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Sample records for recombinant gam82 gametocyte

  1. Eimeria maxima recombinant Gam82 gametocyte antigen vaccine protects against coccidiosis and augments humoral and cell-mediated immunity

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Intestinal infection with Eimeria, the etiologic agent of avian coccidiosis, stimulates protective immunity to subsequent colonization by the homologous parasite, whilst cross-protection against heterologous species is poor. As a first step toward the development of a broad specificity Eimeria vacci...

  2. Perforin-like protein PPLP2 permeabilizes the red blood cell membrane during egress of Plasmodium falciparum gametocytes

    PubMed Central

    Wirth, Christine C; Glushakova, Svetlana; Scheuermayer, Matthias; Repnik, Urska; Garg, Swati; Schaack, Dominik; Kachman, Marika M; Weißbach, Tim; Zimmerberg, Joshua; Dandekar, Thomas; Griffiths, Gareth; Chitnis, Chetan E; Singh, Shailja; Fischer, Rainer; Pradel, Gabriele

    2014-01-01

    Egress of malaria parasites from the host cell requires the concerted rupture of its enveloping membranes. Hence, we investigated the role of the plasmodial perforin-like protein PPLP2 in the egress of Plasmodium falciparum from erythrocytes. PPLP2 is expressed in blood stage schizonts and mature gametocytes. The protein localizes in vesicular structures, which in activated gametocytes discharge PPLP2 in a calcium-dependent manner. PPLP2 comprises a MACPF domain and recombinant PPLP2 has haemolytic activities towards erythrocytes. PPLP2-deficient [PPLP2(−)] merozoites show normal egress dynamics during the erythrocytic replication cycle, but activated PPLP2(−) gametocytes were unable to leave erythrocytes and stayed trapped within these cells. While the parasitophorous vacuole membrane ruptured normally, the activated PPLP2(−) gametocytes were unable to permeabilize the erythrocyte membrane and to release the erythrocyte cytoplasm. In consequence, transmission of PPLP2(−) parasites to the Anopheles vector was reduced. Pore-forming equinatoxin II rescued both PPLP2(−) gametocyte exflagellation and parasite transmission. The pore sealant Tetronic 90R4, on the other hand, caused trapping of activated wild-type gametocytes within the enveloping erythrocytes, thus mimicking the PPLP2(−) loss-of-function phenotype. We propose that the haemolytic activity of PPLP2 is essential for gametocyte egress due to permeabilization of the erythrocyte membrane and depletion of the erythrocyte cytoplasm. PMID:24602217

  3. Strategies for Detection of Plasmodium species Gametocytes

    PubMed Central

    Javati, Sarah; Robinson, Leanne; Betuela, Inoni; Siba, Peter; Beck, Hans-Peter; Mueller, Ivo; Felger, Ingrid

    2013-01-01

    Carriage and density of gametocytes, the transmission stages of malaria parasites, are determined for predicting the infectiousness of humans to mosquitoes. This measure is used for evaluating interventions that aim at reducing malaria transmission. Gametocytes need to be detected by amplification of stage-specific transcripts, which requires RNA-preserving blood sampling. For simultaneous, highly sensitive quantification of both, blood stages and gametocytes, we have compared and optimized different strategies for field and laboratory procedures in a cross sectional survey in 315 5-9 yr old children from Papua New Guinea. qRT-PCR was performed for gametocyte markers pfs25 and pvs25, Plasmodium species prevalence was determined by targeting both, 18S rRNA genes and transcripts. RNA-based parasite detection resulted in a P. falciparum positivity of 24.1%; of these 40.8% carried gametocytes. P. vivax positivity was 38.4%, with 38.0% of these carrying gametocytes. Sensitivity of DNA-based parasite detection was substantially lower with 14.1% for P. falciparum and 19.6% for P. vivax. Using the lower DNA-based prevalence of asexual stages as a denominator increased the percentage of gametocyte-positive infections to 59.1% for P. falciparum and 52.4% for P. vivax. For studies requiring highly sensitive and simultaneous quantification of sexual and asexual parasite stages, 18S rRNA transcript-based detection saves efforts and costs. RNA-based positivity is considerably higher than other methods. On the other hand, DNA-based parasite quantification is robust and permits comparison with other globally generated molecular prevalence data. Molecular monitoring of low density asexual and sexual parasitaemia will support the evaluation of effects of up-scaled antimalarial intervention programs and can also inform about small scale spatial variability in transmission intensity. PMID:24312682

  4. Plasmodium falciparum Gametocyte-Specific Antibody Profiling Reveals Boosting through Natural Infection and Identifies Potential Markers of Gametocyte Exposure.

    PubMed

    Skinner, Jeff; Huang, Chiung-Yu; Waisberg, Michael; Felgner, Philip L; Doumbo, Ogobara K; Ongoiba, Aissata; Kayentao, Kassoum; Traore, Boubacar; Crompton, Peter D; Williamson, Kim C

    2015-11-01

    Malaria elimination efforts would benefit from vaccines that block transmission of Plasmodium falciparum gametocytes from humans to mosquitoes. A clear understanding of gametocyte-specific antibody responses in exposed populations could help determine whether transmission-blocking vaccines (TBV) would be boosted by natural gametocyte exposure, and also inform the development of serologic tools to monitor gametocyte exposure in populations targeted for malaria elimination. To this end, plasma was collected from Malian children and adults before and after the 6-month malaria season and probed against a microarray containing 1,204 P. falciparum proteins. Using publicly available proteomic data, we classified 91 proteins as gametocyte specific and 69 as proteins not expressed by gametocytes. The overall breadth and magnitude of gametocyte-specific IgG responses increased during the malaria season, although they were consistently lower than IgG responses to nongametocyte antigens. Notably, IgG specific for the TBV candidates Pfs48/45 and Pfs230 increased during the malaria season. In addition, IgGs specific for the gametocyte proteins Pfmdv1, Pfs16, PF3D7_1346400, and PF3D7_1024800 were detected in nearly all subjects, suggesting that seroconversion to these proteins may be a sensitive indicator of gametocyte exposure, although further studies are needed to determine the specificity and kinetics of these potential serologic markers. These findings suggest that TBV-induced immunity would be boosted through natural gametocyte exposure, and that antibody responses to particular antigens may reliably indicate gametocyte exposure. PMID:26283330

  5. Identification of inhibitors of Plasmodium falciparum gametocyte development

    PubMed Central

    2013-01-01

    Background Plasmodium falciparum gametocytes, specifically mature stages, are the only stage in man transmissible to the mosquito vector responsible for malaria transmission. Anti-malarial drugs capable of killing these forms are considered essential for the eradication of malaria. The comprehensive profiling of in vitro activity of anti-malarial compounds against both early (I-III) and late (IV-V) stage P. falciparum gametocytes, along with the high throughput screening (HTS) outcomes from the MMV malaria box are described. Method Two anti-gametocyte HTS assays based on confocal fluorescence microscopy, utilizing both a gametocyte specific protein (pfs16-Luc-GFP) and a viability marker (MitoTracker Red CM-H2XRos) (MTR), were used for the measurement of anti-gametocytocidal activity. This combination provided a direct observation of gametocyte number per assay well, whilst defining the viability of each gametocyte imaged. Results IC50 values were obtained for 36 current anti-malarial compounds for activities against asexual, early and late stage gametocytes. The MMV malaria box was screened and actives progressed for IC50 evaluation. Seven % of the “drug-like” and 21% of the “probe-like” compounds from the MMV malaria box demonstrated equivalent activity against both asexual and late stage gametocytes. Conclusions The assays described were shown to selectively identify compounds with gametocytocidal activity and have been demonstrated suitable for HTS with the capability of screening in the order of 20,000 compounds per screening campaign, two to three times per seven-day week. PMID:24206914

  6. The extravascular compartment of the bone marrow: a niche for Plasmodium falciparum gametocyte maturation?

    PubMed Central

    2012-01-01

    Background Plasmodium falciparum immature gametocytes accumulate in the bone marrow, but their exact location in this tissue remains unclear. Methods The stage and deposition pattern of gametocytes was analysed on histological sections of a bone marrow sample collected in a patient with subacute P. falciparum malaria. Results A majority (89%) of immature stages II to IV gametocytes and a minority (29%) of mature stage V gametocytes were observed in extravascular spaces. Discussion and conclusion These observations represent a valuable step towards understanding sequestration patterns of P. falciparum gametocytes and may ultimately lead to novel transmission-blocking interventions. PMID:22905863

  7. How important is gametocyte clearance after malaria therapy?

    PubMed

    Karunajeewa, Harin A; Mueller, Ivo

    2016-01-01

    There has been increasing interest in the role of malaria drugs in preventing malaria transmission from humans to mosquitoes, which would help augment malaria control and elimination strategies. Nevertheless, only one stage in the malaria parasite life cycle, the gametocyte, is infectious to mosquitoes. The Worldwide Antimalarial Resistance Network (WWARN) have analyzed data from 48,840 patients from 141 clinical trials in order to define the nature and determinants of gametocyte clearance following artemisinin combination treatment (ACT) for symptomatic malaria infections. However, the presence of gametocytes does not always predict their infectivity, meaning that the microscopy-based methods used by the WWARN investigators represent an imperfect surrogate marker of transmissibility. Their findings, that some ACTs clear gametocytes faster than others, should be interpreted in light of these limitations and important gaps in our understanding of the biology and epidemiology of malaria transmission.Please see related article: https://bmcmedicine.biomedcentral.com/articles/10.1186/s12916-016-0621-7. PMID:27317420

  8. The fine structure of elongate gametocytes of Leucocytozoon ziemanni (Laveran).

    PubMed

    Kocan, A A; Kocan, K M

    1978-12-01

    In an effort to establish comparative data within the genus Leucocytozoon, elongate gametocytes of L. ziemanni from naturally infected great horned owls (Bubo virginianus) were examined by electron microscopy. Micro- and macrogametocytes proved to be easily distinguishable at the electron microscopic level due to dramatic dimorphism at maturity and cytoplasmic and nuclear morphology. The parasite membrane architecture, number and type of cytoplasmic ribosomes of both micro- and macrogametocytes, presence and arrangement of osmiophilic bodies and electron dense spheres, mitochondrial morphology, endoplasmic reticulum cisternae morphology, mitochondria containing pocket infoldings of the nuclear membrane of the microgametocytes, and cytostome and food vacuole formation compare favorably with available information on L. simondi and L. smithi. Comparative variations exist only in that L. ziemanni gametocytes apparently lack compartmentalization of the cytoplasm by aligned unit membranes and parasite induced separations of the host cell nucleus as reported for L. simondi. PMID:105117

  9. An assay to probe Plasmodium falciparum growth, transmission stage formation and early gametocyte development

    PubMed Central

    Brancucci, Nicolas M B; Goldowitz, Ilana; Buchholz, Kathrin; Werling, Kristine; Marti, Matthias

    2015-01-01

    Conversion from asexual proliferation to sexual differentiation initiates the production of the gametocyte, which is the malaria parasite stage required for human-to-mosquito transmission. This protocol describes an assay designed to probe the effect of drugs or other perturbations on asexual replication, sexual conversion and early gametocyte development in the major human malaria parasite Plasmodium falciparum. Synchronized asexually replicating parasites are induced for gametocyte production by the addition of conditioned medium, and they are then exposed to the treatment of interest during sexual commitment or at any subsequent stage of early gametocyte development. Flow cytometry is used to measure asexual proliferation and gametocyte production via DNA dye staining and the gametocyte-specific expression of a fluorescent protein, respectively. This screening approach may be used to identify and evaluate potential transmission-blocking compounds and to further investigate the mechanism of sexual conversion in malaria parasites. The full protocol can be completed in 11 d. PMID:26134953

  10. Gametocyte clearance dynamics following oral artesunate treatment of uncomplicated falciparum malaria in Malian children

    PubMed Central

    Djimde, Abdoulaye A.; Maiga, Amelia W.; Ouologuem, Dinkorma; Fofana, Bakary; Sagara, Issaka; Dembele, Demba; Toure, Sekou; Sanogo, Kassim; Dama, Souleymane; Sidibe, Bakary; Doumbo, Ogobara K.

    2016-01-01

    Artemisinin-based combination therapies decrease Plasmodium gametocyte carriage. However, the role of artesunate in monotherapy in vivo, the mechanisms involved, and the utility of gametocyte carriage as a potential tool for the surveillance of antimalarial resistance are poorly understood. In 2010–2011, we conducted an open-label, prospective efficacy study of artesunate as monotherapy in children 1–10 years of age with uncomplicated falciparum malaria in Bougoula-Hameau, Mali. Standard oral doses of artesunate were administered for 7 days and patients were followed up for 28 days. The data were compared to a similar study conducted in 2002–2004. Of 100 children enrolled in the 2010–2011 study, 92 were analyzed and compared to 217 children enrolled in the 2002–2004 study. The proportion of gametocyte carriers was unchanged at the end of treatment (23% at baseline vs. 24% on day 7, p = 1.0) and did not significantly decline until day 21 of follow-up (23% vs. 6%, p = 0.003). The mean gametocyte density at inclusion remained unchanged at the end of treatment (12 gametocytes/μL vs. 16 gametocytes/μL, p = 0.6). Overall, 46% of the 71 initial non-carriers had gametocytes detected by day 7. Similar results were found in the 2002–2004 study. In both studies, although gametocyte carriage significantly decreased by the end of the 28-day follow-up, artesunate did not clear mature gametocytes during treatment and did not prevent the appearance of new stage V gametocytes as assessed by light microscopy. Baseline gametocyte carriage was significantly higher 6 years after the deployment of artemisinin-based combination therapies in this setting. PMID:26839003

  11. Gametocyte clearance dynamics following oral artesunate treatment of uncomplicated falciparum malaria in Malian children.

    PubMed

    Djimde, Abdoulaye A; Maiga, Amelia W; Ouologuem, Dinkorma; Fofana, Bakary; Sagara, Issaka; Dembele, Demba; Toure, Sekou; Sanogo, Kassim; Dama, Souleymane; Sidibe, Bakary; Doumbo, Ogobara K

    2016-01-01

    Artemisinin-based combination therapies decrease Plasmodium gametocyte carriage. However, the role of artesunate in monotherapy in vivo, the mechanisms involved, and the utility of gametocyte carriage as a potential tool for the surveillance of antimalarial resistance are poorly understood. In 2010-2011, we conducted an open-label, prospective efficacy study of artesunate as monotherapy in children 1-10 years of age with uncomplicated falciparum malaria in Bougoula-Hameau, Mali. Standard oral doses of artesunate were administered for 7 days and patients were followed up for 28 days. The data were compared to a similar study conducted in 2002-2004. Of 100 children enrolled in the 2010-2011 study, 92 were analyzed and compared to 217 children enrolled in the 2002-2004 study. The proportion of gametocyte carriers was unchanged at the end of treatment (23% at baseline vs. 24% on day 7, p = 1.0) and did not significantly decline until day 21 of follow-up (23% vs. 6%, p = 0.003). The mean gametocyte density at inclusion remained unchanged at the end of treatment (12 gametocytes/μL vs. 16 gametocytes/μL, p = 0.6). Overall, 46% of the 71 initial non-carriers had gametocytes detected by day 7. Similar results were found in the 2002-2004 study. In both studies, although gametocyte carriage significantly decreased by the end of the 28-day follow-up, artesunate did not clear mature gametocytes during treatment and did not prevent the appearance of new stage V gametocytes as assessed by light microscopy. Baseline gametocyte carriage was significantly higher 6 years after the deployment of artemisinin-based combination therapies in this setting. PMID:26839003

  12. The RNA-binding protein Puf1 functions in the maintenance of gametocytes in Plasmodium falciparum.

    PubMed

    Shrestha, Sony; Li, Xiaolian; Ning, Gang; Miao, Jun; Cui, Liwang

    2016-08-15

    Translation control plays an important role in the regulation of gene expression in the malaria parasite Plasmodium falciparum, especially in transition stages between the vertebrate host and mosquito vector. Here, we determined the function of the Puf-family member Puf1 (denoted as PfPuf1 for the P. falciparum protein) during P. falciparum sexual development. We show that PfPuf1 was expressed in all gametocyte stages and at higher levels in female gametocytes. PfPuf1 disruption did not interfere with the asexual erythrocyte cycle of the parasite but resulted in an approximately tenfold decrease of mature gametocytes. In the PfPuf1-disrupted lines, gametocytes appeared normal before stage III but subsequently exhibited a sharp decline in gametocytemia. This was accompanied by a concomitant accumulation of dead and dying late-stage gametocytes, which retained normal gross morphology. In addition, significantly more female gametocytes were lost in the PfPuf1-disrupted lines during development, resulting in a reversed male-to-female sex ratio. These results indicate that PfPuf1 is important for the differentiation and maintenance of gametocytes, especially female gametocytes. PMID:27383769

  13. Antimalarial Iron Chelator FBS0701 Blocks Transmission by Plasmodium falciparum Gametocyte Activation Inhibition

    PubMed Central

    Ferrer, Patricia; Vega-Rodriguez, Joel; Tripathi, Abhai K.; Jacobs-Lorena, Marcelo

    2014-01-01

    Reducing the transmission of the malarial parasite by Anopheles mosquitoes using drugs or vaccines remains a main focus in the efforts to control malaria. Iron chelators have been studied as potential antimalarial drugs due to their activities against different stages of the parasite. The iron chelator FBS0701 affects the development of Plasmodium falciparum early gametocytes and lowers blood-stage parasitemia. Here, we tested the effect of FBS0701 on stage V gametocyte infectivity for mosquitoes. The incubation of stage V gametocytes for up to 3 days with increasing concentrations of FBS0701 resulted in a significant dose-related reduction in mosquito infectivity, as measured by the numbers of oocysts per mosquito. The reduction in mosquito infectivity was due to the inhibition of male and female gametocyte activation. The preincubation of FBS0701 with ferric chloride restored gametocyte infectivity, showing that the inhibitory effect of FBS0701 was quenched by iron. Deferoxamine, another iron chelator, also reduced gametocyte infectivity but to a lesser extent. Finally, the simultaneous administration of drug and gametocytes to mosquitoes without previous incubation did not significantly reduce the numbers of oocysts. These results show the importance of gametocyte iron metabolism as a potential target for new transmission-blocking strategies. PMID:25512427

  14. Molecular evidence for the localization of Plasmodium falciparum immature gametocytes in bone marrow

    PubMed Central

    Aguilar, Ruth; Magallon-Tejada, Ariel; Achtman, Ariel H.; Moraleda, Cinta; Joice, Regina; Cisteró, Pau; Li Wai Suen, Connie S. N.; Nhabomba, Augusto; Macete, Eusebio; Mueller, Ivo; Marti, Matthias; Alonso, Pedro L.; Menéndez, Clara; Schofield, Louis

    2014-01-01

    Plasmodium falciparum immature gametocytes are not observed in peripheral blood. However, gametocyte stages in organs such as bone marrow have never been assessed by molecular techniques, which are more sensitive than optical microscopy. We quantified P falciparum sexual stages in bone marrow (n = 174) and peripheral blood (n = 70) of Mozambican anemic children by quantitative polymerase chain reaction targeting transcripts specific for early (PF14_0748; PHISTa), intermediate (PF13_0247; Pfs48/45), and mature (PF10_0303; Pfs25) gametocytes. Among children positive for the P falciparum housekeeping gene (PF08_0085; ubiquitin-conjugating enzyme gene) in bone marrow (n = 136) and peripheral blood (n = 25), prevalence of immature gametocytes was higher in bone marrow than peripheral blood (early: 95% vs 20%, P < .001; intermediate: 80% vs 16%; P < .001), as were transcript levels (P < .001 for both stages). In contrast, mature gametocytes were more prevalent (100% vs 51%, P < .001) and abundant (P < .001) in peripheral blood than in the bone marrow. Severe anemia (3.57, 95% confidence interval 1.49-8.53) and dyserythropoiesis (6.21, 95% confidence interval 2.24-17.25) were independently associated with a higher prevalence of mature gametocytes in bone marrow. Our results highlight the high prevalence and abundance of early sexual stages in bone marrow, as well as the relationship between hematological disturbances and gametocyte development in this tissue. PMID:24335496

  15. Splenic Retention of Plasmodium falciparum Gametocytes To Block the Transmission of Malaria

    PubMed Central

    Duez, Julien; Holleran, John P.; Ndour, Papa Alioune; Loganathan, Sasdekumar; Amireault, Pascal; Français, Olivier; El Nemer, Wassim; Le Pioufle, Bruno; Amado, Inês F.; Garcia, Sylvie; Chartrel, Nathalie; Le Van Kim, Caroline; Lavazec, Catherine; Avery, Vicky M.

    2015-01-01

    Plasmodium falciparum is transmitted from humans to Anopheles mosquito vectors via the sexual erythrocytic forms termed gametocytes. Erythrocyte filtration through microsphere layers (microsphiltration) had shown that circulating gametocytes are deformable. Compounds reducing gametocyte deformability would induce their splenic clearance, thus removing them from the blood circulation and blocking malaria transmission. The hand-made, single-sample prototype for microsphiltration was miniaturized to a 96-well microtiter plate format, and gametocyte retention in the microsphere filters was quantified by high-content imaging. The stiffening activity of 40 pharmacological compounds was assessed in microtiter plates, using a small molecule (calyculin) as a positive control. The stiffening activity of calyculin was assessed in spleen-mimetic microfluidic chips and in macrophage-depleted mice. Marked mechanical retention (80% to 90%) of mature gametocytes was obtained in microplates following exposure to calyculin at concentrations with no effect on parasite viability. Of the 40 compounds tested, including 20 antimalarials, only 5 endoperoxides significantly increased gametocyte retention (1.5- to 2.5-fold; 24 h of exposure at 1 μM). Mature gametocytes exposed to calyculin accumulated in microfluidic chips and were cleared from the circulation of macrophage-depleted mice as rapidly as heat-stiffened erythrocytes, thus confirming results obtained using the microsphiltration assay. An automated miniaturized approach to select compounds for their gametocyte-stiffening effect has been established. Stiffening induces gametocyte clearance both in vitro and in vivo. Based on physiologically validated tools, this screening cascade can identify novel compounds and uncover new targets to block malaria transmission. Innovative applications in hematology are also envisioned. PMID:25941228

  16. Effect of Fluorescent Dyes on In Vitro-Differentiated, Late-Stage Plasmodium falciparum Gametocytes

    PubMed Central

    Gebru, Tamirat; Mordmüller, Benjamin

    2014-01-01

    Plasmodium falciparum gametocytes are not associated with clinical symptoms, but they are responsible for transmitting the pathogen to mosquitoes. Therefore, gametocytocidal interventions are important for malaria control and resistance containment. Currently available drugs and vaccines are not well suited for that purpose. Several dyes have potent antimicrobial activity, but their use against gametocytes has not been investigated systematically. The gametocytocidal activity of nine synthetic dyes and four control compounds was tested against stage V gametocytes of the laboratory strain 3D7 and three clinical isolates of P. falciparum with a bioluminescence assay. Five of the fluorescent dyes had submicromolar 50% inhibitory concentration (IC50) values against mature gametocytes. Three mitochondrial dyes, MitoRed, dihexyloxacarbocyanine iodide (DiOC6), and rhodamine B, were highly active (IC50s < 200 nM). MitoRed showed the highest activity against gametocytes, with IC50s of 70 nM against 3D7 and 120 to 210 nM against clinical isolates. All compounds were more active against the laboratory strain 3D7 than against clinical isolates. In particular, the endoperoxides artesunate and dihydroartemisinin showed a 10-fold higher activity against 3D7 than against clinical isolates. In contrast to all clinically used antimalarials, several fluorescent dyes had surprisingly high in vitro activity against late-stage gametocytes. Since they also act against asexual blood stages, they shall be considered starting points for the development of new antimalarial lead compounds. PMID:25267675

  17. Antibody responses to surface antigens of Plasmodium falciparum gametocyte-infected erythrocytes and their relation to gametocytaemia.

