week

  5. Pregnancies established from handmade cloned blastocysts reconstructed using skin fibroblasts in buffalo (Bubalus bubalis).

    PubMed

    Shah, R A; George, A; Singh, M K; Kumar, D; Anand, T; Chauhan, M S; Manik, R S; Palta, P; Singla, S K

    2009-05-01

    Handmade cloning (HMC), a simple, micromanipulation-free cloning technique, has been applied for the production of cloned embryos and offspring in many livestock species. The objective of the present study was to compare the effect of donor cell type on developmental competence of HMC embryos and to explore the possibility of establishing pregnancies using these embryos in buffalo. After technical optimization of the HMC procedure for in vitro development of cloned blastocysts, various donor cells were compared for their developmental efficiency. Using buffalo fetal-, newborn-, adult fibroblasts and cumulus cells, blastocyst production rates obtained from reconstructed embryos were 24.0+/-1.8% (35/145), 33.0+/-8.0% (56/163), 21.0+/-9.3% (29/133) and 49.6+/-1.9% (77/154), respectively. Blastocyst rates were higher (P<0.05) in cumulus cell reconstructed embryos in comparison to those derived from fetal or adult fibroblasts. Pregnancy diagnosis (transrectal ultrasonography) was carried out at Day 40 of gestation. Following transfer of HMC embryos reconstructed using newborn fibroblasts 25% (2/8) buffaloes were pregnant and are at Days 201 and 94 of gestation, whereas after transfer of HMC embryos reconstructed using fetal fibroblasts, 20% (1/5) buffaloes were pregnant and are at Day 73 of gestation. In conclusion, HMC could be a simple and efficient technique for the production of cloned embryos for establishing pregnancies in buffalo.

  6. Unified universal quantum cloning machine and fidelities

    SciTech Connect

    Wang Yinan; Shi Handuo; Xiong Zhaoxi; Jing Li; Mu Liangzhu; Ren Xijun; Fan Heng

    2011-09-15

    We present a unified universal quantum cloning machine, which combines several different existing universal cloning machines together, including the asymmetric case. In this unified framework, the identical pure states are projected equally into each copy initially constituted by input and one half of the maximally entangled states. We show explicitly that the output states of those universal cloning machines are the same. One importance of this unified cloning machine is that the cloning procession is always the symmetric projection, which reduces dramatically the difficulties for implementation. Also, it is found that this unified cloning machine can be directly modified to the general asymmetric case. Besides the global fidelity and the single-copy fidelity, we also present all possible arbitrary-copy fidelities.

  7. Effective immune protection of pigs against cysticercosis.

    PubMed

    Nascimento, E; Costa, J O; Guimarães, M P; Tavares, C A

    1995-03-01

    A scolex protein antigen (SPA) was prepared from cysticerci of Taenia solium obtained from naturally infected pigs. Yorkshire pigs were vaccinated with SPA plus incomplete Freund's adjuvant (IFA) or with SPA plus Corynebacterium parvum (CP). Controls were given IFA plus phosphate-buffered saline (PBS) or CP plus PBS. All animals were given three subcutaneous injections at 20-day intervals. Ten days after the third injection, the pigs were fed with 10(4) viable eggs of T. solium. All pigs developed a delayed type hypersensitivity, and a transient eosinophilia after the first dose of vaccine. High titers of specific antibodies were detected in the sera of vaccinated animals and in infected controls. A protection level of 71.43% was recorded in animals vaccinated with SPA plus IFA and of 75.00% in those vaccinated with SPA plus CP.

  8. Swine Influenza (Swine Flu) in Pigs

    MedlinePlus

    ... in the United States since 2005 Prevention Treatment Influenza Types Seasonal Avian Swine Variant Pandemic Other Get ... Submit Button Past Newsletters Key Facts about Swine Influenza (Swine Flu) in Pigs Language: English Español ...

  9. Effective immune protection of pigs against cysticercosis.

    PubMed

    Nascimento, E; Costa, J O; Guimarães, M P; Tavares, C A

    1995-03-01

    A scolex protein antigen (SPA) was prepared from cysticerci of Taenia solium obtained from naturally infected pigs. Yorkshire pigs were vaccinated with SPA plus incomplete Freund's adjuvant (IFA) or with SPA plus Corynebacterium parvum (CP). Controls were given IFA plus phosphate-buffered saline (PBS) or CP plus PBS. All animals were given three subcutaneous injections at 20-day intervals. Ten days after the third injection, the pigs were fed with 10(4) viable eggs of T. solium. All pigs developed a delayed type hypersensitivity, and a transient eosinophilia after the first dose of vaccine. High titers of specific antibodies were detected in the sera of vaccinated animals and in infected controls. A protection level of 71.43% was recorded in animals vaccinated with SPA plus IFA and of 75.00% in those vaccinated with SPA plus CP. PMID:7604530

  10. Development and validation of a highly efficient protocol of porcine somatic cloning using preovulatory embryo transfer in peripubertal gilts.

    PubMed

    Petersen, Björn; Lucas-Hahn, Andrea; Oropeza, Marianne; Hornen, Nadine; Lemme, Erika; Hassel, Petra; Queisser, Anna-Lisa; Niemann, Heiner

    2008-09-01

    The efficiency of porcine somatic nuclear transfer (born piglets/transferred embryos) is low. Here, we report a highly efficient protocol using peripubertal gilts as recipients synchronized to ovulate approximately 24 h after transfer of cloned embryos. Retrospectively, we compared the efficiency of two different synchronization protocols: In group 1, recipient animals were synchronized to ovulate approximately 6 h prior to surgical embryo transfer while in group 2 the animals were treated to ovulate 24 h after embryo transfer. In total, 1562 cloned embryos were transferred to 12 recipients in group 1; two of them became pregnant (16.7%). One pregnancy was lost on day 32, the second pregnancy went to term, and led to the birth of one healthy piglet after Cesarean section. In group 2, 1531 cloned embryos were transferred to 12 recipients. Nine recipients (75.0%) became pregnant as determined by ultrasound scanning on day 25. All pregnancies went to term and delivered a total of 47 live-born piglets. The cloning efficiency of both groups differed significantly (group 1: 0.1%, group 2: 3.1%, p < 0.05). This modified protocol was then applied in subsequent experiments using different types of transgenic and nontransgenic donor cells with similar success rates. Results show that this protocol is robust and highly reproducible, and can thus be employed for routine production of cloned pigs.

  11. No human cloning: a social ethics perspective.

    PubMed

    Cahill, L S

    1999-01-01

    This Essay addresses the negative impact of human cloning on the family, and argues further that market incentives to develop and implement cloning techniques exploit and exacerbate socioeconomic inequities. It suggests that cloning should be prohibited internationally and examines possible routes to that aim. To begin with, it offers some reflections on the nature of moral argument, and on the role of religion in public debate. PMID:12650145

  12. Cloning: pathways to a pluripotent future.

    PubMed

    McLaren, A

    2000-06-01

    In this month's essay, Anne McLaren traces the winding and pitted pathways that connect the early days of the cell theory of biology in the 1830s to the new and unfolding era of cloning science and technology that came to worldwide attention in 1997 with the announcement of the birth of Dolly, the Scottish cloned sheep. The possibilities, including the potential for new medical treatments and perhaps even human cloning, are fantastic ... and ethically charged.

  13. Telomeres and the ethics of human cloning.

    PubMed

    Allhoff, Fritz

    2004-01-01

    In search of a potential problem with cloning, I investigate the phenomenon of telomere shortening which is caused by cell replication; clones created from somatic cells will have shortened telomeres and therefore reach a state of senescence more rapidly. While genetic intervention might fix this problem at some point in the future, I ask whether, absent technological advances, this biological phenomenon undermines the moral permissibility of cloning.