    PubMed

    Dinko, B; King, E; Targett, G A T; Sutherland, C J

    2016-06-01

    An essential element for continuing transmission of Plasmodium falciparum is the availability of mature gametocytes in human peripheral circulation for uptake by mosquitoes. Natural immune responses to circulating gametocytes may play a role in reducing transmission from humans to mosquitoes. Here, antibody recognition of the surface of mature intra-erythrocytic gametocytes produced either by a laboratory-adapted parasite, 3D7, or by a recent clinical isolate of Kenyan origin (HL1204), was evaluated longitudinally in a cohort of Ghanaian school children by flow cytometry. This showed that a proportion of children exhibited antibody responses that recognized gametocyte surface antigens on one or both parasite lines. A subset of the children maintained detectable anti-gametocyte surface antigen (GSA) antibody levels during the 5 week study period. There was indicative evidence that children with anti-GSA antibodies present at enrolment were less likely to have patent gametocytaemia at subsequent visits (odds ratio = 0·29, 95% CI 0·06-1·05; P = 0·034). Our data support the existence of antigens on the surface of gametocyte-infected erythrocytes, but further studies are needed to confirm whether antibodies against them reduce gametocyte carriage. The identification of GSA would allow their evaluation as potential anti-gametocyte vaccine candidates and/or biomarkers for gametocyte carriage. PMID:27084060

  18. Routine in vitro culture of P. falciparum gametocytes to evaluate novel transmission-blocking interventions.

    PubMed

    Delves, Michael J; Straschil, Ursula; Ruecker, Andrea; Miguel-Blanco, Celia; Marques, Sara; Baum, Jake; Sinden, Robert E

    2016-09-01

    The prevention of parasite transmission from the human host to the mosquito has been recognized as a vital tool for malaria eradication campaigns. However, transmission-blocking antimalarial drug and/or vaccine discovery and development is currently hampered by the expense and difficulty of producing mature Plasmodium falciparum gametocytes in vitro-the parasite stage responsible for mosquito infection. Current protocols for P. falciparum gametocyte culture usually require complex parasite synchronization and addition of stimulating and/or inhibitory factors, and they may not have demonstrated the essential property of mosquito infectivity. This protocol details all the steps required for reliable P. falciparum gametocyte production and highlights common factors that influence culture success. The protocol can be completed in 15 d, and particular emphasis is placed upon operating a gametocyte culture facility on a continuous cycle. In addition, we show how functionally viable gametocytes can be used to evaluate transmission-blocking drugs both in a field setting and at high throughput (HTP) for drug discovery. PMID:27560172

  19. Plasmodium falciparum Mating Patterns and Mosquito Infectivity of Natural Isolates of Gametocytes

    PubMed Central

    Morlais, Isabelle; Nsango, Sandrine E.; Toussile, Wilson; Abate, Luc; Annan, Zeinab; Tchioffo, Majoline T.; Cohuet, Anna; Awono-Ambene, Parfait H.; Fontenille, Didier; Rousset, François; Berry, Antoine

    2015-01-01

    Plasmodium falciparum infections in malaria endemic areas often harbor multiple clones of parasites. However, the transmission success of the different genotypes within the mosquito vector has remained elusive so far. The genetic diversity of malaria parasites was measured by using microsatellite markers in gametocyte isolates from 125 asymptomatic carriers. For a subset of 49 carriers, the dynamics of co-infecting genotypes was followed until their development within salivary glands. Also, individual oocysts from midguts infected with blood from 9 donors were genotyped to assess mating patterns. Multiplicity of infection (MOI) was high both in gametocyte isolates and sporozoite populations, reaching up to 10 genotypes. Gametocyte isolates with multiple genotypes gave rise to lower infection prevalence and intensity. Fluctuations of genotype number occurred during the development within the mosquito and sub-patent genotypes, not detected in gametocyte isolates, were identified in the vector salivary glands. The inbreeding coefficient Fis was positively correlated to the oocyst loads, suggesting that P. falciparum parasites use different reproductive strategies according to the genotypes present in the gametocyte isolate. The number of parasite clones within an infection affects the transmission success and the mosquito has an important role in maintaining P. falciparum genetic diversity. Our results emphasize the crucial importance of discriminating between the different genotypes within an infection when studying the A. gambiae natural resistance to P. falciparum, and the need to monitor parasite diversity in areas where malaria control interventions are implemented. PMID:25875840

  20. Plasmodium falciparum gametocyte adhesion to C32 cells via CD36 is inhibited by antibodies to modified band 3.

    PubMed Central

    Rogers, N J; Targett, G A; Hall, B S

    1996-01-01

    Plasmodium falciparum gametocyte-infected erythrocytes are characterized by their ability to sequester in the microvasculature of various organs, primarily the spleen and bone marrow. This phenomenon is thought to play a critical role in the development and survival of the sexual stages. Little is known, however, about ligands on the gametocyte-infected erythrocyte. Infection of erythrocytes with mature asexual stages of P. falciparum (trophozoites and schizonts) has been shown to induce modification of the erythrocyte anion transporter, band 3, and this has been linked to the acquisition of an adherent phenotype. Here, we demonstrate for the first time that immature gametocyte-infected erythrocytes also express modified band 3. In vitro binding assays demonstrate that gametocyte-infected erythrocytes of the 3D7 strain utilize this surface receptor for adhesion to C32 amelanotic melanoma cells via the host cell receptor CD36 (platelet glycoprotein IIIb). Adhesion of gametocyte-infected erythrocytes to CD36-transfected CHO cells is also dependent on modified band 3. However, modified band 3 does not mediate adhesion of gametocyte-infected erythrocytes to intercellular adhesion molecule 1, a second host receptor for gametocytes expressed on C32 cells. PMID:8926098

  1. Plasmodium falciparum Gametocyte Development 1 (Pfgdv1) and Gametocytogenesis Early Gene Identification and Commitment to Sexual Development

    PubMed Central

    Eksi, Saliha; Morahan, Belinda J.; Haile, Yoseph; Furuya, Tetsuya; Jiang, Hongying; Ali, Omar; Xu, Huichun; Kiattibutr, Kirakorn; Suri, Amreena; Czesny, Beata; Adeyemo, Adebowale; Myers, Timothy G.; Sattabongkot, Jetsumon; Su, Xin-zhuan; Williamson, Kim C.

    2012-01-01

    Malaria transmission requires the production of male and female gametocytes in the human host followed by fertilization and sporogonic development in the mosquito midgut. Although essential for the spread of malaria through the population, little is known about the initiation of gametocytogenesis in vitro or in vivo. Using a gametocyte-defective parasite line and genetic complementation, we show that Plasmodium falciparum gametocyte development 1 gene (Pfgdv1), encoding a peri-nuclear protein, is critical for early sexual differentiation. Transcriptional analysis of Pfgdv1 negative and positive parasite lines identified a set of gametocytogenesis early genes (Pfge) that were significantly down-regulated (>10 fold) in the absence of Pfgdv1 and expression was restored after Pfgdv1 complementation. Progressive accumulation of Pfge transcripts during successive rounds of asexual replication in synchronized cultures suggests that gametocytes are induced continuously during asexual growth. Comparison of Pfge gene transcriptional profiles in patient samples divided the genes into two groups differing in their expression in mature circulating gametocytes and providing candidates to evaluate gametocyte induction and maturation separately in vivo. The expression profile of one of the early gametocyte specific genes, Pfge1, correlated significantly with asexual parasitemia, which is consistent with the ongoing induction of gametocytogenesis during asexual growth observed in vitro and reinforces the need for sustained transmission-blocking strategies to eliminate malaria. PMID:23093935

  2. Activity of Herbal Medicines on Plasmodium falciparum Gametocytes: Implications for Malaria Transmission in Ghana

    PubMed Central

    Amoah, Linda Eva; Kakaney, Courage; Kwansa-Bentum, Bethel; Kusi, Kwadwo Asamoah

    2015-01-01

    Background Malaria still remains a major health issue in Ghana despite the introduction of Artemisinin-based combination therapy (ACT) coupled with other preventative measures such as the use of insecticide treated nets (ITNs). The global quest for eradication of malaria has heightened the interest of identifying drugs that target the sexual stage of the parasite, referred to as transmission-blocking drugs. This study aimed at assessing the efficacy and gametocydal effects of some commonly used herbal malaria products in Ghana. Methodology/Principal Findings After identifying herbal anti-malarial products frequently purchased on the Ghanaian market, ten of them were selected and lyophilized. In vitro drug sensitivity testing of different concentrations of the herbal products was carried out on asexual and in vitro generated gametocytes of the 3D7 strain of Plasmodium falciparum. The efficacies of the products were assessed by microscopy. Cultures containing low dose of RT also produced the least number of late stage gametocytes. Two of the herbal products CM and RT inhibited the growth of late stage gametocytes by > 80% at 100 μg/ml whilst KG was the most inhibitory to early stage gametocytes at that same concentration. However at 1 μg/ml, only YF significantly inhibited the survival of late stage gametocytes although at that same concentration YF barely inhibited the survival of early stage gametocytes. Conclusions/Significance Herbal product RT (Aloe schweinfurthii, Khaya senegalensis, Piliostigma thonningii and Cassia siamea) demonstrated properties of a highly efficacious gametocydal product. Low dose of herbal product RT exhibited the highest gametocydal activity and at 100 μg/ml, RT exhibited >80% inhibition of late stage gametocytes. However inhibition of asexual stage parasite by RT was not optimal. Improving the asexual inhibition of RT could convert RT into an ideal antimalarial herbal product. We also found that generally C. sanguinolenta containing

  3. Gametocyte Clearance Kinetics Determined by Quantitative Magnetic Fractionation in Melanesian Children with Uncomplicated Malaria Treated with Artemisinin Combination Therapy

    PubMed Central

    Karl, Stephan; Laman, Moses; Moore, Brioni R.; Benjamin, John; Koleala, Tamarah; Ibam, Clemencia; Kasian, Bernadine; Siba, Peter M.; Waltmann, Andreea; Mueller, Ivo; Woodward, Robert C.; St. Pierre, Timothy G.

    2015-01-01

    Quantitative magnetic fractionation and a published mathematical model were used to characterize between-treatment differences in gametocyte density and prevalence in 70 Papua New Guinean children with uncomplicated Plasmodium falciparum and/or Plasmodium vivax malaria randomized to one of two artemisinin combination therapies (artemether-lumefantrine or artemisinin-naphthoquine) in an intervention trial. There was an initial rise in peripheral P. falciparum gametocyte density with both treatments, but it was more pronounced in the artemisinin-naphthoquine group. Model-derived estimates of the median pretreatment sequestered gametocyte population were 21/μl for artemether-lumefantrine and 61/μl for artemisinin-naphthoquine (P < 0.001). The median time for P. falciparum gametocyte density to fall to <2.5/μl (below which transmission becomes unlikely) was 16 days in the artemether-lumefantrine group and 20 days in artemisinin-naphthoquine group (P < 0.001). Gametocyte prevalence modeling suggested that artemisinin-naphthoquine-treated children became gametocytemic faster (median, 2.2 days) than artemether-lumefantrine-treated children (median, 5.3 days; P < 0.001) and had a longer median P. falciparum gametocyte carriage time per individual (20 versus 13 days; P < 0.001). Clearance of P. vivax gametocytes was rapid (within 3 days) in both groups; however, consistent with the reappearance of asexual forms in the main trial, nearly 40% of children in the artemether-lumefantrine group developed P. vivax gametocytemia between days 28 and 42 compared with 3% of children in the artemisinin-naphthoquine group. These data suggest that artemisinin is less active than artemether against sequestered gametocytes. Greater initial gametocyte release after artemisinin-naphthoquine increases the period of potential P. falciparum transmission by 4 days relative to artemether-lumefantrine, but the longer elimination half-life of naphthoquine than of lumefantrine suppresses P. vivax

  4. Sub-microscopic gametocyte carriage in febrile children living in different areas of Gabon

    PubMed Central

    2013-01-01

    Background Considering malaria prevalence declines in parts of sub-Saharan Africa, such as Gabon, identification of the human infectious reservoir is important for successful malaria control. Microscopic and sub-microscopic parasites contribute to malaria transmission. The aim of the present study was to evaluate the proportion of microscopic and sub-microscopic gametocyte carriers among febrile patients in two different areas of Gabon. Methods Samples from febrile children aged less than 11 years old were collected from February 2008 to January 2009 at two health centres of Gabon. Patients were screened for the presence of asexual Plasmodium falciparum parasites. Gametocyte carriage was determined by microscopy and QT-NASBA. Results Gametocytes were detected in 5.3% (n = 16/304) of children by microscopy compared to 45.7% (n = 139/304) by QT-Nasba. Sub-microscopic gametocyte carriage (ie microscopy negative and QT-Nasba positive) was found in 89.2% (n = 124/139) of patients. Among patients with microscopically detected trophozoites, the proportion of sub-microscopic gametocyte (SMG) carriers was 58.4% (n = 118/202) and 6% in samples from children with negative slides (p < 0.01). In Oyem, where malaria prevalence is three-fold higher than in Owendo, SMG carriage was more frequent (49.0% vs 32.6% in Owendo; p < 0.01). Conclusion Sub-microscopic gametocytaemia is common among Gabonese febrile children. They might strongly contribute to maintain malaria transmission. However, further analysis of sub-microscopic parasite carriage among asymptomatic individuals will be helpful to better characterize malaria transmission. PMID:24168323

  5. Purification Methodology for Viable and Infective Plasmodium vivax Gametocytes That Is Compatible with Transmission-Blocking Assays

    PubMed Central

    Vera, Omaira; Brelas de Brito, Paula; Albrecht, Letusa; Martins-Campos, Keillen Monick; Pimenta, Paulo F. P.; Monteiro, Wuelton M.; Lacerda, Marcus V. G.

    2015-01-01

    Significant progress toward the control of malaria has been achieved, especially regarding Plasmodium falciparum infections. However, the unique biology of Plasmodium vivax hampers current control strategies. The early appearance of P. vivax gametocytes in the peripheral blood and the impossibility of culturing this parasite are major drawbacks. Using blood samples from 40 P. vivax-infected patients, we describe here a methodology to purify viable gametocytes and further infect anophelines. This method opens new avenues to validate transmission-blocking strategies. PMID:26239989

  6. Purification Methodology for Viable and Infective Plasmodium vivax Gametocytes That Is Compatible with Transmission-Blocking Assays.

    PubMed

    Vera, Omaira; Brelas de Brito, Paula; Albrecht, Letusa; Martins-Campos, Keillen Monick; Pimenta, Paulo F P; Monteiro, Wuelton M; Lacerda, Marcus V G; Lopes, Stefanie C P; Costa, Fabio T M

    2015-10-01

    Significant progress toward the control of malaria has been achieved, especially regarding Plasmodium falciparum infections. However, the unique biology of Plasmodium vivax hampers current control strategies. The early appearance of P. vivax gametocytes in the peripheral blood and the impossibility of culturing this parasite are major drawbacks. Using blood samples from 40 P. vivax-infected patients, we describe here a methodology to purify viable gametocytes and further infect anophelines. This method opens new avenues to validate transmission-blocking strategies. PMID:26239989

  7. Inclusion of gametocyte parameters in anti-malarial drug efficacy studies: filling a neglected gap needed for malaria elimination.

    PubMed

    Abdul-Ghani, Rashad; Basco, Leonardo K; Beier, John C; Mahdy, Mohammed A K

    2015-01-01

    Standard anti-malarial drug efficacy and drug resistance assessments neglect the gametocyte parameters in their protocols. With the spread of drug resistance and the absence of clinically proven vaccines, the use of gametocytocidal drugs or drug combinations with transmission-blocking activity is a high priority for malaria control and elimination. However, the limited repertoire of gametocytocidal drugs and induction of gametocytogenesis after treatment with certain anti-malarial drugs necessitate both regular monitoring of gametocytocidal activities of anti-malarial drugs in clinical use and the effectiveness of candidate gametocytocidal agents. Therefore, updating current protocols of anti-malarial drug efficacy is needed to reflect the effects of anti-malarial drugs or drug combinations on gametocyte carriage and gametocyte density along with asexual parasite density. Developing protocols of anti-malarial drug efficacy that include gametocyte parameters related to both microscopic and submicroscopic gametocytaemias is important if drugs or drug combinations are to be strategically used in transmission-blocking interventions in the context of malaria elimination. The present piece of opinion highlights the challenges in gametocyte detection and follow-up and discuss the need for including the gametocyte parameter in anti-malarial efficacy studies. PMID:26481312

  8. Monitoring the Prevalence of Leucocytozoon sabrazesi in Southern China and Testing Tricyclic Compounds against Gametocytes.

    PubMed

    Zhao, Wenting; Pang, Qin; Xu, Ruixue; Liu, Jianwen; Liu, Shengfa; Li, Jian; Su, Xin-Zhuan

    2016-01-01

    Leucocytozoon parasites infect many species of avian hosts, including domestic chicken, and can inflict heavy economic loss on the poultry industry. Two major species of Leucocytozoon parasites have been reported in China, L. sabrazesi and L. caulleryi, although L. sabrazesi appears to be more widespread than L. caulleryi in southern China. The traditional method for detecting Leucocytozoon infection is microscopic examination of blood smears for the presence of mature gametocytes in circulation, which may miss infections with low parasitemia (gametocytemia) or immature gametocytes. Here we developed a PCR-based method to monitor L. sabrazesi infections at seven sites in four provinces of China after testing two PCR primer pairs based on parasite mitochondrial cytochrome b (cytb) and cytochrome c oxidase III (coxIII) genes. We compared the results of PCR detection with those of microscopic observation. As expected, the PCR assays were more sensitive than microscope examination in detecting L. sabrazesi infection and were able to detect parasite DNA after gametocytes disappeared in the blood stream. Using these methods, we investigated monthly dynamics of L. sabrazesi in chickens from a free-range farm in Xiamen, Fujian province of China, over one year. Our results showed that chickens were infected with L. sabrazesi year-round in southern China. Finally, we tested several compounds for potential treatment of Leucocytozoon infections, including primaquine, ketotifen, clomipramine hydrochloride, desipramine hydrochloride, sulfaquinoxaline, and pyrimethamine. Only primaquine had activity against L. sabrazesi gametocytes. Our results provide important information for controlling parasite transmission in southern China and disease management. PMID:27571513

  9. Monitoring the Prevalence of Leucocytozoon sabrazesi in Southern China and Testing Tricyclic Compounds against Gametocytes

    PubMed Central

    Zhao, Wenting; Pang, Qin; Xu, Ruixue; Liu, Jianwen; Liu, Shengfa; Li, Jian; Su, Xin-zhuan

    2016-01-01

    Leucocytozoon parasites infect many species of avian hosts, including domestic chicken, and can inflict heavy economic loss on the poultry industry. Two major species of Leucocytozoon parasites have been reported in China, L. sabrazesi and L. caulleryi, although L. sabrazesi appears to be more widespread than L. caulleryi in southern China. The traditional method for detecting Leucocytozoon infection is microscopic examination of blood smears for the presence of mature gametocytes in circulation, which may miss infections with low parasitemia (gametocytemia) or immature gametocytes. Here we developed a PCR-based method to monitor L. sabrazesi infections at seven sites in four provinces of China after testing two PCR primer pairs based on parasite mitochondrial cytochrome b (cytb) and cytochrome c oxidase III (coxIII) genes. We compared the results of PCR detection with those of microscopic observation. As expected, the PCR assays were more sensitive than microscope examination in detecting L. sabrazesi infection and were able to detect parasite DNA after gametocytes disappeared in the blood stream. Using these methods, we investigated monthly dynamics of L. sabrazesi in chickens from a free-range farm in Xiamen, Fujian province of China, over one year. Our results showed that chickens were infected with L. sabrazesi year-round in southern China. Finally, we tested several compounds for potential treatment of Leucocytozoon infections, including primaquine, ketotifen, clomipramine hydrochloride, desipramine hydrochloride, sulfaquinoxaline, and pyrimethamine. Only primaquine had activity against L. sabrazesi gametocytes. Our results provide important information for controlling parasite transmission in southern China and disease management. PMID:27571513

  10. The Gametocytes of Leucocytozoon sabrazesi Infect Chicken Thrombocytes, Not Other Blood Cells

    PubMed Central

    Zhao, Wenting; Liu, Jianwen; Xu, Ruixue; Zhang, Cui; Pang, Qin; Chen, Xin; Liu, Shengfa; Hong, Lingxian; Yuan, Jing; Li, Xiaotong; Chen, Yixin; Li, Jian; Su, Xin-zhuan

    2015-01-01

    Leucocytozoon parasites infect a large number of avian hosts, including domestic chicken, and cause significant economical loss to the poultry industry. Although the transmission stages of the parasites were observed in avian blood cells more than a century ago, the specific host cell type(s) that the gametocytes infect remain uncertain. Because all the avian blood cells, including red blood cells (RBCs), are nucleated, and the developing parasites dramatically change the morphology of the infected host cells, it has been difficult to identify Leucocytozoon infected host cell(s). Here we use cell-type specific antibodies to investigate the identities of the host cells infected by Leucocytozoon sabrazesi gametocytes. Anti-RBC antibodies stained RBCs membrane strongly, but not the parasite-infected cells, ruling out the possibility of RBCs being the infected host cells. Antibodies recognizing various leukocytes including heterophils, monocytes, lymphocytes, and macrophages did not stain the infected cells either. Antisera raised against a peptide of the parasite cytochrome B (CYTB) stained parasite-infected cells and some leukocytes, particularly cells with a single round nucleus as well as clear/pale cytoplasm suggestive of thrombocytes. Finally, a monoclonal antibody known to specifically bind chicken thrombocytes also stained the infected cells, confirming that L. sabrazesi gametocytes develop within chicken thrombocytes. The identification of L. sabrazesi infected host cell solves a long unresolved puzzle and provides important information for studying parasite invasion of host cells and for developing reagents to interrupt parasite transmission. PMID:26218846

  11. A review of the effects of artemether-lumefantrine on gametocyte carriage and disease transmission

    PubMed Central

    2014-01-01

    While significant advances have been made in the prevention and treatment of malaria in recent years, these successes continue to fall short of the World Health Organization (WHO) goals for malaria control and elimination. For elimination strategies to be effective, limited disease transmission, achieved through rapid reduction in the infectious parasite reservoir and decreased gametocyte carriage, will be critical. Artemisinin-based combination therapy (ACT) forms the cornerstone of WHO-recommended treatment for uncomplicated Plasmodium falciparum malaria, and in combination with other effective interventions will undoubtedly play a vital role in elimination programmes. The gametocytocidal properties of artemisinins are a bonus attribute; there is epidemiological evidence of reductions in malaria incidence and transmission in African regions since the introduction of these agents. Many studies and analyses have specifically investigated the effects of the ACT, artemether-lumefantrine (AL) on gametocyte carriage. In this systematic review of 62 articles published between 1998 and January 2014, the effects of AL on gametocyte carriage and malaria transmission are compared with other artemisinin-based anti-malarials and non-ACT. The impact of AL treatment of asymptomatic carriers on population gametocyte carriage, and the potential future role of AL in malaria elimination initiatives are also considered. Despite the inherent difficulties in comparing data from a range of different studies that also utilized different diagnostic approaches to assess baseline gametocyte counts, the gametocytocidal effect of AL was proportionately consistent across the studies reviewed, suggesting that AL will continue to play a vital role in the treatment of malaria and contribute to clearing the path towards malaria elimination. However, the specific place of AL is the subject of much ongoing research and will undoubtedly be dependent on different demographic and geographical

  12. Epidemiology and Infectivity of Plasmodium falciparum and Plasmodium vivax Gametocytes in Relation to Malaria Control and Elimination

    PubMed Central

    Bousema, Teun; Drakeley, Chris

    2011-01-01

    Summary: Malaria remains a major cause of morbidity and mortality in the tropics, with Plasmodium falciparum responsible for the majority of the disease burden and P. vivax being the geographically most widely distributed cause of malaria. Gametocytes are the sexual-stage parasites that infect Anopheles mosquitoes and mediate the onward transmission of the disease. Gametocytes are poorly studied despite this crucial role, but with a recent resurgence of interest in malaria elimination, the study of gametocytes is in vogue. This review highlights the current state of knowledge with regard to the development and longevity of P. falciparum and P. vivax gametocytes in the human host and the factors influencing their distribution within endemic populations. The evidence for immune responses, antimalarial drugs, and drug resistance influencing infectiousness to mosquitoes is reviewed. We discuss how the application of molecular techniques has led to the identification of submicroscopic gametocyte carriage and to a reassessment of the human infectious reservoir. These components are drawn together to show how control measures that aim to reduce malaria transmission, such as mass drug administration and a transmission-blocking vaccine, might better be deployed. PMID:21482730

  13. Integrated transcriptomic and proteomic analyses of P. falciparum gametocytes: molecular insight into sex-specific processes and translational repression.

    PubMed

    Lasonder, Edwin; Rijpma, Sanna R; van Schaijk, Ben C L; Hoeijmakers, Wieteke A M; Kensche, Philip R; Gresnigt, Mark S; Italiaander, Annet; Vos, Martijn W; Woestenenk, Rob; Bousema, Teun; Mair, Gunnar R; Khan, Shahid M; Janse, Chris J; Bártfai, Richárd; Sauerwein, Robert W

    2016-07-27

    Sexual differentiation of malaria parasites into gametocytes in the vertebrate host and subsequent gamete fertilization in mosquitoes is essential for the spreading of the disease. The molecular processes orchestrating these transitions are far from fully understood. Here, we report the first transcriptome analysis of male and female Plasmodium falciparum gametocytes coupled with a comprehensive proteome analysis. In male gametocytes there is an enrichment of proteins involved in the formation of flagellated gametes; proteins involved in DNA replication, chromatin organization and axoneme formation. On the other hand, female gametocytes are enriched in proteins required for zygote formation and functions after fertilization; protein-, lipid- and energy-metabolism. Integration of transcriptome and proteome data revealed 512 highly expressed maternal transcripts without corresponding protein expression indicating large scale translational repression in P. falciparum female gametocytes for the first time. Despite a high degree of conservation between Plasmodium species, 260 of these 'repressed transcripts' have not been previously described. Moreover, for some of these genes, protein expression is only reported in oocysts and sporozoites indicating that repressed transcripts can be partitioned into short- and long-term storage. Finally, these data sets provide an essential resource for identification of vaccine/drug targets and for further mechanistic studies. PMID:27298255

  14. Manufacture and Testing of a High Field Gradient Magnetic Fractionation System for Quantitative Detection of Plasmodium falciparum Gametocytes

    NASA Astrophysics Data System (ADS)

    Karl, Stephan; Woodward, Robert C.; Davis, Timothy M. E.; St. Pierre, Tim G.

    2010-12-01

    Plasmodium falciparum is the most dangerous of the human malaria parasite species and accounts for millions of clinical episodes of malaria each year in tropical countries. The pathogenicity of Plasmodium falciparum is a result of its ability to infect erythrocytes where it multiplies asexually over 48 h or develops into sexual forms known as gametocytes. If sufficient male and female gametocytes are taken up by a mosquito vector, it becomes infectious. Therefore, the presence and density of gametocytes in human blood is an important indicator of human-to-mosquito transmission of malaria. Recently, we have shown that high field gradient magnetic fractionation improves gametocyte detection in human blood samples. Here we present two important new developments. Firstly we introduce a quantitative approach to replace the previous qualitative method and, secondly, we describe a novel method that enables cost-effective production of the magnetic fractionation equipment required to carry out gametocyte quantification. We show that our custom-made magnetic fractionation equipment can deliver results with similar sensitivity and convenience but for a small fraction of the cost.

  15. Mosquito Feeding Assays to Determine the Infectiousness of Naturally Infected Plasmodium falciparum Gametocyte Carriers

    PubMed Central

    Bousema, Teun; Dinglasan, Rhoel R.; Morlais, Isabelle; Gouagna, Louis C.; van Warmerdam, Travis; Awono-Ambene, Parfait H.; Bonnet, Sarah; Diallo, Mouctar; Coulibaly, Mamadou; Tchuinkam, Timoléon; Mulder, Bert; Targett, Geoff; Drakeley, Chris; Sutherland, Colin; Robert, Vincent; Doumbo, Ogobara; Touré, Yeya; Graves, Patricia M.; Roeffen, Will; Sauerwein, Robert; Birkett, Ashley; Locke, Emily; Morin, Merribeth; Wu, Yimin; Churcher, Thomas S.

    2012-01-01

    Introduction In the era of malaria elimination and eradication, drug-based and vaccine-based approaches to reduce malaria transmission are receiving greater attention. Such interventions require assays that reliably measure the transmission of Plasmodium from humans to Anopheles mosquitoes. Methods We compared two commonly used mosquito feeding assay procedures: direct skin feeding assays and membrane feeding assays. Three conditions under which membrane feeding assays are performed were examined: assays with i) whole blood, ii) blood pellets resuspended with autologous plasma of the gametocyte carrier, and iii) blood pellets resuspended with heterologous control serum. Results 930 transmission experiments from Cameroon, The Gambia, Mali and Senegal were included in the analyses. Direct skin feeding assays resulted in higher mosquito infection rates compared to membrane feeding assays (odds ratio 2.39, 95% confidence interval 1.94–2.95) with evident heterogeneity between studies. Mosquito infection rates in membrane feeding assays and direct skin feeding assays were strongly correlated (p<0.0001). Replacing the plasma of the gametocyte donor with malaria naïve control serum resulted in higher mosquito infection rates compared to own plasma (OR 1.92, 95% CI 1.68–2.19) while the infectiousness of gametocytes may be reduced during the replacement procedure (OR 0.60, 95% CI 0.52–0.70). Conclusions Despite a higher efficiency of direct skin feeding assays, membrane feeding assays appear suitable tools to compare the infectiousness between individuals and to evaluate transmission-reducing interventions. Several aspects of membrane feeding procedures currently lack standardization; this variability makes comparisons between laboratories challenging and should be addressed to facilitate future testing of transmission-reducing interventions. PMID:22936993

  16. Membrane Feeding Assay to Determine the Infectiousness of Plasmodium vivax Gametocytes.

    PubMed

    Sattabongkot, Jetsumon; Kumpitak, Chalermpon; Kiattibutr, Kirakorn

    2015-01-01

    The evaluation of Plasmodium vivax gametocyte infectiousness by the membrane feeding assay is herein described. While P. vivax cannot be cultured and different parasite isolates may infect mosquitoes at different rates, the protocol described in this chapter identifies critical parameters to be considered when performing the assay such as methods for the preparation of the mosquitoes, the size of the blood cup, and the blood volume used. In previous studies the data have shown that the membrane feeding assay is useful for studies of parasite biology, and the effects of transmission blocking drugs and vaccines. PMID:26450382

  17. Nonrandomized Controlled Trial of Artesunate plus Sulfadoxine-Pyrimethamine with or without Primaquine for Preventing Posttreatment Circulation of Plasmodium falciparum Gametocytes

    PubMed Central

    Shah, Naman K.; Schapira, Allan; Juliano, Jonathan J.; Srivastava, Bina; MacDonald, Pia D. M.; Poole, Charles; Anvikar, Anup; Meshnick, Steven R.; Valecha, Neena

    2013-01-01

    Artemisinin combination therapies eliminate immature Plasmodium falciparum gametocytes but not mature gametocytes, which may persist for up to 1 month posttreatment. A single dose of primaquine, which is inexpensive and effective against mature gametocytes, could be added to further reduce the potential for posttreatment parasite transmission. Currently, we have few data regarding the effectiveness or safety of doing so. We collected data from 21 therapeutic efficacy trials of the National Antimalarial Drug Resistance Monitoring System of India conducted during 2009 to 2010, wherein 9 sites used single-dose primaquine (0.75 mg/kg of body weight) administered on day 2 along with artesunate plus sulfadoxine-pyrimethamine (AS+SP) while 12 did not. We estimated the effect of primaquine on posttreatment gametocyte clearance and the total number of gametocyte-weeks as determined by microscopy. We compared the median area under the curve for gametocyte density and reported adverse events. One thousand three hundred thirty-five patients completed the antimalarial drug treatment. Adjusting for region, primaquine increased the rate of gametocyte clearance (hazard ratio, 1.9; 95% confidence interval [CI], 1.1 to 3.3), prevented 45% (95% CI, 19 to 62) of posttreatment gametocyte-weeks, and decreased the area under the gametocyte density curve over the 28-day follow-up compared to AS+SP alone (P value = 0.01). The results were robust to other adjustment sets, and the estimated effect of primaquine increased during sensitivity analysis on the measurement of exposure time. No serious adverse events were detected. In conclusion, the addition of primaquine to AS+SP was effective in reducing the posttreatment presence of P. falciparum gametocytes. Primaquine was well tolerated and could be administered along with an artemisinin combination therapy as the first-line therapy. PMID:23587943

  18. Insight into phagocytosis of mature sexual (gametocyte) stages of Plasmodium falciparum using a human monocyte cell line.

    PubMed

    Bansal, Geetha P; Weinstein, Corey S; Kumar, Nirbhay

    2016-05-01

    During natural infection malaria parasites are injected into the bloodstream of a human host by the bite of an infected female Anopheles mosquito. Both asexual and mature sexual stages of Plasmodium circulate in the blood. Asexual forms are responsible for clinical malaria while sexual stages are responsible for continued transmission via the mosquitoes. Immune responses generated against various life cycle stages of the parasite have important roles in resistance to malaria and in reducing malaria transmission. Phagocytosis of free merozoites and erythrocytic asexual stages has been well studied, but very little is known about similar phagocytic clearance of mature sexual stages, which are critical for transmission. We evaluated phagocytic uptake of mature sexual (gametocyte) stage parasites by a human monocyte cell line in the absence of immune sera. We found that intact mature stages do not undergo phagocytosis, unless they are either killed or freed from erythrocytes. In view of this observation, we propose that the inability of mature gametocytes to be phagocytized may actually result in malaria transmission advantage. On the other hand, mature gametocytes that are not transmitted to mosquitoes during infection will eventually die and undergo phagocytosis, initiating immune responses that may have transmission blocking potential. A better understanding of early phagocytic clearance and immune responses to gametocytes may identify additional targets for transmission blocking strategies. PMID:26851166

  19. Malaria proteases mediate inside-out egress of gametocytes from red blood cells following parasite transmission to the mosquito.

    PubMed

    Sologub, Ludmilla; Kuehn, Andrea; Kern, Selina; Przyborski, Jude; Schillig, Rebecca; Pradel, Gabriele

    2011-06-01

    Malaria parasites reside in human erythrocytes within a parasitophorous vacuole. The parasites are transmitted from the human to the mosquito by the uptake of intraerythrocytic gametocytes during a blood meal, which in the midgut become activated by external stimuli and subsequently egress from the enveloping erythrocyte. Gametocyte egress is a crucial step for the parasite to prepare for fertilization, but the molecular mechanisms of egress are not well understood. Via electron microscopy, we show that Plasmodium falciparum gametocytes exit the erythrocyte by an inside-out type of egress. The parasitophorous vacuole membrane (PVM) ruptures at multiple sites within less than a minute following activation, a process that requires a temperature drop and parasite contact with xanthurenic acid. PVM rupture can also be triggered by the ionophore nigericin and is sensitive to the cysteine protease inhibitor E-64d. Following PVM rupture the subpellicular membrane begins to disintegrate. This membrane is specific to malaria gametocytes, and disintegration is impaired by the aspartic protease inhibitor EPNP and the cysteine/serine protease inhibitor TLCK. Approximately 15 min post activation, the erythrocyte membrane ruptures at a single breaking point, which can be inhibited by inhibitors TLCK and TPCK. In all cases inhibitor treatment results in interrupted gametogenesis. PMID:21501358

  20. Luciferase-Based, High-Throughput Assay for Screening and Profiling Transmission-Blocking Compounds against Plasmodium falciparum Gametocytes.

    PubMed

    Lucantoni, Leonardo; Fidock, David A; Avery, Vicky M

    2016-04-01

    The discovery of new antimalarial drugs able to target both the asexual and gametocyte stages ofPlasmodium falciparumis critical to the success of the malaria eradication campaign. We have developed and validated a robust, rapid, and cost-effective high-throughput reporter gene assay to identify compounds active against late-stage (stage IV and V) gametocytes. The assay, which is suitable for testing compound activity at incubation times up to 72 h, demonstrates excellent quality and reproducibility, with averageZ' values of 0.85 ± 0.01. We used the assay to screen more than 10,000 compounds from three chemically diverse libraries. The screening outcomes highlighted the opportunity to use collections of compounds with known activity against the asexual stages of the parasites as a starting point for gametocytocidal activity detection in order to maximize the chances of identifying gametocytocidal compounds. This assay extends the capabilities of our previously reported luciferase assay, which tested compounds against early-stage gametocytes, and opens possibilities to profile the activities of gametocytocidal compounds over the entire course of gametocytogenesis. PMID:26787698

  1. In Vitro Activities of Primaquine-Schizonticide Combinations on Asexual Blood Stages and Gametocytes of Plasmodium falciparum

    PubMed Central

    Cabrera, Mynthia

    2015-01-01

    Currently, the World Health Organization recommends addition of a 0.25-mg base/kg single dose of primaquine (PQ) to artemisinin combination therapies (ACTs) for Plasmodium falciparum malaria as a gametocytocidal agent for reducing transmission. Here, we investigated the potential interactions of PQ with the long-lasting components of the ACT drugs for eliminating the asexual blood stages and gametocytes of in vitro-cultured P. falciparum strains. Using the SYBR green I assay for asexual parasites and a flow cytometry-based assay for gametocytes, we determined the interactions of PQ with the schizonticides chloroquine, mefloquine, piperaquine, lumefantrine, and naphthoquine. With the sums of fractional inhibitory concentrations and isobolograms, we were able to determine mostly synergistic interactions for the various PQ and schizonticide combinations on the blood stages of P. falciparum laboratory strains. The synergism in inhibiting asexual stages and gametocytes was highly evident with PQ-naphthoquine, whereas synergism was moderate for the PQ-piperaquine, PQ-chloroquine, and PQ-mefloquine combinations. We have detected potentially antagonistic interactions between PQ and lumefantrine under certain drug combination ratios, suggesting that precautions might be needed when PQ is added as the gametocytocide to the artemether-lumefantrine ACT (Coartem). PMID:26416869

  2. In Vitro Activities of Primaquine-Schizonticide Combinations on Asexual Blood Stages and Gametocytes of Plasmodium falciparum.

    PubMed

    Cabrera, Mynthia; Cui, Liwang

    2015-12-01

    Currently, the World Health Organization recommends addition of a 0.25-mg base/kg single dose of primaquine (PQ) to artemisinin combination therapies (ACTs) for Plasmodium falciparum malaria as a gametocytocidal agent for reducing transmission. Here, we investigated the potential interactions of PQ with the long-lasting components of the ACT drugs for eliminating the asexual blood stages and gametocytes of in vitro-cultured P. falciparum strains. Using the SYBR green I assay for asexual parasites and a flow cytometry-based assay for gametocytes, we determined the interactions of PQ with the schizonticides chloroquine, mefloquine, piperaquine, lumefantrine, and naphthoquine. With the sums of fractional inhibitory concentrations and isobolograms, we were able to determine mostly synergistic interactions for the various PQ and schizonticide combinations on the blood stages of P. falciparum laboratory strains. The synergism in inhibiting asexual stages and gametocytes was highly evident with PQ-naphthoquine, whereas synergism was moderate for the PQ-piperaquine, PQ-chloroquine, and PQ-mefloquine combinations. We have detected potentially antagonistic interactions between PQ and lumefantrine under certain drug combination ratios, suggesting that precautions might be needed when PQ is added as the gametocytocide to the artemether-lumefantrine ACT (Coartem). PMID:26416869

  3. Risk factors for Plasmodium falciparum and Plasmodium vivax gametocyte carriage in Papua New Guinean children with uncomplicated malaria.

    PubMed

    Karl, Stephan; Laman, Moses; Moore, Brioni R; Benjamin, John M; Salib, Mary; Lorry, Lina; Maripal, Samuel; Siba, Peter; Robinson, Leanne J; Mueller, Ivo; Davis, Timothy M E

    2016-08-01

    There are limited data on gametocytaemia risk factors before/after treatment with artemisinin combination therapy in children from areas with transmission of multiple Plasmodium species. We utilised data from a randomised trial comparing artemether-lumefantrine (AL) and artemisinin-naphthoquine (AN) in 230 Papua New Guinean children aged 0.5-5 years with uncomplicated malaria in whom determinants of gametocytaemia by light microscopy were assessed at baseline using logistic regression and during follow-up using multilevel mixed effects modelling. Seventy-four (32%) and 18 (8%) children presented with P. falciparum and P. vivax gametocytaemia, respectively. Baseline P. falciparum gametocytaemia was associated with Hackett spleen grade 1 (odds ratio (95% CI) 4.01 (1.60-10.05) vs grade 0; P<0.001) and haemoglobin (0.95 (0.92-0.97) per 1g/L increase; P<0.001), and P. falciparum asexual parasitaemia in slide-positive cases (0.36 (0.19-0.68) for a 10-fold increase; P=0.002). Baseline P. vivax gametocytaemia was associated with Hackett grade 2 (12.66 (1.31-122.56); P=0.028), mixed P. falciparum/vivax infection (0.16 (0.03-1.00); P=0.050), P. vivax asexual parasitaemia (5.68 (0.98-33.04); P=0.053) and haemoglobin (0.94 (0.88-1.00); P=0.056). For post-treatment P. falciparum gametocytaemia, independent predictors were AN vs AL treatment (4.09 (1.43-11.65)), haemoglobin (0.95 (0.93-0.97)), presence/absence of P. falciparum asexual forms (3.40 (1.66-0.68)) and day post-treatment (0.086 (0.82-0.90)) (P<0.001). Post-treatment P. vivax gametocytaemia was predicted by presence of P. vivax asexual forms (596 (12-28,433); P<0.001). Consistent with slow P. falciparum gametocyte maturation, low haemoglobin, low asexual parasite density and higher spleen grading, markers of increased prior infection exposure/immunity, were strong associates of pre-treatment gametocyte positivity. The persistent inverse association between P. falciparum gametocytaemia and haemoglobin during follow

  4. λ Recombination and Recombineering.

    PubMed

    Murphy, Kenan C

    2016-05-01

    The bacteriophage λ Red homologous recombination system has been studied over the past 50 years as a model system to define the mechanistic details of how organisms exchange DNA segments that share extended regions of homology. The λ Red system proved useful as a system to study because recombinants could be easily generated by co-infection of genetically marked phages. What emerged from these studies was the recognition that replication of phage DNA was required for substantial Red-promoted recombination in vivo, and the critical role that double-stranded DNA ends play in allowing the Red proteins access to the phage DNA chromosomes. In the past 16 years, however, the λ Red recombination system has gained a new notoriety. When expressed independently of other λ functions, the Red system is able to promote recombination of linear DNA containing limited regions of homology (∼50 bp) with the Escherichia coli chromosome, a process known as recombineering. This review explains how the Red system works during a phage infection, and how it is utilized to make chromosomal modifications of E. coli with such efficiency that it changed the nature and number of genetic manipulations possible, leading to advances in bacterial genomics, metabolic engineering, and eukaryotic genetics. PMID:27223821

  5. Morphological and morphometrical characterization of gametocytes of Hepatozoon procyonis Richards, 1961 (Protista, Apicomplexa) from a Brazilian wild procionid Nasua nasua and Procyon cancrivorus (Carnivora, Procyonidae).

    PubMed

    Soares Ferreira Rodrigues, André Flávio; Daemon, Erik; Massard, Carlos Luiz

    2007-01-01

    The species Hepatozoon procyonis Richards, 1961 was described in Procyon lotor in the USA and then in other reports in the USA, while in Panama H. procyonis has been described in Procyon cancrivorus. The objective of this paper is to report the occurrence of this species in the Brazilian procionids P. cancrivorus and Nasua nausa and to describe the morphology and morphometrics of the gametocytes. The analysis was based on blood smears, stained with Giemsa, which were examined under a photonic microscope. The morphometry was done with an ocular micrometer. It was based on the morphological characteristics and morphometric data on the gametocyte. It can be concluded that the species of the genus Hepatozoon that occurs in Brazilian procionids is the same as that occurring in procionids in Central and North America. PMID:16941187

  6. Frequencies of dhfr/dhps multiple mutations and Plasmodium falciparum submicroscopic gametocyte carriage in Gabonese pregnant women following IPTp-SP implementation.

    PubMed

    Bouyou-Akotet, Marielle K; Tshibola, Marie-Louise; Mawili-Mboumba, Denise P; Nzong, Julie; Bahamontes-Rosa, Noemi; Tsoumbou-Bakana, Gladys; Kombila, Maryvonne

    2015-06-01

    This study analyzed the relationship between intermittent preventive treatment with sulfadoxine-pyrimethamine (SP) (IPTp-SP), the rate of multiple resistant parasites and of submicroscopic gametocyte carriage among pregnant women at the beginning of IPTp implementation in Gabon (2005) and six years after (2011). The detection of pfdhfr and pfdhps gene mutations was performed by PCR-RFLP in Plasmodium (P.) falciparum positive samples collected from pregnant women in 2005 and 2011. Gametocytes carriage was detected by Pfs25mRNA amplification using QT-NASBA. Data were analyzed according to the time of collection (study period) and IPTp-SP doses. The proportion of isolates with at least a triple Pfdhfr mutation (n = 39/42, 92.9% versus 100%, n = 78/78)) and of those isolates with the S108N/C59R/N51I/S436A/A437G multiple mutation (17.9% versus 75.6%) significantly increased between 2005 and 2011 (p<0.01). Mutations I164L and A581G were not found, while higher proportions of 436 and 437 mutations were detected in 2011.A trend toward a higher frequency of isolates with five mutations was observed in women who received two SP doses (p<0.01). Pfs25mRNA was found in 6.8 % (n = 3/44) and 34.6% (n = 27/78) of the samples collected in 2005 and 2011 respectively (p<0.01). In 2011, 74.0% (n = 20/27) of women with detected submicroscopic gametocytes carried parasites with the S108N/C59R/N51/S436A/A437G multiple mutation. All the ten delivering women who received three IPTp-SP doses had a submicroscopic Plasmodium falciparum infection, but none had detected gametocytes. Following IPTp-SP implementation, an increase in the frequency of multiple mutant parasites and of submicroscopic gametocyte carriage was observed among pregnant women living in Gabon. PMID:26203988

  7. Genetic Recombination

    ERIC Educational Resources Information Center

    Whitehouse, H. L. K.