  14. Cloning: pathways to a pluripotent future.

    PubMed

    McLaren, A

    2000-06-01

    In this month's essay, Anne McLaren traces the winding and pitted pathways that connect the early days of the cell theory of biology in the 1830s to the new and unfolding era of cloning science and technology that came to worldwide attention in 1997 with the announcement of the birth of Dolly, the Scottish cloned sheep. The possibilities, including the potential for new medical treatments and perhaps even human cloning, are fantastic ... and ethically charged. PMID:10877698

  15. Experimental evidence of hepatitis A virus infection in pigs.

    PubMed

    Song, Young-Jo; Park, Woo-Jung; Park, Byung-Joo; Kwak, Sang-Woo; Kim, Yong-Hyeon; Lee, Joong-Bok; Park, Seung-Yong; Song, Chang-Seon; Lee, Sang-Won; Seo, Kun-Ho; Kang, Young-Sun; Park, Choi-Kyu; Song, Jae-Young; Choi, In-Soo

    2016-04-01

    Hepatitis A virus (HAV) is the leading cause of acute viral hepatitis worldwide, with HAV infection being restricted to humans and nonhuman primates. In this study, HAV infection status was serologically determined in domestic pigs and experimental infections of HAV were attempted to verify HAV infectivity in pigs. Antibodies specific to HAV or HAV-like agents were detected in 3.5% of serum samples collected from pigs in swine farms. When the pigs were infected intravenously with 2 × 10(5) 50% tissue culture infectious dose (TCID50 ) of HAV, shedding of the virus in feces, viremia, and seroconversion were detected. In pigs orally infected with the same quantity of HAV, viral shedding was detected only in feces. HAV genomic RNA was detected in the liver and bile of intravenously infected pigs, but only in the bile of orally infected pigs. In further experiments, pigs were intravenously infected with 6 × 10(5) TCID50 of HAV. Shedding of HAV in feces, along with viremia and seroconversion, were confirmed in infected pigs but not in sentinel pigs. HAV genomic RNA was detected in the liver, bile, spleen, lymph node, and kidney of the infected pigs. HAV antigenomic RNA was detected in the spleen of one HAV-infected pig, suggesting HAV replication in splenic cells. Infiltration of inflammatory cells was observed in the livers of infected pigs but not in controls. This is the first experimental evidence to demonstrate that human HAV strains can infect pigs.

  16. Assessing pig body language: agreement and consistency between pig farmers, veterinarians, and animal activists.

    PubMed

    Wemelsfelder, F; Hunter, A E; Paul, E S; Lawrence, A B

    2012-10-01

    This study investigates the interobserver and intraobserver reliability of qualitative behavior assessments (QBA) of individual pigs by 3 observer groups selected for their diverging backgrounds, experience, and views of pigs. Qualitative behavior assessment is a "whole animal" assessment approach that characterizes the demeanor of an animal as an expressive body language, using descriptors such as relaxed, anxious, or content. This paper addresses the concern that use of such descriptors in animal science may be prone to distortion by observer-related bias. Using a free-choice profiling methodology, 12 pig farmers, 10 large animal veterinarians, and 10 animal protectionists were instructed to describe and score the behavioral expressions of 10 individual pigs (sus scrofa) in 2 repeat sets of 10 video clips, showing these pigs in interaction with a human female. They were also asked to fill in a questionnaire gauging their experiences with and views on pigs. Pig scores were analyzed with generalized procrustes analysis and effect of treatment on these scores with ANOVA. Questionnaire scores were analyzed with a χ(2) test or ANOVA. Observers achieved consensus both within and among observer groups (P < 0.001), identifying 2 main dimensions of pig expression (dim1: playful/confident-cautious/timid; dim2: aggressive/nervous-relaxed/bored), on which pig scores for different observer groups were highly correlated (pearson r > 0.90). The 3 groups also repeated their assessments of individual pigs with high precision (r > 0.85). Animal protectionists used a wider quantitative range in scoring individual pigs on dimension 2 than the other groups (P < 0.001); however, this difference did not distort the strong overall consistency of characterizations by observers of individual pigs. Questionnaire results indicated observer groups to differ in various ways, such as daily and lifetime contact with pigs (P < 0.001), some aspects of affection and empathy for pigs (P < 0

  17. Single-nucleotide polymorphisms and activity analysis of the promoter and enhancer of the pig lactase gene.

    PubMed

    Du, Hai-Ting; Zhu, Hong-Yan; Wang, Jia-Mei; Zhao, Wei; Tao, Xiao-Li; Ba, Cai-Feng; Tian, Yu-Min; Su, Yu-Hong