    1973-01-01

    Discusses the mechanisms of genetic recombination with particular emphasis on the study of the fungus Sordaria brevicollis. The study of recombination is facilitated by the use of mutants of this fungus in which the color of the ascospores is affected. (JR)

  8. Cosmological Recombination

    NASA Astrophysics Data System (ADS)

    Wong, Wan Yan

    2008-11-01

    In this thesis we focus on studying the physics of cosmological recombination and how the details of recombination affect the Cosmic Microwave Background (CMB) anisotropies. We present a detailed calculation of the spectral line distortions on the CMB spectrum arising from the Lyman-alpha and the lowest two-photon transitions in the recombination of hydrogen (H), and the corresponding lines from helium (He). The peak of these distortions mainly comes from the Lyman-alpha transition and occurs at about 170 microns, which is the Wien part of the CMB. The major theoretical limitation for extracting cosmological parameters from the CMB sky lies in the precision with which we can calculate the cosmological recombination process. With this motivation, we perform a multi-level calculation of the recombination of H and He with the addition of the spin-forbidden transition for neutral helium (He I), plus the higher order two-photon transitions for H and among singlet states of He I. We find that the inclusion of the spin-forbidden transition results in more than a percent change in the ionization fraction, while the other transitions give much smaller effects. Last we modify RECFAST by introducing one more parameter to reproduce recent numerical results for the speed-up of helium recombination. Together with the existing hydrogen `fudge factor', we vary these two parameters to account for the remaining dominant uncertainties in cosmological recombination. By using a Markov Chain Monte Carlo method with Planck forecast data, we find that we need to determine the parameters to better than 10% for He I and 1% for H, in order to obtain negligible effects on the cosmological parameters.

  9. Recombinant Pvs48/45 Antigen Expressed in E. coli Generates Antibodies that Block Malaria Transmission in Anopheles albimanus Mosquitoes

    PubMed Central

    Arévalo-Herrera, Myriam; Vallejo, Andrés F.; Rubiano, Kelly; Solarte, Yezid; Marin, Catherin; Castellanos, Angélica; Céspedes, Nora; Herrera, Sócrates

    2015-01-01

    Transmission of malaria parasites from humans to Anopheles mosquitoes can be inhibited by specific antibodies elicited during malaria infection, which target surface Plasmodium gametocyte/gamete proteins. Some of these proteins may have potential for vaccine development. Pvs48/45 is a P. vivax gametocyte surface antigen orthologous to Pfs48/45, which may play a role during parasite fertilization and thus has potential for transmission blocking (TB) activity. Here we describe the expression of a recombinant Pvs48/45 protein expressed in Escherichia coli as a ∼60kDa construct which we tested for antigenicity using human sera and for its immunogenicity and transmission blocking activity of specific anti-mouse and anti-monkey Pvs48/45 antibodies. The protein reacted with sera of individuals from malaria-endemic areas and in addition induced specific IgG antibody responses in BALB/c mice and Aotus l. griseimembra monkeys. Sera from both immunized animal species recognized native P. vivax protein in Western blot (WB) and immunofluorescence assays. Moreover, sera from immunized mice and monkeys produced significant inhibition of parasite transmission to An. Albimanus mosquitoes as shown by membrane feeding assays. Results indicate the presence of reactive epitopes in the Pvs48/45 recombinant product that induce antibodies with TB activity. Further testing of this protein is ongoing to determine its vaccine potential. PMID:25775466

  10. Spectrum Recombination.

    ERIC Educational Resources Information Center

    Greenslade, Thomas B., Jr.

    1984-01-01

    Describes several methods of executing lecture demonstrations involving the recombination of the spectrum. Groups the techniques into two general classes: bringing selected portions of the spectrum together using lenses or mirrors and blurring the colors by rapid movement or foreshortening. (JM)

  11. CRISPR-Cas9-modified pfmdr1 protects Plasmodium falciparum asexual blood stages and gametocytes against a class of piperazine-containing compounds but potentiates artemisinin-based combination therapy partner drugs.

    PubMed

    Ng, Caroline L; Siciliano, Giulia; Lee, Marcus C S; de Almeida, Mariana J; Corey, Victoria C; Bopp, Selina E; Bertuccini, Lucia; Wittlin, Sergio; Kasdin, Rachel G; Le Bihan, Amélie; Clozel, Martine; Winzeler, Elizabeth A; Alano, Pietro; Fidock, David A

    2016-08-01

    Emerging resistance to first-line antimalarial combination therapies threatens malaria treatment and the global elimination campaign. Improved therapeutic strategies are required to protect existing drugs and enhance treatment efficacy. We report that the piperazine-containing compound ACT-451840 exhibits single-digit nanomolar inhibition of the Plasmodium falciparum asexual blood stages and transmissible gametocyte forms. Genome sequence analyses of in vitro-derived ACT-451840-resistant parasites revealed single nucleotide polymorphisms in pfmdr1, which encodes a digestive vacuole membrane-bound ATP-binding cassette transporter known to alter P. falciparum susceptibility to multiple first-line antimalarials. CRISPR-Cas9 based gene editing confirmed that PfMDR1 point mutations mediated ACT-451840 resistance. Resistant parasites demonstrated increased susceptibility to the clinical drugs lumefantrine, mefloquine, quinine and amodiaquine. Stage V gametocytes harboring Cas9-introduced pfmdr1 mutations also acquired ACT-451840 resistance. These findings reveal that PfMDR1 mutations can impart resistance to compounds active against asexual blood stages and mature gametocytes. Exploiting PfMDR1 resistance mechanisms provides new opportunities for developing disease-relieving and transmission-blocking antimalarials. PMID:27073104

  12. Recombinant protein production technology

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Recombinant protein production is an important technology for antibody production, biochemical activity study, and structural determination during the post-genomic era. Limiting factors in recombinant protein production include low-level protein expression, protein precipitation, and loss of protein...

  13. Therapeutic Recombinant Monoclonal Antibodies

    ERIC Educational Resources Information Center

    Bakhtiar, Ray

    2012-01-01

    During the last two decades, the rapid growth of biotechnology-derived techniques has led to a myriad of therapeutic recombinant monoclonal antibodies with significant clinical benefits. Recombinant monoclonal antibodies can be obtained from a number of natural sources such as animal cell cultures using recombinant DNA engineering. In contrast to…

  14. Recombineering homologous recombination constructs in Drosophila.

    PubMed

    Carreira-Rosario, Arnaldo; Scoggin, Shane; Shalaby, Nevine A; Williams, Nathan David; Hiesinger, P Robin; Buszczak, Michael

    2013-01-01

    The continued development of techniques for fast, large-scale manipulation of endogenous gene loci will broaden the use of Drosophila melanogaster as a genetic model organism for human-disease related research. Recent years have seen technical advancements like homologous recombination and recombineering. However, generating unequivocal null mutations or tagging endogenous proteins remains a substantial effort for most genes. Here, we describe and demonstrate techniques for using recombineering-based cloning methods to generate vectors that can be used to target and manipulate endogenous loci in vivo. Specifically, we have established a combination of three technologies: (1) BAC transgenesis/recombineering, (2) ends-out homologous recombination and (3) Gateway technology to provide a robust, efficient and flexible method for manipulating endogenous genomic loci. In this protocol, we provide step-by-step details about how to (1) design individual vectors, (2) how to clone large fragments of genomic DNA into the homologous recombination vector using gap repair, and (3) how to replace or tag genes of interest within these vectors using a second round of recombineering. Finally, we will also provide a protocol for how to mobilize these cassettes in vivo to generate a knockout, or a tagged gene via knock-in. These methods can easily be adopted for multiple targets in parallel and provide a means for manipulating the Drosophila genome in a timely and efficient manner. PMID:23893070

  15. Photoionization and Recombination

    NASA Technical Reports Server (NTRS)

    Nahar, Sultana N.

    2000-01-01

    Theoretically self-consistent calculations for photoionization and (e + ion) recombination are described. The same eigenfunction expansion for the ion is employed in coupled channel calculations for both processes, thus ensuring consistency between cross sections and rates. The theoretical treatment of (e + ion) recombination subsumes both the non-resonant recombination ("radiative recombination"), and the resonant recombination ("di-electronic recombination") processes in a unified scheme. In addition to the total, unified recombination rates, level-specific recombination rates and photoionization cross sections are obtained for a large number of atomic levels. Both relativistic Breit-Pauli, and non-relativistic LS coupling, calculations are carried out in the close coupling approximation using the R-matrix method. Although the calculations are computationally intensive, they yield nearly all photoionization and recombination parameters needed for astrophysical photoionization models with higher precision than hitherto possible, estimated at about 10-20% from comparison with experimentally available data (including experimentally derived DR rates). Results are electronically available for over 40 atoms and ions. Photoionization and recombination of He-, and Li-like C and Fe are described for X-ray modeling. The unified method yields total and complete (e+ion) recombination rate coefficients, that can not otherwise be obtained theoretically or experimentally.

  16. Recombination of cluster ions

    NASA Technical Reports Server (NTRS)

    Johnsen, Rainer

    1993-01-01

    Some of our recent work on molecular band emissions from recombination of molecular dimer ions (N4(+) and CO(+) CO) is discussed. Much of the experimental work was done by Y. S. Cao; the results on N4(+) recombination have been published. A brief progress report is given on our ongoing measurements of neutral products of recombination using the flowing-afterglow Langmuir-probe technique in conjunction with laser-induced fluorescence.

  17. Recombination in electron coolers

    NASA Astrophysics Data System (ADS)

    Wolf, A.; Gwinner, G.; Linkemann, J.; Saghiri, A. A.; Schmitt, M.; Schwalm, D.; Grieser, M.; Beutelspacher, M.; Bartsch, T.; Brandau, C.; Hoffknecht, A.; Müller, A.; Schippers, S.; Uwira, O.; Savin, D. W.

    2000-02-01

    An introduction to electron-ion recombination processes is given and recent measurements are described as examples, focusing on low collision energies. Discussed in particular are fine-structure-mediated dielectronic recombination of fluorine-like ions, the moderate recombination enhancement by factors of typically 1.5-4 found for most ion species at relative electron-ion energies below about 10 meV, and the much larger enhancement occurring for specific highly charged ions of complex electronic structure, apparently caused by low-energy dielectronic recombination resonances. Recent experiments revealing dielectronic resonances with very large natural width are also described.

  18. Genetic recombination. [Escherichia coli

    SciTech Connect

    Stahl, F.W.

    1987-02-01

    The molecular pathways of gene recombination are explored and compared in studies of the model organisms, Escherichia coli and phase lambda. In the discussion of data from these studies it seems that recombination varies with the genetic idiosyncrasies of the organism and may also vary within a single organism.

  19. [Recombinant antibodies against bioweapons].

    PubMed

    Thullier, Philippe; Pelat, Thibaut; Vidal, Dominique

    2009-12-01

    The threat posed by bioweapons (BW) could lead to the re-emergence of such deadly diseases as plague or smallpox, now eradicated from industrialized countries. The development of recombinant antibodies allows tackling this risk because these recombinant molecules are generally well tolerated in human medicine, may be utilized for prophylaxis and treatment, and because antibodies neutralize many BW. Recombinant antibodies neutralizing the lethal toxin of anthrax, botulinum toxins and the smallpox virus have in particular been isolated recently, with different technologies. Our approach, which uses phage-displayed immune libraries built from non-human primates (M. fascicularis) to obtain recombinant antibodies, which may later be super-humanized (germlinized), has allowed us to obtain such BWs-neutralizing antibodies. PMID:20035695

  20. Activated recombinant adenovirus proteinases

    DOEpatents

    Anderson, C.W.; Mangel, W.F.

    1999-08-10

    This application describes methods and expression constructs for producing activatable recombinant adenovirus proteinases. Purified activatable recombinant adenovirus proteinases and methods of purification are described. Activated adenovirus proteinases and methods for obtaining activated adenovirus proteinases are further included. Isolated peptide cofactors of adenovirus proteinase activity, methods of purifying and identifying peptide cofactors are also described. Antibodies immunoreactive with adenovirus proteinases, immunospecific antibodies, and methods for preparing them are also described. Other related methods and materials are also described. 29 figs.

  1. Activated recombinant adenovirus proteinases

    DOEpatents

    Anderson, Carl W.; Mangel, Walter F.

    1999-08-10

    This application describes methods and expression constructs for producing activatable recombinant adenovirus proteinases. Purified activatable recombinant adenovirus proteinases and methods of purification are described. Activated adenovirus proteinases and methods for obtaining activated adenovirus proteinases are further included. Isolated peptide cofactors of adenovirus proteinase activity, methods of purifying and identifying said peptide cofactors are also described. Antibodies immunoreactive with adenovirus proteinases, immunospecific antibodies, and methods for preparing them are also described. Other related methods and materials are also described.

  2. Regulation of Meiotic Recombination

    SciTech Connect

    Gregory p. Copenhaver

    2011-11-09

    Meiotic recombination results in the heritable rearrangement of DNA, primarily through reciprocal exchange between homologous chromosome or gene conversion. In plants these events are critical for ensuring proper chromosome segregation, facilitating DNA repair and providing a basis for genetic diversity. Understanding this fundamental biological mechanism will directly facilitate trait mapping, conventional plant breeding, and development of genetic engineering techniques that will help support the responsible production and conversion of renewable resources for fuels, chemicals, and the conservation of energy (1-3). Substantial progress has been made in understanding the basal recombination machinery, much of which is conserved in organisms as diverse as yeast, plants and mammals (4, 5). Significantly less is known about the factors that regulate how often and where that basal machinery acts on higher eukaryotic chromosomes. One important mechanism for regulating the frequency and distribution of meiotic recombination is crossover interference - or the ability of one recombination event to influence nearby events. The MUS81 gene is thought to play an important role in regulating the influence of interference on crossing over. The immediate goals of this project are to use reverse genetics to identify mutants in two putative MUS81 homologs in the model plant Arabidopsis thaliana, characterize those mutants and initiate a novel forward genetic screen for additional regulators of meiotic recombination. The long-term goal of the project is to understand how meiotic recombination is regulated in higher eukaryotes with an emphasis on the molecular basis of crossover interference. The ability to monitor recombination in all four meiotic products (tetrad analysis) has been a powerful tool in the arsenal of yeast geneticists. Previously, the qrt mutant of Arabidopsis, which causes the four pollen products of male meiosis to remain attached, was developed as a facile system

  3. Meiotic recombination mechanisms.

    PubMed

    Grelon, Mathilde

    2016-01-01

    Meiosis is a specialized cell division at the origin of the haploid cells that eventually develop into the gametes. It therefore lies at the heart of Mendelian heredity. Recombination and redistribution of the homologous chromosomes arising during meiosis constitute an important source of genetic diversity, conferring to meiosis a particularly important place in the evolution and the diversification of the species. Our understanding of the molecular mechanisms governing meiotic recombination has considerably progressed these last decades, benefiting from complementary approaches led on various model species. An overview of these mechanisms will be provided as well as a discussion on the implications of these recent discoveries. PMID:27180110

  4. Recombinant renewable polyclonal antibodies

    PubMed Central

    Ferrara, Fortunato; D’Angelo, Sara; Gaiotto, Tiziano; Naranjo, Leslie; Tian, Hongzhao; Gräslund, Susanne; Dobrovetsky, Elena; Hraber, Peter; Lund-Johansen, Fridtjof; Saragozza, Silvia; Sblattero, Daniele; Kiss, Csaba; Bradbury, Andrew RM

    2015-01-01

    Only a small fraction of the antibodies in a traditional polyclonal antibody mixture recognize the target of interest, frequently resulting in undesirable polyreactivity. Here, we show that high-quality recombinant polyclonals, in which hundreds of different antibodies are all directed toward a target of interest, can be easily generated in vitro by combining phage and yeast display. We show that, unlike traditional polyclonals, which are limited resources, recombinant polyclonal antibodies can be amplified over one hundred million-fold without losing representation or functionality. Our protocol was tested on 9 different targets to demonstrate how the strategy allows the selective amplification of antibodies directed toward desirable target specific epitopes, such as those found in one protein but not a closely related one, and the elimination of antibodies recognizing common epitopes, without significant loss of diversity. These recombinant renewable polyclonal antibodies are usable in different assays, and can be generated in high throughput. This approach could potentially be used to develop highly specific recombinant renewable antibodies against all human gene products. PMID:25530082

  5. Recombinant DNA for Teachers.

    ERIC Educational Resources Information Center

    Duvall, James G., III

    1992-01-01

    A science teacher describes his experience at a workshop to learn to teach the Cold Spring Harbor DNA Science Laboratory Protocols. These protocols lead students through processes for taking E. coli cells and transforming them into a new antibiotic resistant strain. The workshop featured discussions of the role of DNA recombinant technology in…

  6. Recombineering Pseudomonas syringae

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Here we report the identification of functions that promote genomic recombination of linear DNA introduced into Pseudomonas cells by electroporation. The genes encoding these functions were identified in Pseudomonas syringae pv. syringae B728a based on similarity to the lambda Red Exo/Beta and RecE...

  7. Oligonucleotide recombination in bacteria

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Today, there are more than 1,500 completed or draft bacterial genome sequences available for public access. To functionally analyze these genomes and to test the hypotheses that are generated from the sequence information we require new and generically useful tools. Recombineering (genetic engineer...

  8. Genomic homologous recombination in planta.

    PubMed Central

    Gal, S; Pisan, B; Hohn, T; Grimsley, N; Hohn, B

    1991-01-01

    A system for monitoring intrachromosomal homologous recombination in whole plants is described. A multimer of cauliflower mosaic virus (CaMV) sequences, arranged such that CaMV could only be produced by recombination, was integrated into Brassica napus nuclear DNA. This set-up allowed scoring of recombination events by the appearance of viral symptoms. The repeated homologous regions were derived from two different strains of CaMV so that different recombinant viruses (i.e. different recombination events) could be distinguished. In most of the transgenic plants, a single major virus species was detected. About half of the transgenic plants contained viruses of the same type, suggesting a hotspot for recombination. The remainder of the plants contained viruses with cross-over sites distributed throughout the rest of the homologous sequence. Sequence analysis of two recombinant molecules suggest that mismatch repair is linked to the recombination process. Images PMID:2026150

  9. Recombinant vaccines against leptospirosis.

    PubMed

    Dellagostin, Odir A; Grassmann, André A; Hartwig, Daiane D; Félix, Samuel R; da Silva, Éverton F; McBride, Alan J A

    2011-11-01

    Leptospirosis is an important neglected infectious disease that occurs in urban environments, as well as in rural regions worldwide. Rodents, the principal reservoir hosts of pathogenic Leptospira spp., and other infected animals shed the bacteria in their urine. During occupational or even recreational activities, humans that come into direct contact with infected animals or with a contaminated environment, particularly water, are at risk of infection. Prevention of urban leptospirosis is largely dependent on sanitation measures that are often difficult to implement, especially in developing countries. Vaccination with inactivated whole-cell preparations (bacterins) has limited efficacy due to the wide antigenic variation of the pathogen. Intensive efforts towards developing improved recombinant vaccines are ongoing. During the last decade, many reports on the evaluation of recombinant vaccines have been published. Partial success has been obtained with some surface-exposed protein antigens. The combination of protective antigens and new adjuvants or delivery systems may result in the much-needed effective vaccine. PMID:22048111

  10. Recombinant influenza vaccines.

    PubMed

    Sedova, E S; Shcherbinin, D N; Migunov, A I; Smirnov, Iu A; Logunov, D Iu; Shmarov, M M; Tsybalova, L M; Naroditskiĭ, B S; Kiselev, O I; Gintsburg, A L

    2012-10-01

    This review covers the problems encountered in the construction and production of new recombinant influenza vaccines. New approaches to the development of influenza vaccines are investigated; they include reverse genetics methods, production of virus-like particles, and DNA- and viral vector-based vaccines. Such approaches as the delivery of foreign genes by DNA- and viral vector-based vaccines can preserve the native structure of antigens. Adenoviral vectors are a promising gene-delivery platform for a variety of genetic vaccines. Adenoviruses can efficiently penetrate the human organism through mucosal epithelium, thus providing long-term antigen persistence and induction of the innate immune response. This review provides an overview of the practicability of the production of new recombinant influenza cross-protective vaccines on the basis of adenoviral vectors expressing hemagglutinin genes of different influenza strains. PMID:23346377

  11. Site directed recombination

    DOEpatents

    Jurka, Jerzy W.

    1997-01-01

    Enhanced homologous recombination is obtained by employing a consensus sequence which has been found to be associated with integration of repeat sequences, such as Alu and ID. The consensus sequence or sequence having a single transition mutation determines one site of a double break which allows for high efficiency of integration at the site. By introducing single or double stranded DNA having the consensus sequence flanking region joined to a sequence of interest, one can reproducibly direct integration of the sequence of interest at one or a limited number of sites. In this way, specific sites can be identified and homologous recombination achieved at the site by employing a second flanking sequence associated with a sequence proximal to the 3'-nick.

  12. The recombination epoch revisited

    NASA Technical Reports Server (NTRS)

    Krolik, Julian H.

    1989-01-01

    Previous studies of cosmological recombination have shown that this process produces as a by-product a highly superthermal population of Ly-alpha photons which retard completion of recombination. Cosmological redshifting was thought to determine the frequency distribution of the photons, while two-photon decay of hydrogen's 2s state was thought to control their numbers. It is shown here that frequency diffusion due to photon scattering dominate the cosmological redshift in the frequency range near line center which fixes the ratio of ground state to excited state population, while incoherent scattering into the far-red damping wing effectively destroys Ly-alpha photons as a rate which is competitive with two-photon decay. The former effect tends to hold back recombination, while the latter tends to accelerate it; the net results depends on cosmological parameters, particularly the combination Omega(b) h/sq rt (2q0), where Omega(b) is the fraction of the critical density provided by baryons.

  13. Bacterial Recombineering: Genome Engineering via Phage-Based Homologous Recombination.

    PubMed

    Pines, Gur; Freed, Emily F; Winkler, James D; Gill, Ryan T

    2015-11-20

    The ability to specifically modify bacterial genomes in a precise and efficient manner is highly desired in various fields, ranging from molecular genetics to metabolic engineering and synthetic biology. Much has changed from the initial realization that phage-derived genes may be employed for such tasks to today, where recombineering enables complex genetic edits within a genome or a population. Here, we review the major developments leading to recombineering becoming the method of choice for in situ bacterial genome editing while highlighting the various applications of recombineering in pushing the boundaries of synthetic biology. We also present the current understanding of the mechanism of recombineering. Finally, we discuss in detail issues surrounding recombineering efficiency and future directions for recombineering-based genome editing. PMID:25856528

  14. Dielectronic recombination theory

    SciTech Connect

    LaGattuta, K.J.

    1991-12-31

    A theory now in wide use for the calculation of dielectronic recombination cross sections ({sigma}{sup DR}) and rate coefficients ({alpha}{sup DR}) was one introduced originally by Feshbach for nuclear physics applications, and then later adapted for atomic scattering problems by Hahn. In the following, we briefly review this theory in a very general form, which allows one to account for the effects of overlapping and interacting resonances, as well as continuum-continuum coupling. An extension of our notation will then also allow for the inclusion of the effects of direct radiative recombination, along with a treatment of the interference between radiative and dielectronic recombination. Other approaches to the calculation of {sigma}{sup DR} have been described by Fano and by Seaton. We will not consider those theories here. Calculations of {alpha}{sup DR} have progressed considerably over the last 25 years, since the early work of Burgess. Advances in the reliability of theoretical predictions have also been promoted recently b a variety of direct laboratory measurements of {sigma}{sup DR}. While the measurements of {sigma}{sup DR} for {delta}n {ne} 0 excitations have tended to agree very well with calculations, the case of {delta}n = 0 has been much problematic. However, by invoking a mechanism originally proposed by Jacobs, which takes into account the effect of stray electric fields on high Rydberg states (HRS) participating in the DR process, new calculations have improved the agreement between theory and experiment for these cases. Nevertheless, certain discrepancies still remain.

  15. Recombinant Collagenlike Proteins

    NASA Technical Reports Server (NTRS)

    Fertala, Andzej

    2007-01-01

    A group of collagenlike recombinant proteins containing high densities of biologically active sites has been invented. The method used to express these proteins is similar to a method of expressing recombinant procollagens and collagens described in U. S. Patent 5,593,859, "Synthesis of human procollagens and collagens in recombinant DNA systems." Customized collagenous proteins are needed for biomedical applications. In particular, fibrillar collagens are attractive for production of matrices needed for tissue engineering and drug delivery. Prior to this invention, there was no way of producing customized collagenous proteins for these and other applications. Heretofore, collagenous proteins have been produced by use of such biological systems as yeasts, bacteria, and transgenic animals and plants. These products are normal collagens that can also be extracted from such sources as tendons, bones, and hides. These products cannot be made to consist only of biologically active, specific amino acid sequences that may be needed for specific applications. Prior to this invention, it had been established that fibrillar collagens consist of domains that are responsible for such processes as interaction with cells, binding of growth factors, and interaction with a number of structural proteins present in the extracellular matrix. A normal collagen consists of a sequence of domains that can be represented by a corresponding sequence of labels, e.g., D1D2D3D4. A collagenlike protein of the present invention contains regions of collagen II that contain multiples of a single domain (e.g., D1D1D1D1 or D4D4D4D4) chosen for its specific biological activity. By virtue of the multiplicity of the chosen domain, the density of sites having that specific biological activity is greater than it is in a normal collagen. A collagenlike protein according to this invention can thus be made to have properties that are necessary for tissue engineering.

  16. Did the universe recombine?

    NASA Technical Reports Server (NTRS)

    Bartlett, James G.; Stebbins, Albert

    1991-01-01

    The Zel'dovich-Sunyaev model-independent arguments for the existence of a neutral hydrogen phase is reviewed in light of new limits on the Compton y parameter from COBE. It is concluded that with baryon densities compatible with standard cosmological nucleosynthesis, the universe could have remained fully ionized throughout its history without producing a detectable spectral distortion. It is argued that it is unlikely that spectral observations of the cosmic microwave background will ever require the universe to have recombined for flat cosmologies.

  17. Expression of Recombinant Antibodies

    PubMed Central

    Frenzel, André; Hust, Michael; Schirrmann, Thomas

    2013-01-01

    Recombinant antibodies are highly specific detection probes in research, diagnostics, and have emerged over the last two decades as the fastest growing class of therapeutic proteins. Antibody generation has been dramatically accelerated by in vitro selection systems, particularly phage display. An increasing variety of recombinant production systems have been developed, ranging from Gram-negative and positive bacteria, yeasts and filamentous fungi, insect cell lines, mammalian cells to transgenic plants and animals. Currently, almost all therapeutic antibodies are still produced in mammalian cell lines in order to reduce the risk of immunogenicity due to altered, non-human glycosylation patterns. However, recent developments of glycosylation-engineered yeast, insect cell lines, and transgenic plants are promising to obtain antibodies with “human-like” post-translational modifications. Furthermore, smaller antibody fragments including bispecific antibodies without any glycosylation are successfully produced in bacteria and have advanced to clinical testing. The first therapeutic antibody products from a non-mammalian source can be expected in coming next years. In this review, we focus on current antibody production systems including their usability for different applications. PMID:23908655

  18. Recombinant factor VIIa.

    PubMed

    Aitken, Michael G

    2004-01-01

    Human coagulation factor (F) VII is a single chain protease that circulates in the blood as a weakly active zymogen at concentrations of approximately 10 nmol/L. When converted to the active 2 chain form (FVIIa), it is a powerful initiator of haemostasis. Recombinant factor VIIa (rFVIIa, eptacog alfa, NovoSeven) is a genetically engineered product that was first introduced in 1988 for the treatment of patients with haemophilia A and B with high inhibitory antibody titres to factors VIII and IX. Recent reports in the form of case studies and series, and early trial data, have suggested a role for rFVIIa across a diverse range of indications including bleeding associated with trauma, surgery, thrombocytopaenia, liver disease and oral anticoagulant toxicity. This review describes the physiology of the coagulation pathway and in particular the role of recombinant factor VIIa. It will also focus on the emerging role of rFVIIa in both trauma and non-trauma bleeding and its potential use in the ED. PMID:15537408

  19. Unraveling recombination rate evolution using ancestral recombination maps

    PubMed Central

    Munch, Kasper; Schierup, Mikkel H; Mailund, Thomas

    2014-01-01

    Recombination maps of ancestral species can be constructed from comparative analyses of genomes from closely related species, exemplified by a recently published map of the human-chimpanzee ancestor. Such maps resolve differences in recombination rate between species into changes along individual branches in the speciation tree, and allow identification of associated changes in the genomic sequences. We describe how coalescent hidden Markov models are able to call individual recombination events in ancestral species through inference of incomplete lineage sorting along a genomic alignment. In the great apes, speciation events are sufficiently close in time that a map can be inferred for the ancestral species at each internal branch - allowing evolution of recombination rate to be tracked over evolutionary time scales from speciation event to speciation event. We see this approach as a way of characterizing the evolution of recombination rate and the genomic properties that influence it. PMID:25043668

  20. Algebraic theory of recombination spaces.

    PubMed

    Stadler, P F; Wagner, G P

    1997-01-01

    A new mathematical representation is proposed for the configuration space structure induced by recombination, which we call "P-structure." It consists of a mapping of pairs of objects to the power set of all objects in the search space. The mapping assigns to each pair of parental "genotypes" the set of all recombinant genotypes obtainable from the parental ones. It is shown that this construction allows a Fourier decomposition of fitness landscapes into a superposition of "elementary landscapes." This decomposition is analogous to the Fourier decomposition of fitness landscapes on mutation spaces. The elementary landscapes are obtained as eigenfunctions of a Laplacian operator defined for P-structures. For binary string recombination, the elementary landscapes are exactly the p-spin functions (Walsh functions), that is, the same as the elementary landscapes of the string point mutation spaces (i.e., the hypercube). This supports the notion of a strong homomorphism between string mutation and recombination spaces. However, the effective nearest neighbor correlations on these elementary landscapes differ between mutation and recombination and among different recombination operators. On average, the nearest neighbor correlation is higher for one-point recombination than for uniform recombination. For one-point recombination, the correlations are higher for elementary landscapes with fewer interacting sites as well as for sites that have closer linkage, confirming the qualitative predictions of the Schema Theorem. We conclude that the algebraic approach to fitness landscape analysis can be extended to recombination spaces and provides an effective way to analyze the relative hardness of a landscape for a given recombination operator. PMID:10021760

  1. Recombinant Human Erythropoietin

    PubMed Central

    Bartels, Claudia; Späte, Kira; Krampe, Henning

    2008-01-01

    Treatment of multiple sclerosis (MS) is still unsatisfactory and essentially non-existing for the progressive course of the disease. Recombinant human erythropoietin (EPO) may be a promising neuroprotective/neuroregenerative treatment of MS. In the nervous system, EPO acts anti-apoptotic, antioxidative, anti-inflammatory, neurotrophic and plasticity-modulating. Beneficial effects have been shown in animal models of various neurological and psychiatric diseases, including different models of experimental autoimmune encephalomyelitis. EPO is also effective in human brain disease, as shown in double-blind placebo-controlled clinical studies on ischemic stroke and chronic schizophrenia. An exploratory study on chronic progressive MS yielded lasting improvement in motor and cognitive performance upon high-dose long-term EPO treatment. PMID:21180577

  2. Recombinant glucose uptake system

    DOEpatents

    Ingrahm, Lonnie O.; Snoep, Jacob L.; Arfman, Nico

    1997-01-01

    Recombinant organisms are disclosed that contain a pathway for glucose uptake other than the pathway normally utilized by the host cell. In particular, the host cell is one in which glucose transport into the cell normally is coupled to PEP production. This host cell is transformed so that it uses an alternative pathway for glucose transport that is not coupled to PEP production. In a preferred embodiment, the host cell is a bacterium other than Z. mobilis that has been transformed to contain the glf and glk genes of Z. mobilis. By uncoupling glucose transport into the cell from PEP utilization, more PEP is produced for synthesis of products of commercial importance from a given quantity of biomass supplied to the host cells.

  3. The recombination of genetic material

    SciTech Connect

    Low, K.B.

    1988-01-01

    Genetic recombination is the major mechanism by which new arrangements of genetic elements are produced in all living organisms, from the simplest bacterial viruses to humans. This volume presents an overview of the types of recombination found in prokaryotes and eukaryotes.

  4. Coalescent Simulation of Intracodon Recombination

    PubMed Central

    Arenas, Miguel; Posada, David

    2010-01-01

    The coalescent with recombination is a very useful tool in molecular population genetics. Under this framework, genealogies often represent the evolution of the substitution unit, and because of this, the few coalescent algorithms implemented for the simulation of coding sequences force recombination to occur only between codons. However, it is clear that recombination is expected to occur most often within codons. Here we have developed an algorithm that can evolve coding sequences under an ancestral recombination graph that represents the genealogies at each nucleotide site, thereby allowing for intracodon recombination. The algorithm is a modification of Hudson's coalescent in which, in addition to keeping track of events occurring in the ancestral material that reaches the sample, we need to keep track of events occurring in ancestral material that does not reach the sample but that is produced by intracodon recombination. We are able to show that at typical substitution rates the number of nonsynonymous changes induced by intracodon recombination is small and that intracodon recombination does not generally result in inflated estimates of the overall nonsynonymous/synonymous substitution ratio (ω). On the other hand, recombination can bias the estimation of ω at particular codons, resulting in apparent rate variation among sites and in the spurious identification of positively selected sites. Importantly, in this case, allowing for variable synonymous rates across sites greatly reduces the false-positive rate and recovers statistical power. Finally, coalescent simulations with intracodon recombination could be used to better represent the evolution of nuclear coding genes or fast-evolving pathogens such as HIV-1.We have implemented this algorithm in a computer program called NetRecodon, freely available at http://darwin.uvigo.es. PMID:19933876

  5. Delayed recombination and standard rulers

    SciTech Connect

    De Bernardis, Francesco; Melchiorri, Alessandro; Bean, Rachel; Galli, Silvia; Silk, Joseph I.; Verde, Licia

    2009-02-15

    Measurements of baryonic acoustic oscillations (BAOs) in galaxy surveys have been recognized as a powerful tool for constraining dark energy. However, this method relies on the knowledge of the size of the acoustic horizon at recombination derived from cosmic microwave background (CMB) anisotropy measurements. This estimate is typically derived assuming a standard recombination scheme; additional radiation sources can delay recombination altering the cosmic ionization history and the cosmological inferences drawn from CMB and BAO data. In this paper we quantify the effect of delayed recombination on the determination of dark energy parameters from future BAO surveys such as the Baryon Oscillation Spectroscopic Survey and the Wide-Field Multi-Object Spectrograph. We find the impact to be small but still not negligible. In particular, if recombination is nonstandard (to a level still allowed by CMB data), but this is ignored, future surveys may incorrectly suggest the presence of a redshift-dependent dark energy component. On the other hand, in the case of delayed recombination, adding to the analysis one extra parameter describing deviations from standard recombination does not significantly degrade the error bars on dark energy parameters and yields unbiased estimates. This is due to the CMB-BAO complementarity.

  6. Three Decades of Recombinant DNA.

    ERIC Educational Resources Information Center

    Palmer, Jackie

    1985-01-01

    Discusses highlights in the development of genetic engineering, examining techniques with recombinant DNA, legal and ethical issues, GenBank (a national database of nucleic acid sequences), and other topics. (JN)

  7. Controlled Release from Recombinant Polymers

    PubMed Central

    Price, Robert; Poursaid, Azadeh; Ghandehari, Hamidreza

    2014-01-01

    Recombinant polymers provide a high degree of molecular definition for correlating structure with function in controlled release. The wide array of amino acids available as building blocks for these materials lend many advantages including biorecognition, biodegradability, potential biocompatibility, and control over mechanical properties among other attributes. Genetic engineering and DNA manipulation techniques enable the optimization of structure for precise control over spatial and temporal release. Unlike the majority of chemical synthetic strategies used, recombinant DNA technology has allowed for the production of monodisperse polymers with specifically defined sequences. Several classes of recombinant polymers have been used for controlled drug delivery. These include, but are not limited to, elastin-like, silk-like, and silk-elastinlike proteins, as well as emerging cationic polymers for gene delivery. In this article, progress and prospects of recombinant polymers used in controlled release will be reviewed. PMID:24956486

  8. Recombinant DNA means and method

    SciTech Connect

    Alford, B.L.; Mao, J.I.; Moir, D.T.; Taunton-Rigby, A.; Vovis, G.F.

    1987-05-19

    This patent describes a transformed living cell selected from the group consisting of fungi, yeast and bacteria, and containing genetic material derived from recombinant DNA material and coding for bovine rennin.

  9. Stable recombination hotspots in birds.

    PubMed

    Singhal, Sonal; Leffler, Ellen M; Sannareddy, Keerthi; Turner, Isaac; Venn, Oliver; Hooper, Daniel M; Strand, Alva I; Li, Qiye; Raney, Brian; Balakrishnan, Christopher N; Griffith, Simon C; McVean, Gil; Przeworski, Molly

    2015-11-20

    The DNA-binding protein PRDM9 has a critical role in specifying meiotic recombination hotspots in mice and apes, but it appears to be absent from other vertebrate species, including birds. To study the evolution and determinants of recombination in species lacking the gene that encodes PRDM9, we inferred fine-scale genetic maps from population resequencing data for two bird species: the zebra finch, Taeniopygia guttata, and the long-tailed finch, Poephila acuticauda. We found that both species have recombination hotspots, which are enriched near functional genomic elements. Unlike in mice and apes, most hotspots are shared between the two species, and their conservation seems to extend over tens of millions of years. These observations suggest that in the absence of PRDM9, recombination targets functional features that both enable access to the genome and constrain its evolution. PMID:26586757

  10. Recombination device for storage batteries

    DOEpatents

    Kraft, H.; Ledjeff, K.

    1984-01-01

    A recombination device including a gas-tight enclosure connected to receive the discharge gases from a rechargeable storage battery. Catalytic material for the recombination of hydrogen and oxygen to form water is supported within the enclosure. The enclosure is sealed from the atmosphere by a liquid seal including two vertical chambers interconnected with an inverted U-shaped overflow tube. The first chamber is connected at its upper portion to the enclosure and the second chamber communicates at its upper portion with the atmosphere. If the pressure within the enclosure differs as overpressure or vacuum by more than the liquid level, the liquid is forced into one of the two chambers and the overpressure is vented or the vacuum is relieved. The recombination device also includes means for returning recombined liquid to the battery and for absorbing metal hydrides.

  11. Recombination device for storage batteries

    DOEpatents

    Kraft, Helmut; Ledjeff, Konstantin

    1985-01-01

    A recombination device including a gas-tight enclosure connected to receive he discharge gases from a rechargeable storage battery. Catalytic material for the recombination of hydrogen and oxygen to form water is supported within the enclosure. The enclosure is sealed from the atmosphere by a liquid seal including two vertical chambers interconnected with an inverted U-shaped overflow tube. The first chamber is connected at its upper portion to the enclosure and the second chamber communicates at its upper portion with the atmosphere. If the pressure within the enclosure differs as overpressure or vacuum by more than the liquid level, the liquid is forced into one of the two chambers and the overpressure is vented or the vacuum is relieved. The recombination device also includes means for returning recombined liquid to the battery and for absorbing metal hydrides.

  12. [Antithrombotic recombinant antibodies].

    PubMed

    Muzard, Julien; Loyau, Stéphane; Ajzenberg, Nadine; Billiald, Philippe; Jandrot-Perrus, Martine

    2006-01-01

    Coronary syndromes, stroke and other ischaemic arterial diseases are the leading cause of death in the world and will probably remain it at least until 2020. Cardiovascular diseases kill 17 million people each year with an expected increase to 20 million in 2020 and 24 million in 2030. The global impact of recurrence and death during the 6 months following an acute coronary syndrome remains at 8-15% in the present state of medical practice. Acute ischaemic syndromes have a common aetiology that is the formation of a platelet-rich clot at the site of severe coronary stenosis and of eroded atherosclerotic plaques. Therapy consists of medical treatments associating thrombolysis, antiplatelet drugs, and the re-opening of the coronary artery by angioplasty. But these treatments do not prevent morbidity and mortality reaching 15% at 6 months. Finally the treatment of stroke is very limited. There is thus a real clinical need to improve existing treatments and to discover new molecules. Platelet activation is a critical step in ischaemic cardiovascular diseases. This is the reason why antiplatelet drugs are most often prescribed in these cases. Currently, only one recombinant antithrombotic antibody is used in therapy. This is a chimeric Fab, c7E3 or abciximab, which inhibits the final phase of platelet aggregation. Abciximab is prescribed in acute coronary syndromes treated by angioplasty. However, treatment by abciximab can induce severe complications, principally, hemorrages and thrombopenia. Other platelet receptors involved in the earlier steps of platelet activation, such as the phases of contact with and of activation by the subendothelium matrix, have been identified as potential targets for the development of antithrombotic antibodies and are described in this revue. PMID:17652972

  13. Genetic recombination in Streptomyces griseus.

    PubMed Central

    Parag, Y

    1978-01-01

    Low-frequency (10(-6)) genetic recombination was observed in a cephamycin-producing strain of Streptomyces griseus. The recombinants were predominantly heteroclones. Heteroclone analysis was performed involving four heteroclones of one cross. In 100 mutants correlation was found between the type of auxotrophy and the level of antibiotic activity. A cross of this strain with a streptomycin-producing strain of S. griesus is described. PMID:415037

  14. [Vaccine application of recombinant herpesviruses].

    PubMed

    Yokoyama, N; Xuan, X; Mikami, T

    2000-04-01

    Recently, genetic engineering using recombinant DNA techniques has been applied to design new viral vaccines in order to reduce some problems which the present viral vaccines have. Up to now, many viruses have been investigated for development of recombinant attenuated vaccines or live viral vectors for delivery of foreign genes coding immunogenic antigens. In this article, we introduced the new vaccine strategy using genetically engineered herpesviruses. PMID:10774221

  15. Combinatorics in Recombinational Population Genomics

    NASA Astrophysics Data System (ADS)

    Parida, Laxmi

    The work that I will discuss is motivated by the need for understanding, and processing, the manifestations of recombination events in chromosome sequences. In this talk, we focus on two related problems. First, we explore the very general problem of reconstructability of pedigree history. How plausible is it to unravel the history of a complete unit (chromosome) of inheritance? The second problem deals with reconstructing the recombinational history of a collection of chromosomes.

  16. Delayed recombination and cosmic parameters

    NASA Astrophysics Data System (ADS)

    Galli, Silvia; Bean, Rachel; Melchiorri, Alessandro; Silk, Joseph

    2008-09-01

    Current cosmological constraints from cosmic microwave background anisotropies are typically derived assuming a standard recombination scheme, however additional resonance and ionizing radiation sources can delay recombination, altering the cosmic ionization history and the cosmological inferences drawn from the cosmic microwave background data. We show that for recent observations of the cosmic microwave background anisotropy, from the Wilkinson microwave anisotropy probe satellite mission (WMAP) 5-year survey and from the arcminute cosmology bolometer array receiver experiment, additional resonance radiation is nearly degenerate with variations in the spectral index, ns, and has a marked effect on uncertainties in constraints on the Hubble constant, age of the universe, curvature and the upper bound on the neutrino mass. When a modified recombination scheme is considered, the redshift of recombination is constrained to z*=1078±11, with uncertainties in the measurement weaker by 1 order of magnitude than those obtained under the assumption of standard recombination while constraints on the shift parameter are shifted by 1σ to R=1.734±0.028. From the WMAP5 data we obtain the following constraints on the resonance and ionization sources parameters: γα<0.39 and γi<0.058 at 95% c.l.. Although delayed recombination limits the precision of parameter estimation from the WMAP satellite, we demonstrate that this should not be the case for future, smaller angular scales measurements, such as those by the Planck satellite mission.

  17. Ethanol production by recombinant hosts

    DOEpatents

    Ingram, Lonnie O.; Beall, David S.; Burchhardt, Gerhard F. H.; Guimaraes, Walter V.; Ohta, Kazuyoshi; Wood, Brent E.; Shanmugam, Keelnatham T.

    1995-01-01

    Novel plasmids comprising genes which code for the alcohol dehydrogenase and pyruvate decarboxylase are described. Also described are recombinant hosts which have been transformed with genes coding for alcohol dehydrogenase and pyruvate. By virtue of their transformation with these genes, the recombinant hosts are capable of producing significant amounts of ethanol as a fermentation product. Also disclosed are methods for increasing the growth of recombinant hosts and methods for reducing the accumulation of undesirable metabolic products in the growth medium of these hosts. Also disclosed are recombinant host capable of producing significant amounts of ethanol as a fermentation product of oligosaccharides and plasmids comprising genes encoding polysaccharases, in addition to the genes described above which code for the alcohol dehydrogenase and pyruvate decarboxylase. Further, methods are described for producing ethanol from oligomeric feedstock using the recombinant hosts described above. Also provided is a method for enhancing the production of functional proteins in a recombinant host comprising overexpressing an adhB gene in the host. Further provided are process designs for fermenting oligosaccharide-containing biomass to ethanol.

  18. Ethanol production by recombinant hosts

    DOEpatents

    Fowler, David E.; Horton, Philip G.; Ben-Bassat, Arie

    1996-01-01

    Novel plasmids comprising genes which code for the alcohol dehydrogenase and pyruvate decarboxylase are described. Also described are recombinant hosts which have been transformed with genes coding for alcohol dehydrogenase and pyruvate. By virtue of their transformation with these genes, the recombinant hosts are capable of producing significant amounts of ethanol as a fermentation product. Also disclosed are methods for increasing the growth of recombinant hosts and methods for reducing the accumulation of undesirable metabolic products in the growth medium of these hosts. Also disclosed are recombinant host capable of producing significant amounts of ethanol as a fermentation product of oligosaccharides and plasmids comprising genes encoding polysaccharases, in addition to the genes described above which code for the alcohol dehydrogenase and pyruvate decarboxylase. Further, methods are described for producing ethanol from oligomeric feedstock using the recombinant hosts described above. Also provided is a method for enhancing the production of functional proteins in a recombinant host comprising overexpressing an adhB gene in the host. Further provided are process designs for fermenting oligosaccharide-containing biomass to ethanol.