    2014-07-15

    Lactose intolerance in northern Europeans is strongly associated with a single-nucleotide polymorphism (SNP) located 14 kb upstream of the human lactase gene: -13,910 C/T. We examined whether SNPs in the 5' flanking region of the pig lactase gene are similar to those in the human gene and whether these polymorphisms play a functional role in regulating pig lactase gene expression. The 5' flanking region of the lactase gene from several different breeds of pigs was cloned and analyzed for gene regulatory activity of a luciferase reporter gene. One SNP was found in the enhancer region (-797 G/A) and two were found in the promoter region (-308G/C and -301 A/G). The promoter C-308,G-301(Pro-CG) strongly promotes the expression of the lactase gene, but the promoter G-308,A-301(Pro-GA) does not. The enhancer A-797(Enh-A) genotype for Pro-GA can significantly enhance promoter activity, but has an inhibitory effect on Pro-CG. The Enhancer G-797(Enh-G) has a significant inhibitory effect on both promoters. In conclusion, the order of effectiveness on the pig lactase gene is Enh-A+Pro-GA>Enh-A/G+Pro-CG>Enh-G+Pro-GA.

  18. Single-nucleotide polymorphisms and activity analysis of the promoter and enhancer of the pig lactase gene.

    PubMed

    Du, Hai-Ting; Zhu, Hong-Yan; Wang, Jia-Mei; Zhao, Wei; Tao, Xiao-Li; Ba, Cai-Feng; Tian, Yu-Min; Su, Yu-Hong

    2014-07-15

    Lactose intolerance in northern Europeans is strongly associated with a single-nucleotide polymorphism (SNP) located 14 kb upstream of the human lactase gene: -13,910 C/T. We examined whether SNPs in the 5' flanking region of the pig lactase gene are similar to those in the human gene and whether these polymorphisms play a functional role in regulating pig lactase gene expression. The 5' flanking region of the lactase gene from several different breeds of pigs was cloned and analyzed for gene regulatory activity of a luciferase reporter gene. One SNP was found in the enhancer region (-797 G/A) and two were found in the promoter region (-308G/C and -301 A/G). The promoter C-308,G-301(Pro-CG) strongly promotes the expression of the lactase gene, but the promoter G-308,A-301(Pro-GA) does not. The enhancer A-797(Enh-A) genotype for Pro-GA can significantly enhance promoter activity, but has an inhibitory effect on Pro-CG. The Enhancer G-797(Enh-G) has a significant inhibitory effect on both promoters. In conclusion, the order of effectiveness on the pig lactase gene is Enh-A+Pro-GA>Enh-A/G+Pro-CG>Enh-G+Pro-GA. PMID:24809963

  19. Naegleria: another pathogenic ameba studies in germfree guinea pigs.

    PubMed

    Phillips, B P

    1974-09-01

    Free-living amebas of the genus Naegleria, of world-wide distribution and long considered harmless, have been linked etiologically with 57 fatal cases of primary amebic meningoencephalitis during the last decade. Naegleria from cultures derived from one of these fatal cases in Richmond, Virginia, have been inoculated intranasally, intraorally, into the conjunctival sac near the inner canthus of the eyes, and into induced skin lesions in adult germfree guinea pigs. Of 33 animals inoculated intranasally with 18 to 31 amebas, 31 developed a fatal encephalitis. There was considerable destruction of tissues of the cerebellum and the cerebrum and including the olfactory lobes. The meninges were involved to varying degrees in most of the animals. None of the animals inoculated by the three other routes developed either symptoms or lesions. PMID:4451226

  20. Cloning of spin-coherent states

    SciTech Connect

    Demkowicz-Dobrzanski, Rafal; Kus, Marek; Wodkiewicz, Krzysztof

    2004-01-01

    We consider optimal cloning of the spin coherent states in Hilbert spaces of different dimensionality d. We give explicit form of optimal cloning transformation for spin coherent states in the three-dimensional space, analytical results for the fidelity of the optimal cloning in d=3 and d=4 as well as numerical results for higher dimensions. In the low-dimensional case we construct the corresponding completely positive maps and exhibit their structure with the help of Jamiolkowski isomorphism. This allows us to formulate some conjectures about the form of optimal coherent cloning completely positive maps in arbitrary dimension.