  19. Delayed recombination and cosmic parameters

    SciTech Connect

    Galli, Silvia; Melchiorri, Alessandro; Bean, Rachel; Silk, Joseph

    2008-09-15

    Current cosmological constraints from cosmic microwave background anisotropies are typically derived assuming a standard recombination scheme, however additional resonance and ionizing radiation sources can delay recombination, altering the cosmic ionization history and the cosmological inferences drawn from the cosmic microwave background data. We show that for recent observations of the cosmic microwave background anisotropy, from the Wilkinson microwave anisotropy probe satellite mission (WMAP) 5-year survey and from the arcminute cosmology bolometer array receiver experiment, additional resonance radiation is nearly degenerate with variations in the spectral index, n{sub s}, and has a marked effect on uncertainties in constraints on the Hubble constant, age of the universe, curvature and the upper bound on the neutrino mass. When a modified recombination scheme is considered, the redshift of recombination is constrained to z{sub *}=1078{+-}11, with uncertainties in the measurement weaker by 1 order of magnitude than those obtained under the assumption of standard recombination while constraints on the shift parameter are shifted by 1{sigma} to R=1.734{+-}0.028. From the WMAP5 data we obtain the following constraints on the resonance and ionization sources parameters: {epsilon}{sub {alpha}}<0.39 and {epsilon}{sub i}<0.058 at 95% c.l.. Although delayed recombination limits the precision of parameter estimation from the WMAP satellite, we demonstrate that this should not be the case for future, smaller angular scales measurements, such as those by the Planck satellite mission.

  20. Recombination Drives Vertebrate Genome Contraction

    PubMed Central

    Nam, Kiwoong; Ellegren, Hans

    2012-01-01

    Selective and/or neutral processes may govern variation in DNA content and, ultimately, genome size. The observation in several organisms of a negative correlation between recombination rate and intron size could be compatible with a neutral model in which recombination is mutagenic for length changes. We used whole-genome data on small insertions and deletions within transposable elements from chicken and zebra finch to demonstrate clear links between recombination rate and a number of attributes of reduced DNA content. Recombination rate was negatively correlated with the length of introns, transposable elements, and intergenic spacer and with the rate of short insertions. Importantly, it was positively correlated with gene density, the rate of short deletions, the deletion bias, and the net change in sequence length. All these observations point at a pattern of more condensed genome structure in regions of high recombination. Based on the observed rates of small insertions and deletions and assuming that these rates are representative for the whole genome, we estimate that the genome of the most recent common ancestor of birds and lizards has lost nearly 20% of its DNA content up until the present. Expansion of transposable elements can counteract the effect of deletions in an equilibrium mutation model; however, since the activity of transposable elements has been low in the avian lineage, the deletion bias is likely to have had a significant effect on genome size evolution in dinosaurs and birds, contributing to the maintenance of a small genome. We also demonstrate that most of the observed correlations between recombination rate and genome contraction parameters are seen in the human genome, including for segregating indel polymorphisms. Our data are compatible with a neutral model in which recombination drives vertebrate genome size evolution and gives no direct support for a role of natural selection in this process. PMID:22570634

  1. Recombination processes in ionised plasmas

    NASA Astrophysics Data System (ADS)

    Bastin, Robert

    The observational analysis of astrophysical plasmas relies on accurate calculations of the atomic processes involved. The recombination spectra of singly ionised oxygen (O il) and carbon (C il) present excellent tools for investigating regions such as planetary nebulae and H II regions. In this thesis, detailed treatments of the recombination processes of both O II and C II are presented. Using the R-matrix solution to the close coupling equations, I present the results of accurate photoionisation calculations. Bound state energy levels are determined and oscillator strengths calculated for both species. Recombination coefficients were evalu ated for low n and 1, for C II in LS-coupling, and 0 II in intermediate coupling, taking particular care to treat resonances effectively. Sample photoionisation cross-sections are presented for both species, and compared to previous work. A complete radiative-cascade model is treated for both species, in order to determine line emissivities under nebular conditions at a wide range of temperatures and densities. Collisional effects are treated for C II, along with, for the first time, the effects of high temperature dielectronic recombination, allowing the modelling of regions of much higher electron temperature than previous work. The O II calculations were performed under intermediate coupling for the first time, allowing the effects of non-statistical popula tions of the parent ion fine-structure levels and dielectronic recombination onto bound states within this fine-structure to be taken into account in line emissivities. Detailed comparison with previous theoretical work was made for both species. The application of the C II and 0 n recombination spectra to determining tempera ture and densities from the observed spectra of a number of ionised nebulae is considered. The potential for using the new recombination spectra as diagnostic tools to solve some of the key problems in the study of ionised nebulae is demonstrated.

  2. Recombination at the DNA level. Abstracts

    SciTech Connect

    Not Available

    1984-01-01

    Abstracts of papers in the following areas are presented: (1) chromosome mechanics; (2) yeast systems; (3) mammalian homologous recombination; (4) transposons; (5) Mu; (6) plant transposons/T4 recombination; (7) topoisomerase, resolvase, and gyrase; (8) Escherichia coli general recombination; (9) recA; (10) repair; (11) eucaryotic enzymes; (12) integration and excision of bacteriophage; (13) site-specific recombination; and (14) recombination in vitro. (ACR)

  3. PROGENITORS OF RECOMBINING SUPERNOVA REMNANTS

    SciTech Connect

    Moriya, Takashi J.

    2012-05-01

    Usual supernova remnants have either ionizing plasma or plasma in collisional ionization equilibrium, i.e., the ionization temperature is lower than or equal to the electron temperature. However, the existence of recombining supernova remnants, i.e., supernova remnants with ionization temperature higher than the electron temperature, has been recently confirmed. One suggested way to have recombining plasma in a supernova remnant is to have a dense circumstellar medium at the time of the supernova explosion. If the circumstellar medium is dense enough, collisional ionization equilibrium can be established in the early stage of the evolution of the supernova remnant and subsequent adiabatic cooling, which occurs after the shock wave gets out of the dense circumstellar medium, makes the electron temperature lower than the ionization temperature. We study the circumstellar medium around several supernova progenitors and show which supernova progenitors can have a circumstellar medium dense enough to establish collisional ionization equilibrium soon after the explosion. We find that the circumstellar medium around red supergiants (especially massive ones) and the circumstellar medium dense enough to make Type IIn supernovae can establish collisional ionization equilibrium soon after the explosion and can evolve to become recombining supernova remnants. Wolf-Rayet stars and white dwarfs have the possibility to be recombining supernova remnants but the fraction is expected to be very small. As the occurrence rate of the explosions of red supergiants is much higher than that of Type IIn supernovae, the major progenitors of recombining supernova remnants are likely to be red supergiants.

  4. Recombinant allergens for specific immunotherapy.

    PubMed

    Cromwell, Oliver; Häfner, Dietrich; Nandy, Andreas

    2011-04-01

    Recombinant DNA technology provides the means for producing allergens that are equivalent to their natural counterparts and also genetically engineered variants with reduced IgE-binding activity. The proteins are produced as chemically defined molecules with consistent structural and immunologic properties. Several hundred allergens have been cloned and expressed as recombinant proteins, and these provide the means for making a very detailed diagnosis of a patient's sensitization profile. Clinical development programs are now in progress to assess the suitability of recombinant allergens for both subcutaneous and sublingual immunotherapy. Recombinant hypoallergenic variants, which are developed with the aim of increasing the doses that can be administered while at the same time reducing the risks for therapy-associated side effects, are also in clinical trials for subcutaneous immunotherapy. Grass and birch pollen preparations have been shown to be clinically effective, and studies with various other allergens are in progress. Personalized or patient-tailored immunotherapy is still a very distant prospect, but the first recombinant products based on single allergens or defined mixtures could reach the market within the next 5 years. PMID:21377719

  5. Stable recombination hotspots in birds

    PubMed Central

    Singhal, Sonal; Leffler, Ellen M.; Sannareddy, Keerthi; Turner, Isaac; Venn, Oliver; Hooper, Daniel M.; Strand, Alva I.; Li, Qiye; Raney, Brian; Balakrishnan, Christopher N.; Griffith, Simon C.; McVean, Gil; Przeworski, Molly

    2016-01-01

    The DNA-binding protein PRDM9 has a critical role in specifying meiotic recombination hotspots in mice and apes, but appears to be absent from other vertebrate species, including birds. To study the evolution and determinants of recombination in species lacking PRDM9, we inferred fine-scale genetic maps from population resequencing data for two bird species, the zebra finch Taeniopygia guttata and the long-tailed finch Poephila acuticauda. We find that both species have hotspots, which are enriched near functional genomic elements. Unlike in mice and apes, the two species share most hotspots, with conservation seemingly extending over tens of millions of years. These observations suggest that in the absence of PRDM9, recombination targets functional features that both enable access to the genome and constrain its evolution. PMID:26586757

  6. Recombinant snake venom prothrombin activators.

    PubMed

    Lövgren, Ann

    2013-01-01

    Three prothrombin activators; ecarin, which was originally isolated from the venom of the saw-scaled viper Echis carinatus, trocarin from the rough-scaled snake Tropidechis carinatus, and oscutarin from the Taipan snake Oxyuranus scutellatus, were expressed in mammalian cells with the purpose to obtain recombinant prothrombin activators that could be used to convert prothrombin to thrombin. We have previously reported that recombinant ecarin can efficiently generate thrombin without the need for additional cofactors, but does not discriminate non-carboxylated prothrombin from biologically active γ-carboxylated prothrombin. Here we report that recombinant trocarin and oscutarin could not efficiently generate thrombin without additional protein co-factors. We confirm that both trocarin and oscutarin are similar to human coagulation Factor X (FX), explaining the need for additional cofactors. Sequencing of a genomic fragment containing 7 out of the 8 exons coding for oscutarin further confirmed the similarity to human FX. PMID:23111318

  7. The Dissociative Recombination of OH(+)

    NASA Technical Reports Server (NTRS)

    Guberman, Steven L.

    1995-01-01

    Theoretical quantum chemical calculations of the cross sections and rates for the dissociative recombination of the upsilon = 0 level of the ground state of OH(+) show that recombination occurs primarily along the 2 (2)Pi diabatic route. The products are 0((1)D) and a hot H atom with 6.1 eV kinetic energy. The coupling to the resonances is very small and the indirect recombination mechanism plays only a minor role. The recommended value for the rate coefficient is (6.3 +/- 0.7) x 10(exp -9)x (T(e)/1300)(exp -0.48) cu.cm/s for 10 less than T(e) less than 1000 K.

  8. Current Drive in Recombining Plasma

    SciTech Connect

    P.F. Schmit and N.J. Fisch

    2012-05-15

    The Langevin equations describing the average collisional dynamics of suprathermal particles in nonstationary plasma remarkably admit an exact analytical solution in the case of recombining plasma. The current density produced by arbitrary particle fluxes is derived including the effect of charge recombination. Since recombination has the effect of lowering the charge density of the plasma, thus reducing the charged particle collisional frequencies, the evolution of the current density can be modified substantially compared to plasma with fixed charge density. The current drive efficiency is derived and optimized for discrete and continuous pulses of current, leading to the discovery of a nonzero "residual" current density that persists indefinitely under certain conditions, a feature not present in stationary plasmas.

  9. Selenium incorporation using recombinant techniques

    SciTech Connect

    Walden, Helen

    2010-04-01

    An overview of techniques for recombinant incorporation of selenium and subsequent purification and crystallization of the resulting labelled protein. Using selenomethionine to phase macromolecular structures is common practice in structure determination, along with the use of selenocysteine. Selenium is consequently the most commonly used heavy atom for MAD. In addition to the well established recombinant techniques for the incorporation of selenium in prokaryal expression systems, there have been recent advances in selenium labelling in eukaryal expression, which will be discussed. Tips and things to consider for the purification and crystallization of seleno-labelled proteins are also included.

  10. Classifications and comparisons of multilocus recombination distributions

    PubMed Central

    Karlin, Samuel; Liberman, Uri

    1978-01-01

    Various classifications and representations of multilocus recombination structures are delineated based on generalized notions of linkage values and recombination rates. An important class of recombination distributions (called the count-location chiasma process) is parameterized by a distribution of the number of crossover events and, for each such crossover count, by a conditional distribution of crossover locations. A number of properties of this recombination structure are developed. A multilocus definition of a “natural” recombination range is set forth. Orderings among recombination distributions in the multilocus setting are also discussed. Comparisons are made in terms of complete linkage, free assortment and noninterference schemes serving as standards. PMID:16592601

  11. Improving recombinant protein purification yield

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Production of adequate amounts of recombinant proteins is essential for antibody production, biochemical activity study, and structural determination during the post-genomic era. It’s technologically challenging and a limiting factor for tung oil research because analytical reagents such as high qua...

  12. CRMAGE: CRISPR Optimized MAGE Recombineering

    PubMed Central

    Ronda, Carlotta; Pedersen, Lasse Ebdrup; Sommer, Morten O. A.; Nielsen, Alex Toftgaard

    2016-01-01

    A bottleneck in metabolic engineering and systems biology approaches is the lack of efficient genome engineering technologies. Here, we combine CRISPR/Cas9 and λ Red recombineering based MAGE technology (CRMAGE) to create a highly efficient and fast method for genome engineering of Escherichia coli. Using CRMAGE, the recombineering efficiency was between 96.5% and 99.7% for gene recoding of three genomic targets, compared to between 0.68% and 5.4% using traditional recombineering. For modulation of protein synthesis (small insertion/RBS substitution) the efficiency was increased from 6% to 70%. CRMAGE can be multiplexed and enables introduction of at least two mutations in a single round of recombineering with similar efficiencies. PAM-independent loci were targeted using degenerate codons, thereby making it possible to modify any site in the genome. CRMAGE is based on two plasmids that are assembled by a USER-cloning approach enabling quick and cost efficient gRNA replacement. CRMAGE furthermore utilizes CRISPR/Cas9 for efficient plasmid curing, thereby enabling multiple engineering rounds per day. To facilitate the design process, a web-based tool was developed to predict both the λ Red oligos and the gRNAs. The CRMAGE platform enables highly efficient and fast genome editing and may open up promising prospective for automation of genome-scale engineering. PMID:26797514

  13. Inhomogeneous recombinations during cosmic reionization

    NASA Astrophysics Data System (ADS)

    Sobacchi, Emanuele; Mesinger, Andrei

    2014-05-01

    By depleting the ionizing photon budget available to expand cosmic H II regions, recombining systems (or Lyman limit systems) can have a large impact during (and following) cosmic reionization. Unfortunately, directly resolving such structures in large-scale reionization simulations is computationally impractical. Instead, here we implement a subgrid prescription for tracking inhomogeneous recombinations in the intergalactic medium. Building on previous work parametrizing photoheating feedback on star formation, we present large-scale, seminumeric reionization simulations which self-consistently track the local (subgrid) evolution of both sources and sinks of ionizing photons. Our simple, single-parameter model naturally results in both an extended reionization and a modest, slowly evolving emissivity, consistent with observations. Recombinations are instrumental in slowing the growth of large H II regions, and damping the rapid rise of the ionizing background in the late stages of (and following) reionization. As a result, typical H II regions are smaller by factors of ˜2 to 3 throughout reionization. The large-scale (k ≲ 0.2 Mpc-1) ionization power spectrum is suppressed by factors of ≳2-3 in the second half of reionization. Therefore properly modelling recombinations is important in interpreting virtually all reionization observables, including upcoming interferometry with the redshifted 21cm line. Consistent with previous works, we find the clumping factor of ionized gas to be C H II ˜ 4 at the end of reionization.

  14. Recombinant DNA: History of the Controversy.

    ERIC Educational Resources Information Center

    Vigue, Charles L.; Stanziale, William G.

    1979-01-01

    The hazards associated with recombinant DNA research are presented along with some social implications and the development of recombinant DNA research guidelines by the National Institutes of Health. (SA)

  15. Antibacterial activity of recombinant murine beta interferon.

    PubMed Central

    Fujiki, T; Tanaka, A

    1988-01-01

    Recombinant murine beta interferon was protective and therapeutic for mice against Listeria monocytogenes infection in vivo. The recombinant murine beta interferon caused enhanced H2O2 release by macrophages in vivo, but not in vitro. PMID:3343048

  16. [Recombination in Drosophila in space flight].

    PubMed

    Filatova, L P; Vaulina, E N; Lapteva, N Sh; Grozdova, T Ia

    1988-04-01

    An experiment with Drosophila melanogaster males was performed aboard the Artificial Satellite "Kosmos-1667". Mutagenic effects of a 7-day space flight on intergene recombination in chromosome 2 were studied. The space flight factors decreased the frequency of recombination. A model experiment on a laboratory centrifuge demonstrated insignificant increase in recombination frequency caused by acceleration. PMID:3135244

  17. Selection of Recombinant Human Antibodies.

    PubMed

    Tomszak, Florian; Weber, Susanne; Zantow, Jonas; Schirrmann, Thomas; Hust, Michael; Frenzel, André

    2016-01-01

    Since the development of therapeutic antibodies the demand of recombinant human antibodies is steadily increasing. Traditionally, therapeutic antibodies were generated by immunization of rat or mice, the generation of hybridoma clones, cloning of the antibody genes and subsequent humanization and engineering of the lead candidates. In the last few years, techniques were developed that use transgenic animals with a human antibody gene repertoire. Here, modern recombinant DNA technologies can be combined with well established immunization and hybridoma technologies to generate already affinity maturated human antibodies. An alternative are in vitro technologies which enabled the generation of fully human antibodies from antibody gene libraries that even exceed the human antibody repertoire. Specific antibodies can be isolated from these libraries in a very short time and therefore reduce the development time of an antibody drug at a very early stage.In this review, we describe different technologies that are currently used for the in vitro and in vivo generation of human antibodies. PMID:27236551

  18. Chemical kinetics of geminal recombination

    SciTech Connect

    Levin, P.P.; Khudyakov, I.V.; Brin, E.F.; Kuz'min, V.A.

    1988-09-01

    The kinetics of geminal recombination of triplet radical pairs formed in photoreduction of benzophenone by p-cresol in glycerin solution was studied by pulsed laser photolysis. The experiments were conducted at several temperatures and in a constant magnetic field of H = 0.34 T. The parameters in six kinetic equations describing geminal recombination were determined with a computer. The values of the sums of the squares of the residual deviations of the approximation were obtained. It was found that the kinetics are best described by the functions proposed by Noyes and Shushin. It was shown that it is necessary to use the mutual diffusion coefficient of the radicals, which is significantly smaller than the sum of the estimations of the experimental values of the radical diffusion coefficients, for describing the kinetics due to the correlations of the molecular motions of the radicals in the cage.

  19. Recombination Catalysts for Hypersonic Fuels

    NASA Technical Reports Server (NTRS)

    Chinitz, W.

    1998-01-01

    The goal of commercially-viable access to space will require technologies that reduce propulsion system weight and complexity, while extracting maximum energy from the products of combustion. This work is directed toward developing effective nozzle recombination catalysts for the supersonic and hypersonic aeropropulsion engines used to provide such access to space. Effective nozzle recombination will significantly reduce rk=le length (hence, propulsion system weight) and reduce fuel requirements, further decreasing the vehicle's gross lift-off weight. Two such catalysts have been identified in this work, barium and antimony compounds, by developing chemical kinetic reaction mechanisms for these materials and determining the engine performance enhancement for a typical flight trajectory. Significant performance improvements are indicated, using only 2% (mole or mass) of these compounds in the combustor product gas.

  20. Surface recombination statistics at traps

    NASA Astrophysics Data System (ADS)

    Landsberg, P. T.; Abrahams, M. S.

    1983-09-01

    The Shockley-Read-Hall recombination statistics was recently generalised by Dhariwal, Kothari and Jain to include the effect of a finite time of relaxation before the captured carrier settles into its ground state, and by Landsberg to allow for Auger effects and so-called "extra" carriers supplied to the semiconductor from the outside. The combined result of these effects is studied here theoretically, together with the consideration of a simple distribution of trap states. It is found that the surface recombination velocity s has the usual minimum in the near intrinsic state and that s passes through a maximum as a function of excess electron concentration. Both extrema are enhanced if the trap states are distributed over an energy range. Experimental plots of s as a function of excess electron and hole concentrations should yield insight concerning the numerical importance of (a) Auger effects with the participation of traps and (b) relaxation times.

  1. Recombinant Toxins for Cancer Treatment

    NASA Astrophysics Data System (ADS)

    Pastan, Ira; Fitzgerald, David

    1991-11-01

    Recombinant toxins target cell surface receptors and antigens on tumor cells. They kill by mechanisms different from conventional chemotherapy, so that cross resistance to conventional chemotherapeutic agents should not be a problem. Furthermore, they are not mutagens and should not induce secondary malignancies or accelerate progression of benign malignancies. They can be mass-produced cheaply in bacteria as homogeneous proteins. Either growth factor-toxin fusions or antibody-toxin fusions can be chosen, depending on the cellular target.

  2. Introductory experiments in recombinant DNA.

    PubMed

    Tait, R C

    2000-07-01

    Nine practical exercises demonstrate the basic principles in recombinant DNA. The exercises explain the principles that DNA equals genes and that changes in DNA cause changes in genetic properties. The aim is to provide a teaching resource that can be used to illustrate the theory and applications of molecular biology to highschool students, undergraduate students, medics, dentists, doctors, nurses, life scientists, and anyone learning the basics of DNA technology. PMID:11471559

  3. Recombinant vector and eukaryotic host transformed thereby

    SciTech Connect

    Sugden, W.M.

    1987-08-11

    A recombinant plasmid is described comprising: a segment from a first plasmid which is not a lymphotrophic herpes virus segment and which facilitates the replication of the recombinant plasmid in a prokaryotic host; a segment from a lymphotrophic herpes virus which is linked to the first plasmid segment such that is a capable of assisting in maintaining the recombinant plasmid as a plasmid if the recombinant plasmid is inserted into a eukaryotic host that has been transformed by the lymphotrophic herpes virus; and a foreign eukaryotic gene component linked as part of the recombinant plasmid.

  4. A coalescent model of recombination hotspots.

    PubMed Central

    Wiuf, Carsten; Posada, David

    2003-01-01

    Recent experimental findings suggest that the assumption of a homogeneous recombination rate along the human genome is too naive. These findings point to block-structured recombination rates; certain regions (called hotspots) are more prone than other regions to recombination. In this report a coalescent model incorporating hotspot or block-structured recombination is developed and investigated analytically as well as by simulation. Our main results can be summarized as follows: (1) The expected number of recombination events is much lower in a model with pure hotspot recombination than in a model with pure homogeneous recombination, (2) hotspots give rise to large variation in recombination rates along the genome as well as in the number of historical recombination events, and (3) the size of a (nonrecombining) block in the hotspot model is likely to be overestimated grossly when estimated from SNP data. The results are discussed with reference to the current debate about block-structured recombination and, in addition, the results are compared to genome-wide variation in recombination rates. A number of new analytical results about the model are derived. PMID:12750351

  5. Nondisjunction of chromosome 15: Origin and recombination

    SciTech Connect

    Robinson, W.P.; Bernasconi, F.; Schinzel, A.A.; Mutirangura, A.; Ledbetter, D.H. ); Langlois, S. ); Morris, M.A.; Malcolm, S.

    1993-09-01

    Thirty-two cases of uniparental disomy (UPD), ascertained from Prader-Willi syndrome patients (N=27) and Angelman syndrome patients (N-5), are used to investigate the pattern of recombination associated with nondisjunction of chromosome 15. In addition, the meiotic stage of nondisjunction is inferred by using markers mapping near the centromere. Two basic approaches to the analysis of recombination in specific pairwise intervals along the chromosome. This method shows a significant reduction in recombination for two of five intervals examined. Second, the observed frequency of each recombinant class (i.e., zero, one, two, three, or more observable crossovers) is compared with expected values. This is useful for testing whether the reduction in recombination can be attributed solely to a proportion of cases with no recombination at all (because of asynapsis), with the remaining groups showing normal recombination (or even excess recombination), or whether recombination is uniformly reduced. Analysis of maternal UPD(15) data shows a slight reduction in the multiple-recombinant classes, with a corresponding increase in both the zero- and one-recombinant classes over expected values. The majority, more than 82%, of the extra chromosomes in maternal UPD(15) cases are due to meiotic I nondisjunction events. In contrast, more paternal UPD(15) cases so far examined appear to have a postzygotic origin of the extra paternal chromosome. 33 refs., 1 fig., 7 tabs.

  6. Effect of gamma radiation on retroviral recombination.

    PubMed

    Hu, W S; Temin, H M

    1992-07-01

    To elucidate the mechanism(s) of retroviral recombination, we exposed virions to gamma radiation prior to infecting target cells. By using previously described spleen necrosis virus-based vectors containing multiple markers, recombinant proviruses were studied after a single round of retrovirus replication. The current models of retroviral recombination predict that breaking virion RNA should promote minus-strand recombination (forced copy-choice model), decrease or not affect plus-strand recombination (strand displacement/assimilation model), and shift plus-strand recombination towards the 3' end of the genome. However, we found that while gamma irradiation of virions reduced the amount of recoverable viral RNA, it did not primarily cause breaks. Thus, the frequency of selected recombinants was not significantly altered with greater doses of radiation. In spite of this, the irradiation did decrease the number of recombinants with only one internal template switch. As a result, the average number of additional internal template switches in the recombinant proviruses increased from 0.7 to 1.4 as infectivity decreased to 6%. The unselected internal template switches tended to be 5' of the selected crossover even in the recombinants from irradiated viruses, inconsistent with a plus-strand recombination mechanism. PMID:1602553

  7. Bacteriophage recombination systems and biotechnical applications.

    PubMed

    Nafissi, Nafiseh; Slavcev, Roderick

    2014-04-01

    Bacteriophage recombination systems have been widely used in biotechnology for modifying prokaryotic species, for creating transgenic animals and plants, and more recently, for human cell gene manipulation. In contrast to homologous recombination, which benefits from the endogenous recombination machinery of the cell, site-specific recombination requires an exogenous source of recombinase in mammalian cells. The mechanism of bacteriophage evolution and their coexistence with bacterial cells has become a point of interest ever since bacterial viruses' life cycles were first explored. Phage recombinases have already been exploited as valuable genetic tools and new phage enzymes, and their potential application to genetic engineering and genome manipulation, vectorology, and generation of new transgene delivery vectors, and cell therapy are attractive areas of research that continue to be investigated. The significance and role of phage recombination systems in biotechnology is reviewed in this paper, with specific focus on homologous and site-specific recombination conferred by the coli phages, λ, and N15, the integrase from the Streptomyces phage, ΦC31, the recombination system of phage P1, and the recently characterized recombination functions of Yersinia phage, PY54. Key steps of the molecular mechanisms involving phage recombination functions and their application to molecular engineering, our novel exploitations of the PY54-derived recombination system, and its application to the development of new DNA vectors are discussed. PMID:24442504

  8. Recombinant DNA technology in apple.

    PubMed

    Gessler, Cesare; Patocchi, Andrea

    2007-01-01

    This review summarizes the achievements of almost 20 years of recombinant DNA technology applied to apple, grouping the research results into the sections: developing the technology, insect resistance, fungal disease resistance, self-incompatibility, herbicide resistance, fire blight resistance, fruit ripening, allergens, rooting ability, and acceptance and risk assessment. The diseases fire blight, caused by Erwinia amylovora, and scab, caused by Venturia inaequalis, were and still are the prime targets. Shelf life improvement and rooting ability of rootstocks are also relevant research areas. The tools to create genetically modified apples of added value to producers, consumers, and the environment are now available. PMID:17522823

  9. Recombinant erythropoietin in clinical practice

    PubMed Central

    Ng, T; Marx, G; Littlewood, T; Macdougall, I

    2003-01-01

    The introduction of recombinant human erythropoietin (RHuEPO) has revolutionised the treatment of patients with anaemia of chronic renal disease. Clinical studies have demonstrated that RHuEPO is also useful in various non-uraemic conditions including haematological and oncological disorders, prematurity, HIV infection, and perioperative therapies. Besides highlighting both the historical and functional aspects of RHuEPO, this review discusses the applications of RHuEPO in clinical practice and the potential problems of RHuEPO treatment. PMID:12897214

  10. Annealing Vs. Invasion in Phage λ Recombination

    PubMed Central

    Stahl, M. M.; Thomason, L.; Poteete, A. R.; Tarkowski, T.; Kuzminov, A.; Stahl, F. W.

    1997-01-01

    Genetic recombination catalyzed by λ's Red pathway was studied in rec(+) and recA mutant bacteria by examining both intracellular λ DNA and mature progeny particles. Recombination of nonreplicating phage chromosomes was induced by double-strand breaks delivered at unique sites in vivo. In rec(+) cells, cutting only one chromosome gave nearly maximal stimulation of recombination; the recombinants formed contained relatively short hybrid regions, suggesting strand invasion. In contrast, in recA mutant cells, cutting the two parental chromosomes at non-allelic sites was required for maximal stimulation; the recombinants formed tended to be hybrid over the entire region between the two cuts, implying strand annealing. We conclude that, in the absence of RecA and the presence of non-allelic DNA ends, the Red pathway of λ catalyzes recombination primarily by annealing. PMID:9383045

  11. Recombinant DNA production of spider silk proteins

    PubMed Central

    Tokareva, Olena; Michalczechen-Lacerda, Valquíria A; Rech, Elíbio L; Kaplan, David L

    2013-01-01

    Spider dragline silk is considered to be the toughest biopolymer on Earth due to an extraordinary combination of strength and elasticity. Moreover, silks are biocompatible and biodegradable protein-based materials. Recent advances in genetic engineering make it possible to produce recombinant silks in heterologous hosts, opening up opportunities for large-scale production of recombinant silks for various biomedical and material science applications. We review the current strategies to produce recombinant spider silks. PMID:24119078

  12. Recombinant bacteria for mosquito control.

    PubMed

    Federici, B A; Park, H-W; Bideshi, D K; Wirth, M C; Johnson, J J

    2003-11-01

    Bacterial insecticides have been used for the control of nuisance and vector mosquitoes for more than two decades. Nevertheless, due primarily to their high cost and often only moderate efficacy, these insecticides remain of limited use in tropical countries where mosquito-borne diseases are prevalent. Recently, however, recombinant DNA techniques have been used to improve bacterial insecticide efficacy by markedly increasing the synthesis of mosquitocidal proteins and by enabling new endotoxin combinations from different bacteria to be produced within single strains. These new strains combine mosquitocidal Cry and Cyt proteins of Bacillus thuringiensis with the binary toxin of Bacillus sphaericus, improving efficacy against Culex species by 10-fold and greatly reducing the potential for resistance through the presence of Cyt1A. Moreover, although intensive use of B. sphaericus against Culex populations in the field can result in high levels of resistance, most of this can be suppressed by combining this bacterial species with Cyt1A; the latter enables the binary toxin of this species to enter midgut epithelial cells via the microvillar membrane in the absence of a midgut receptor. The availability of these novel strains and newly discovered mosquitocidal proteins, such as the Mtx toxins of B. sphaericus, offers the potential for constructing a range of recombinant bacterial insecticides for more effective control of the mosquito vectors of filariasis, Dengue fever and malaria. PMID:14506223

  13. Dissociative recombination in planetary ionospheres

    NASA Technical Reports Server (NTRS)

    Fox, J. L.

    1993-01-01

    Ionization in planetary atmospheres can be produced by solar photoionization, photoelectron impact ionization, and, in auroral regions, by impact of precipitating particles. This ionization is lost mainly in dissociative recombination (DR) of molecular ions. Although atomic ions cannot undergo DR, they can be transformed locally through ion-molecule reactions into molecular ions, or they may be transported vertically or horizontally to regions of the atmosphere where such transformations are possible. Because DR reactions tend to be very exothermic, they can be an important source of kinetically or internally excited fragments. In interplanetary thermospheres, the neutral densities decrease exponentially with altitude. Below the homopause (or turbopause), the atmosphere is assumed to be throughly mixed by convection and/or turbulence. Above the homopause, diffusion is the major transport mechanism, and each species is distributed according to its mass, with the logarithmic derivative of the density with repect to altitude given approximately by -1/H, where H = kT/mg is the scale height. In this expression, T is the neutral temperature, g is the local acceleratiion of gravity, and m is the mass of the species. Thus lighter species become relatively more abundant, and heavier species less abundant, as the altitude increases. This variation of the neutral composition can lead to changes in the ion composition; furthermore, as the neutral densities decrease, dissociative recombination becomes more important relative to ion-neutral reactions as a loss mechanism for molecular ions.

  14. Human Insulin from Recombinant DNA Technology

    NASA Astrophysics Data System (ADS)

    Johnson, Irving S.

    1983-02-01

    Human insulin produced by recombinant DNA technology is the first commercial health care product derived from this technology. Work on this product was initiated before there were federal guidelines for large-scale recombinant DNA work or commercial development of recombinant DNA products. The steps taken to facilitate acceptance of large-scale work and proof of the identity and safety of such a product are described. While basic studies in recombinant DNA technology will continue to have a profound impact on research in the life sciences, commercial applications may well be controlled by economic conditions and the availability of investment capital.

  15. Dielectronic recombination of tungsten ions

    NASA Astrophysics Data System (ADS)

    Li, Bowen; O’Sullivan, Gerry; Dong, Chenzhong; Chen, Ximeng

    2016-08-01

    Ab initio calculations of dielectronic recombination rate coefficients of Ne-, Pd- and Ag-like tungsten have been performed. Energy levels, radiative transition probabilities and autoionization rates were calculated using the Flexible Atomic Code. The contributions from different channels to the total rate coefficients are discussed. The present calculated rate coefficients are compared with other calculations where available. Excellent agreement has been found for Ne-like W while a large discrepancy was found for Pd-like W, which implies that more ab initio calculations and experimental measurements are badly needed. Further calculations demonstrated that the influence of configuration interaction is small while nonresonant radiative stabilizing (NRS) contribution to doubly excited non-autoionizing states are vital. The data obtained are expected to be useful for modeling plasmas for fusion applications, especially for the ITER community, which makes experimental verification even more essential.

  16. Steric effects on geminate recombinations

    SciTech Connect

    Traylor, T.G.; Taube, D.; Jongeward, K.A.; Magde, D. )

    1990-09-12

    Steric effects on the binding of isonitrile ligands to iron(II) porphyrins were investigated by picosecond flash photolysis. Two different types of steric effects were distinguished and characterized: (1) steric restrictions to porphyrin planarity and (2) blocking of the pathway for ligand approach. Heme planarity was restricted by coordinating 1,2-dimethylimidazole trans to the ligand binding site being investigated. Blocking of the binding site was explored by using adamantane heme 6,6-cyclophane, in which the adamantane moiety forms a cap over the binding site. Results of picosecond kinetic measurements demonstrate that the first effect, heme nonplanarity or trans strain, influences the bond-making step, whereas the second effect, ligand blocking, involves a conformational preequilibrium prior to bond making. Relevance of these findings for contact pair recombination, in general, and for heme protein ligation, in particular, are discussed.

  17. Modeling Interference in Genetic Recombination

    PubMed Central

    McPeek, M. S.; Speed, T. P.

    1995-01-01

    In analyzing genetic linkage data it is common to assume that the locations of crossovers along a chromosome follow a Poisson process, whereas it has long been known that this assumption does not fit the data. In many organisms it appears that the presence of a crossover inhibits the formation of another nearby, a phenomenon known as ``interference.'' We discuss several point process models for recombination that incorporate position interference but assume no chromatid interference. Using stochastic simulation, we are able to fit the models to a multilocus Drosophila dataset by the method of maximum likelihood. We find that some biologically inspired point process models incorporating one or two additional parameters provide a dramatically better fit to the data than the usual ``no-interference'' Poisson model. PMID:7713406

  18. Deleterious background selection with recombination

    SciTech Connect

    Hudson, R.R.; Kaplan, N.L.

    1995-12-01

    An analytic expression for the expected nucleotide diversity is obtained for a neutral locus in a region with deleterious mutation and recombination. Our analytic results are used to predict levels of variation for the entire third chromosome of Drosophila melanogaster. The predictions are consistent with the low levels of variation that have been observed at loci near the centromeres of the third chromosome of D. melanogaster. However, the low levels of variation observed near the tips of this chromosome are not predicted using currently available estimates of the deleterious mutation rate and of selection coefficients. If considerably smaller selection coefficients are assumed, the low observed levels of variation at the tips of the third chromosome are consistent with the background selection model. 33 refs., 4 figs., 1 tab.

  19. Recombinant prolylcarboxypeptidase activates plasma prekallikrein.

    PubMed

    Shariat-Madar, Zia; Mahdi, Fakhri; Schmaier, Alvin H

    2004-06-15

    The serine protease prolylcarboxypeptidase (PRCP), isolated from human umbilical vein endothelial cells (HUVECs), is a plasma prekallikrein (PK) activator. PRCP cDNA was cloned in pMT/BIP/V5-HIS-C, transfected into Schneider insect (S2) cells, and purified from serum-free media. Full-length recombinant PRCP (rPRCP) activates PK when bound to high-molecular-weight kininogen (HK). Recombinant PRCP is inhibited by leupeptin, angiotensin II, bradykinin, anti-PRCP, diisopropyl-fluorophosphonate (DFP), phenylmethylsulfonyl fluoride (PMSF), and Z-Pro-Proaldehyde-dimethyl acetate, but not by 1 mM EDTA (ethylenediaminetetraacetic acid), bradykinin 1-5, or angiotensin 1-7. Corn trypsin inhibitor binds to prekallikrein to prevent rPRCP activation, but it does not directly inhibit the active site of either enzyme. Unlike factor XIIa, the ability of rPRCP to activate PK is blocked by angiotensin II, not by neutralizing antibody to factor XIIa. PRCP antigen is detected on HUVEC membranes using flow cytometry and laser scanning confocal microscopy. PRCP antigen does not colocalize with LAMP1 on nonpermeabilized HUVECs, but it partially colocalizes in permeabilized cells. PRCP colocalizes with all the HK receptors, gC1qR, uPAR, and cytokeratin 1 antigen, on nonpermeabilized HUVECs. PRCP activity and antigen expression on cultured HUVECs are blocked by a morpholino antisense oligonucleotide. These investigations indicate that rPRCP is functionally identical to isolated HUVEC PRCP and is a major HUVEC membrane-expressed, PK-activating enzyme detected in the intravascular compartment. PMID:14996700

  20. Fundamental Studies of Recombinant Hydrogenases

    SciTech Connect

    Adams, Michael W

    2014-01-25

    This research addressed the long term goals of understanding the assembly and organization of hydrogenase enzymes, of reducing them in size and complexity, of determining structure/function relationships, including energy conservation via charge separation across membranes, and in screening for novel H2 catalysts. A key overall goal of the proposed research was to define and characterize minimal hydrogenases that are produced in high yields and are oxygen-resistant. Remarkably, in spite of decades of research carried out on hydrogenases, it is not possible to readily manipulate or design the enzyme using molecular biology approaches since a recombinant form produced in a suitable host is not available. Such resources are essential if we are to understand what constitutes a “minimal” hydrogenase and design such catalysts with certain properties, such as resistance to oxygen, extreme stability and specificity for a given electron donor. The model system for our studies is Pyrococcus furiosus, a hyperthermophile that grows optimally at 100°C, which contains three different nickel-iron [NiFe-] containing hydrogenases. Hydrogenases I and II are cytoplasmic while the other, MBH, is an integral membrane protein that functions to both evolve H2 and pump protons. Three important breakthroughs were made during the funding period with P. furiosus soluble hydrogenase I (SHI). First, we produced an active recombinant form of SHI in E. coli by the co-expression of sixteen genes using anaerobically-induced promoters. Second, we genetically-engineered P. furiosus to overexpress SHI by an order of magnitude compared to the wild type strain. Third, we generated the first ‘minimal’ form of SHI, one that contained two rather than four subunits. This dimeric form was stable and active, and directly interacted with a pyruvate-oxidizing enzyme with any intermediate electron carrier. The research resulted in five peer-reviewed publications.

  1. RNA recombination in a coronavirus: recombination between viral genomic RNA and transfected RNA fragments.

    PubMed Central

    Liao, C L; Lai, M M

    1992-01-01

    Mouse hepatitis virus (MHV), a coronavirus, has been shown to undergo a high frequency of RNA recombination both in tissue culture and in animal infection. So far, RNA recombination has been demonstrated only between genomic RNAs of two coinfecting viruses. To understand the mechanism of RNA recombination and to further explore the potential of RNA recombination, we studied whether recombination could occur between a replicating MHV RNA and transfected RNA fragments. We first used RNA fragments which represented the 5' end of genomic-sense sequences of MHV RNA for transfection. By using polymerase chain reaction amplification with two specific primers, we were able to detect recombinant RNAs which incorporated the transfected fragment into the 5' end of the viral RNA in the infected cells. Surprisingly, even the anti-genomic-sense RNA fragments complementary to the 5' end of MHV genomic RNA could also recombine with the MHV genomic RNAs. This observation suggests that RNA recombination can occur during both positive- and negative-strand RNA synthesis. Furthermore, the recombinant RNAs could be detected in the virion released from the infected cells even after several passages of virus in tissue culture cells, indicating that these recombinant RNAs represented functional virion RNAs. The crossover sites of these recombinants were detected throughout the transfected RNA fragments. However, when an RNA fragment with a nine-nucleotide (CUUUAUAAA) deletion immediately downstream of a pentanucleotide (UCUAA) repeat sequence in the leader RNA was transfected into MHV-infected cells, most of the recombinants between this RNA and the MHV genome contained crossover sites near this pentanucleotide repeat sequence. In contrast, when exogenous RNAs with the intact nine-nucleotide sequence were used in similar experiments, the crossover sites of recombinants in viral genomic RNA could be detected at more-downstream sites. This study demonstrated that recombination can occur

  2. Recombining without Hotspots: A Comprehensive Evolutionary Portrait of Recombination in Two Closely Related Species of Drosophila

    PubMed Central

    Smukowski Heil, Caiti S.; Ellison, Chris; Dubin, Matthew; Noor, Mohamed A.F.

    2015-01-01

    Meiotic recombination rate varies across the genome within and between individuals, populations, and species in virtually all taxa studied. In almost every species, this variation takes the form of discrete recombination hotspots, determined in some mammals by a protein called PRDM9. Hotspots and their determinants have a profound effect on the genomic landscape, and share certain features that extend across the tree of life. Drosophila, in contrast, are anomalous in their absence of hotspots, PRDM9, and other species-specific differences in the determination of recombination. To better understand the evolution of meiosis and general patterns of recombination across diverse taxa, we present a truly comprehensive portrait of recombination across time, combining recently published cross-based contemporary recombination estimates from each of two sister species with newly obtained linkage-disequilibrium-based historic estimates of recombination from both of these species. Using Drosophila pseudoobscura and Drosophila miranda as a model system, we compare recombination rate between species at multiple scales, and we suggest that Drosophila replicate the pattern seen in human–chimpanzee in which recombination rate is conserved at broad scales. We also find evidence of a species-wide recombination modifier(s), resulting in both a present and historic genome-wide elevation of recombination rates in D. miranda, and identify broad scale effects on recombination from the presence of an inversion. Finally, we reveal an unprecedented view of the distribution of recombination in D. pseudoobscura, illustrating patterns of linked selection and where recombination is taking place. Overall, by combining these estimation approaches, we highlight key similarities and differences in recombination between Drosophila and other organisms. PMID:26430062

  3. Oligo Recombination in Gram Negative Bacteria

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Homologous recombination is important for bacterial survival because it simultaneously provides genomic stability as well as genomic plasticity. Of the mechanistic pathways for homologous recombination, those mediated by RecA are the most thoroughly characterized and are understood to be structural...

  4. Dissociative recombination of highly symmetric polyatomic ions.

    PubMed

    Douguet, Nicolas; Orel, Ann E; Greene, Chris H; Kokoouline, Viatcheslav

    2012-01-13

    A general first-principles theory of dissociative recombination is developed for highly symmetric molecular ions and applied to H(3)O(+) and CH(3)(+), which play an important role in astrophysical, combustion, and laboratory plasma environments. The theoretical cross sections obtained for the dissociative recombination of the two ions are in good agreement with existing experimental data from storage ring experiments. PMID:22324682

  5. High efficiency recombineering in lactic acid bacteria

    PubMed Central

    van Pijkeren, Jan-Peter; Britton, Robert A.

    2012-01-01

    The ability to efficiently generate targeted point mutations in the chromosome without the need for antibiotics, or other means of selection, is a powerful strategy for genome engineering. Although oligonucleotide-mediated recombineering (ssDNA recombineering) has been utilized in Escherichia coli for over a decade, the successful adaptation of ssDNA recombineering to Gram-positive bacteria has not been reported. Here we describe the development and application of ssDNA recombineering in lactic acid bacteria. Mutations were incorporated in the chromosome of Lactobacillus reuteri and Lactococcus lactis without selection at frequencies ranging between 0.4% and 19%. Whole genome sequence analysis showed that ssDNA recombineering is specific and not hypermutagenic. To highlight the utility of ssDNA recombineering we reduced the intrinsic vancomymycin resistance of L. reuteri >100-fold. By creating a single amino acid change in the d-Ala-d-Ala ligase enzyme we reduced the minimum inhibitory concentration for vancomycin from >256 to 1.5 µg/ml, well below the clinically relevant minimum inhibitory concentration. Recombineering thus allows high efficiency mutagenesis in lactobacilli and lactococci, and may be used to further enhance beneficial properties and safety of strains used in medicine and industry. We expect that this work will serve as a blueprint for the adaptation of ssDNA recombineering to other Gram-positive bacteria. PMID:22328729

  6. Oligonucleotide recombination in gram negative bacteria

    Technology Transfer Automated Retrieval System (TEKTRAN)

    This report describes several key aspects of a novel form of RecA-independent homologous recombination. We found that synthetic single stranded DNA oligonucleotides (oligos) introduced into bacteria by transformation can site-specifically recombine with bacterial chromosomes in the absence of any a...

  7. Dielectronic recombination lines of C{sup +}

    SciTech Connect

    Sochi, Taha Storey, Peter J.

    2013-11-15

    The present paper presents atomic data generated to investigate the recombination lines of C II in the spectra of planetary nebulae. These data include energies of bound and autoionizing states, oscillator strengths and radiative transition probabilities, autoionization probabilities, and recombination coefficients. The R-matrix method of electron scattering theory was used to describe the C{sup 2+} plus electron system.

  8. Recombinant Vaccinia Virus: Immunization against Multiple Pathogens

    NASA Astrophysics Data System (ADS)

    Perkus, Marion E.; Piccini, Antonia; Lipinskas, Bernard R.; Paoletti, Enzo

    1985-09-01

    The coding sequences for the hepatitis B virus surface antigen, the herpes simplex virus glycoprotein D, and the influenza virus hemagglutinin were inserted into a single vaccinia virus genome. Rabbits inoculated intravenously or intradermally with this polyvalent vaccinia virus recombinant produced antibodies reactive to all three authentic foreign antigens. In addition, the feasibility of multiple rounds of vaccination with recombinant vaccinia virus was demonstrated.

  9. Recombinant Swinepox Virus for Veterinary Vaccine Development.

    PubMed

    Fan, Hong-Jie; Lin, Hui-Xing

    2016-01-01

    Poxvirus-vectors have been widely used in vaccine development for several important human and animal diseases; some of these vaccines have been licensed and used extensively. Swinepox virus (SPV) is well suited to develop recombinant vaccines because of its large packaging capacity for recombinant DNA, its host range specificity, and its ability to induce appropriate immune responses. PMID:26458836

  10. Recombinant DNA encoding a desulfurization biocatalyst

    DOEpatents

    Rambosek, J.; Piddington, C.S.; Kovacevich, B.R.; Young, K.D.; Denome, S.A.

    1994-10-18

    This invention relates to a recombinant DNA molecule containing a gene or genes which encode a biocatalyst capable of desulfurizing a fossil fuel which contains organic sulfur molecules. For example, the present invention encompasses a recombinant DNA molecule containing a gene or genes of a strain of Rhodococcus rhodochrous. 13 figs.

  11. Recombinant DNA encoding a desulfurization biocatalyst

    DOEpatents

    Rambosek, John; Piddington, Chris S.; Kovacevich, Brian R.; Young, Kevin D.; Denome, Sylvia A.

    1994-01-01

    This invention relates to a recombinant DNA molecule containing a gene or genes which encode a biocatalyst capable of desulfurizing a fossil fuel which contains organic sulfur molecules. For example, the present invention encompasses a recombinant DNA molecule containing a gene or genes of a strain of Rhodococcus rhodochrous.

  12. V(D)J recombination deficiencies.

    PubMed

    de Villartay, Jean-Pierre

    2009-01-01

    V(D)J recombination not only comprises the molecular mechanism that insures diversity of the immune system but also constitutes a critical checkpoint in the developmental program of B- and T-lymphocytes. The analysis of human patients with Severe Combined Immune Deficiency (SCID) has contributed to the understanding of the biochemistry of the V(D)J recombination reaction. The molecular study V(D)J recombination settings in humans, mice and in cellular mutants has allowed to unravel the process of Non Homologous End Joining (NHEJ), one of the key pathway that insure proper repair of DNA double strand breaks (dsb), whether they occur during V(D)J recombination or secondary to other DNA injuries. Two NHEJ factors, Artemis and Cernunnos, were indeed discovered through the study of human V(D)J recombination defective human SCID patients. PMID:19731800

  13. Biochemistry of Meiotic Recombination: Formation, Processing, and Resolution of Recombination Intermediates

    PubMed Central

    Ehmsen, Kirk T.

    2009-01-01

    Meiotic recombination ensures accurate chromosome segregation during the first meiotic division and provides a mechanism to increase genetic heterogeneity among the meiotic products. Unlike homologous recombination in somatic (vegetative) cells, where sister chromatid interactions prevail and crossover formation is avoided, meiotic recombination is targeted to involve homologs, resulting in crossovers to connect the homologs before anaphase of the first meiotic division. The mechanisms responsible for homolog choice and crossover control are poorly understood, but likely involve meiosis-specific recombination proteins, as well as meiosis-specific chromosome organization and architecture. Much progress has been made to identify and biochemically characterize many of the proteins acting during meiotic recombination. This review will focus on the proteins that generate and process heteroduplex DNA, as well as those that process DNA junctions during meiotic recombination, with particular attention to how recombination activities promote crossover resolution between homologs. PMID:20098639

  14. GENERATION OF RECOMBINANT BACULOVIRUS VIA LIPOSOME MEDIATED TRANSFECTION

    EPA Science Inventory

    Baculovirus expression vectors have become a popular method of producing recombinant proteins. Production of recombinant virus requires the transfection of both the native viral DNA and a transfer plasmid into insect cells where recombination takes place. While several methods of...

  15. Charge recombination and thermoluminescence in photosystem II.

    PubMed

    Rappaport, Fabrice; Cuni, Aude; Xiong, Ling; Sayre, Richard; Lavergne, Jérôme

    2005-03-01

    In the recombination process of Photosystem II (S(2)Q(A)(-)-->S(1)Q(A)) the limiting step is the electron transfer from the reduced primary acceptor pheophytin Ph(-) to the oxidized primary donor P(+) and the rate depends on the equilibrium constant between states S(2)PPhQ(A)(-) and S(1)P(+)Ph(-)Q(A). Accordingly, mutations that affect the midpoint potential of Ph or of P result in a modified recombination rate. A strong correlation is observed between the effects on the recombination rate and on thermoluminescence (TL, the light emission from S(2)Q(A)(-) during a warming ramp): a slower recombination corresponds to a large enhancement and higher temperature of the TL peak. The current theory of TL does not account for these effects, because it is based on the assumption that the rate-limiting step coincides with the radiative process. When implementing the known fact that the radiative pathway represents a minor leak, the modified TL theory readily accounts qualitatively for the observed behavior. However, the peak temperature is still lower than predicted from the temperature-dependence of recombination. We argue that this reflects the heterogeneity of the recombination process combined with the enhanced sensitivity of TL to slower components. The recombination kinetics are accurately fitted as a sum of two exponentials and we show that this is not due to a progressive stabilization of the charge-separated state, but to a pre-existing conformational heterogeneity. PMID:15653722

  16. Electron Recombination in a Dense Hydrogen Plasma

    SciTech Connect

    Jana, M.R.; Johnstone, C.; Kobilarcik, T.; Koizumi, G.M.; Moretti, A.; Popovic, M.; Tollestrup, A.V.; Yonehara, K.; Leonova, M.A.; Schwarz, T.A.; Chung, M.; /Unlisted /IIT, Chicago /Fermilab /MUONS Inc., Batavia /Turin Polytechnic

    2012-05-01

    A high pressure hydrogen gas filled RF cavity was subjected to an intense proton beam to study the evolution of the beam induced plasma inside the cavity. Varying beam intensities, gas pressures and electric fields were tested. Beam induced ionized electrons load the cavity, thereby decreasing the accelerating gradient. The extent and duration of this degradation has been measured. A model of the recombination between ionized electrons and ions is presented, with the intent of producing a baseline for the physics inside such a cavity used in a muon accelerator. Analysis of the data taken during the summer of 2011 shows that self recombination takes place in pure hydrogen gas. The decay of the number of electrons in the cavity once the beam is turned off indicates self recombination rather than attachment to electronegative dopants or impurities. The cross section of electron recombination grows for larger clusters of hydrogen and so at the equilibrium of electron production and recombination in the cavity, processes involving H{sub 5}{sup +} or larger clusters must be taking place. The measured recombination rates during this time match or exceed the analytic predicted values. The accelerating gradient in the cavity recovers fully in time for the next beam pulse of a muon collider. Exactly what the recombination rate is and how much the gradient degrades during the 60 ns muon collider beam pulse will be extrapolated from data taken during the spring of 2012.

  17. Intraspecific variation of recombination rate in maize

    PubMed Central

    2013-01-01

    Background In sexually reproducing organisms, meiotic crossovers ensure the proper segregation of chromosomes and contribute to genetic diversity by shuffling allelic combinations. Such genetic reassortment is exploited in breeding to combine favorable alleles, and in genetic research to identify genetic factors underlying traits of interest via linkage or association-based approaches. Crossover numbers and distributions along chromosomes vary between species, but little is known about their intraspecies variation. Results Here, we report on the variation of recombination rates between 22 European maize inbred lines that belong to the Dent and Flint gene pools. We genotype 23 doubled-haploid populations derived from crosses between these lines with a 50 k-SNP array and construct high-density genetic maps, showing good correspondence with the maize B73 genome sequence assembly. By aligning each genetic map to the B73 sequence, we obtain the recombination rates along chromosomes specific to each population. We identify significant differences in recombination rates at the genome-wide, chromosome, and intrachromosomal levels between populations, as well as significant variation for genome-wide recombination rates among maize lines. Crossover interference analysis using a two-pathway modeling framework reveals a negative association between recombination rate and interference strength. Conclusions To our knowledge, the present work provides the most comprehensive study on intraspecific variation of recombination rates and crossover interference strength in eukaryotes. Differences found in recombination rates will allow for selection of high or low recombining lines in crossing programs. Our methodology should pave the way for precise identification of genes controlling recombination rates in maize and other organisms. PMID:24050704

  18. Advances in recombinant antibody manufacturing.

    PubMed

    Kunert, Renate; Reinhart, David

    2016-04-01

    Since the first use of Chinese hamster ovary (CHO) cells for recombinant protein expression, production processes have steadily improved through numerous advances. In this review, we have highlighted several key milestones that have contributed to the success of CHO cells from the beginning of their use for monoclonal antibody (mAb) expression until today. The main factors influencing the yield of a production process are the time to accumulate a desired amount of biomass, the process duration, and the specific productivity. By comparing maximum cell densities and specific growth rates of various expression systems, we have emphasized the limiting parameters of different cellular systems and comprehensively described scientific approaches and techniques to improve host cell lines. Besides the quantitative evaluation of current systems, the quality-determining properties of a host cell line, namely post-translational modifications, were analyzed and compared to naturally occurring polyclonal immunoglobulin fractions from human plasma. In summary, numerous different expression systems for mAbs are available and also under scientific investigation. However, CHO cells are the most frequently investigated cell lines and remain the workhorse for mAb production until today. PMID:26936774

  19. Recombinant protein production and streptomycetes.

    PubMed

    Anné, Jozef; Maldonado, Bárbara; Van Impe, Jan; Van Mellaert, Lieve; Bernaerts, Kristel

    2012-04-30

    The biopharmaceutical market has come a long way since 1982, when the first biopharmaceutical product, recombinant human insulin, was launched. Just over 200 biopharma products have already gained approval. The global market for biopharmaceuticals which is currently valued at over US$99 billion has been growing at an impressive compound annual growth rate over the previous years. To produce these biopharmaceuticals and other industrially important heterologous proteins, different prokaryotic and eukaryotic expression systems are used. All expression systems have some advantages as well as some disadvantages that should be considered in selecting which one to use. Choosing the best one requires evaluating the options--from yield to glycosylation, to proper folding, to economics of scale-up. No host cell from which all the proteins can be universally expressed in large quantities has been found so far. Therefore, it is important to provide a variety of host-vector expression systems in order to increase the opportunities to screen for the most suitable expression conditions or host cell. In this overview, we focus on Streptomyces lividans, a Gram-positive bacterium with a proven excellence in secretion capacity, as host for heterologous protein production. We will discuss its advantages and disadvantages, and how with systems biology approaches strains can be developed to better producing cell factories. PMID:21777629

  20. Dissociative Recombination without a Curve Crossing

    NASA Technical Reports Server (NTRS)

    Guberman, Steven L.

    1994-01-01

    Ab initio calculations show that a curve crossing is not always needed for a high dissociative- recombination cross section. For HeH(+), in which no neutral states cross the ion potential curve, dissociative recombination is driven by the nuclear kinetic-energy operator on adiabatic potential curves. The kinetic-energy derivative operator allows for capture into repulsive curves that are outside of the classical turning points for the nuclear motion. The dominant dissociative route is the C (2)Sigma(+) state leading to H(n = 2) atoms. An analogous mechanism is proposed for the dissociative recombination of H3(+).

  1. Recombination of N4(+) ions with electrons

    NASA Technical Reports Server (NTRS)

    Cao, Y. S.; Johnsen, R.

    1991-01-01

    Using a modified high-pressure-afterglow/mass spectrometer apparatus similar to that described by Lee and Johnsen (1989), spectroscopic observations of afterglow helium plasmas, with N2 as a minor additive, were carried out in order to verify the mechanism suggested by Bates (1991) for dissociative recombination of electrons with N4(+) ions. It was found that dissociative recombination of electrons with N4(+) ions results in the formation of N2 molecules in the C 3Pi(u) (v = 0,1) state, with the recombination rate coefficient of (2.6 +/- 0.3) x 10 exp -6 cu cm/sec at 300 K.

  2. Spacecraft thermal energy accommodation from atomic recombination

    NASA Technical Reports Server (NTRS)

    Carleton, Karen L.; Marinelli, William J.

    1991-01-01

    Measurements of atomic recombination probabilities important in determining energy release to reusable spacecraft thermal protection surfaces during reentry are presented. An experimental apparatus constructed to examine recombination of atomic oxygen from thermal protection and reference materials at reentry temperatures is described. The materials are examined under ultrahigh vacuum conditions to develop and maintain well characterized surface conditions that are free of contamination. When compared with stagnation point heat transfer measurements performed in arc jet facilities, these measurements indicate that a significant fraction of the excess energy available from atom recombination is removed from the surface as metastable O2.

  3. Bacterial genome remodeling through bacteriophage recombination.

    PubMed

    Menouni, Rachid; Hutinet, Geoffrey; Petit, Marie-Agnès; Ansaldi, Mireille

    2015-01-01

    Bacteriophages co-exist and co-evolve with their hosts in natural environments. Virulent phages lyse infected cells through lytic cycles, whereas temperate phages often remain dormant and can undergo lysogenic or lytic cycles. In their lysogenic state, prophages are actually part of the host genome and replicate passively in rhythm with host division. However, prophages are far from being passive residents: they can modify or bring new properties to their host. In this review, we focus on two important phage-encoded recombination mechanisms, i.e. site-specific recombination and homologous recombination, and how they remodel bacterial genomes. PMID:25790500

  4. Mapping Recombination Initiation Sites Using Chromatin Immunoprecipitation.

    PubMed

    He, Yan; Wang, Minghui; Sun, Qi; Pawlowski, Wojciech P

    2016-01-01

    Genome-wide maps of recombination sites provide valuable information not only on the recombination pathway itself but also facilitate the understanding of genome dynamics and evolution. Here, we describe a chromatin immunoprecipitation (ChIP) protocol to map the sites of recombination initiation in plants with maize used as an example. ChIP is a method that allows identification of chromosomal sites occupied by specific proteins. Our protocol utilizes RAD51, a protein involved in repair of double-strand breaks (DSBs) that initiate meiotic recombination, to identify DSB formation hotspots. Chromatin is extracted from meiotic flowers, sheared and enriched in fragments bound to RAD51. Genomic location of the protein is then identified by next-generation sequencing. This protocol can also be used in other species of plants, animals, and fungi. PMID:27511175

  5. Constraints from jet calculus on quark recombination

    SciTech Connect

    Jones, L.M.; Lassila, K.E.; Sukhatme, U.; Willen, D.

    1981-02-01

    Within the quantum-chromodynamic jet-calculus formalism, we deduce an equation describing recombination of quarks and antiquarks into mesons within a quark or gluon jet. This equation relates the recombination function R(x/sub 1/,x/sub 2/,x) used in current literature to the fragmentation function for producing that same meson out of the parton initiating the jet. We submit currently used recombination functions to our consistency test, taking as input mainly the u-quark fragmentation ''data'' into ..pi../sup +/ mesons. The qq-bar..--> pi.. recombination functions popular in the literature are consistent with measured fragmentation functions, but they must be supplemented by other contributions to provide the full D..pi../sup +//sub u/. We also discuss the Q/sup 2/ dependence of the resulting fragmentation functions.

  6. The Kinetics of Nitrogen Atom Recombination

    ERIC Educational Resources Information Center

    Brown, G. Ronald; Winkler, C. A.

    1977-01-01

    Describes a study of the kinetics of the recombination of nitrogen atoms in which concentration-time relations are determined directly by utilizing visual observations of emissions to make gas phase titrations of N atoms with NO. (MLH)

  7. Chemical recombination in an expansion tube

    NASA Technical Reports Server (NTRS)

    Bakos, Robert J.; Morgan, Richard G.

    1994-01-01

    The note describes the theoretical basis of chemical recombination in an expansion tube which simulates energy, Reynolds number, and stream chemistry at near-orbital velocities. Expansion tubes can satisfy ground-based hypersonic propulsion and aerothermal testing requirements.

  8. Recombinant Human Papillomavirus (HPV) Nonavalent Vaccine

    Cancer.gov

    This page contains brief information about recombinant human papillomavirus (HPV) nonavalent vaccine and a collection of links to more information about the use of this vaccine, research results, and ongoing clinical trials.

  9. Recombinant Human Papillomavirus (HPV) Bivalent Vaccine

    Cancer.gov

    This page contains brief information about recombinant human papillomavirus (HPV) bivalent vaccine and a collection of links to more information about the use of this vaccine, research results, and ongoing clinical trials.

  10. Recombinant Human Papillomavirus (HPV) Quadrivalent Vaccine

    Cancer.gov

    This page contains brief information about recombinant human papillomavirus (HPV) quadrivalent vaccine and a collection of links to more information about the use of this vaccine, research results, and ongoing clinical trials.

  11. Fermentations with new recombinant organisms

    SciTech Connect

    Bothast, R.J.; Nichols, N.N.; Dien, B.S.

    1999-10-01

    US fuel ethanol production in 1998 exceeded the record production of 1.4 billion gallons set in 1995. Most of this ethanol was produced from over 550 million bushels of corn. Expanding fuel ethanol production will require developing lower-cost feedstocks, and only lignocellulosic feedstocks are available in sufficient quantities to substitute for corn starch. Major technical hurdles to converting lignocellulose to ethanol include the lack of low-cost efficient enzymes for saccharification of biomass to fermentable sugars and the development of microorganisms for the fermentation of these mixed sugars. To date, the most successful research approaches to develop novel biocatalysts that will efficiently ferment mixed sugar syrups include isolation of novel yeasts that ferment xylose, genetic engineering of Escherichia coli and other gram negative bacteria for ethanol production, and genetic engineering of Saccharomyces cerevisiae and Zymomonas mobilis for pentose utilization. The authors have evaluated the fermentation of corn fiber hydrolyzates by the various strains developed. E. coli K011, E. coli SL40, E. coli FBR3, Zymomonas CP4 (pZB5), and Saccharomyces 1400 (pLNH32) fermented corn fiber hydrolyzates to ethanol in the range of 21--34 g/L with yields ranging from 0.41 to 0.50 g of ethanol per gram of sugar consumed. Progress with new recombinant microorganisms has been rapid and will continue with the eventual development of organisms suitable for commercial ethanol production. Each research approach holds considerable promise, with the possibility existing that different industrially hardened strains may find separate applications in the fermentation of specific feedstocks.

  12. Recombination walking: Genetic selection of clones from pooled libraries of yeast artificial chromosomes by homologous recombination

    SciTech Connect

    Miller, A.M.; Savinelli, E.A.; Couture, S.M.; Hannigan, G.M.; Han, Z.; Selden, R.F.; Treco, D.A. )

    1993-09-01

    Recombination walking is based on the genetic selection of specific human clones from a yeast artificial chromosome (YAC) library by homologous recombination. The desired clone is selected from a pooled (unorderd) YAC library, eliminating labor-intensive steps typically used in organizing and maintaining ordered YAC libraries. Recombination walking represents an efficient approach to library screening and is well suited for chromosome-walking approaches to the isolation of genes associated with common diseases. 29 refs., 4 figs., 1 tab.

  13. Recombination-deficient mutant of Streptococcus faecalis

    SciTech Connect

    Yagi, Y.; Clewell, D.B.

    1980-08-01

    An ultraviolet radiation-sensitive derivative of Streptococcus faecalis strain JH2-2 was isolated and found to be deficient in recombination, using a plasmid-plasmid recombination system. The strain was sensitive to chemical agents which interact with deoxyribonucleic acid and also underwent deoxyribonucleic acid degradation after ultraviolet irradiation. Thus, the mutant has properties similar to those of recA strains of Escherichia coli.

  14. Tunnel surface recombination in optoelectronic device modeling

    NASA Astrophysics Data System (ADS)

    Ptashchenko, Alexander A.; Ptashchenko, Fedor A.

    1997-08-01

    The rate of tunnel surface recombination (TSR) in a p-n structure has been calculated as a function of the excitation level and temperature in a semiclassical approximation under the assumption that the excess energy of a recombining electron is transferred to phonons or to a photon. The approximating analytical expressions obtained are applied in calculations of the effect of TSR on the characteristics of photodiodes, solar cells, light-emitting diodes and diode lasers.

  15. Co-factor activated recombinant adenovirus proteinases

    DOEpatents

    Anderson, Carl W.; Mangel, Walter F.

    1996-08-06

    This application describes methods and expression constructs for producing activatable recombinant adenovirus proteinases. Purified activatable recombinant adenovirus proteinases and methods of purification are described. Activated adenovirus proteinases and methods for obtaining activated adenovirus proteinases are further included. Isolated peptide cofactors of adenovirus proteinase activity, methods of purifying and identifying said peptide cofactors are also described. Antibodies immunoreactive with adenovirus proteinases, immunospecific antibodies, and methods for preparing them are also described. Other related methods and materials are also described.

  16. Co-factor activated recombinant adenovirus proteinases

    DOEpatents

    Anderson, C.W.; Mangel, W.F.

    1996-08-06

    This application describes methods and expression constructs for producing activatable recombinant adenovirus proteinases. Purified activatable recombinant adenovirus proteinases and methods of purification are described. Activated adenovirus proteinases and methods for obtaining activated adenovirus proteinases are further included. Isolated peptide cofactors of adenovirus proteinase activity, methods of purifying and identifying the peptide cofactors are also described. Antibodies immunoreactive with adenovirus proteinases, immunospecific antibodies, and methods for preparing them are also described. Other related methods and materials are also described. 29 figs.

  17. Recombination-generation currents in degenerate semiconductors

    NASA Technical Reports Server (NTRS)

    Von Roos, O.

    1978-01-01

    The classical Shockley-Read-Hall theory of free carrier recombination and generation via traps is extended to degenerate semiconductors. A concise and simple expression is found which avoids completely the concept of a Fermi level, a concept which is alien to nonequilibrium situations. Assumptions made in deriving the recombination generation current are carefully delineated and are found to be basically identical to those made in the original theory applicable to nondegenerate semiconductors.

  18. Detecting the cosmological recombination signal from space

    NASA Astrophysics Data System (ADS)

    Desjacques, Vincent; Chluba, Jens; Silk, Joseph; de Bernardis, Francesco; Doré, Olivier

    2015-08-01

    Spectral distortions of the cosmic microwave background (CMB) have recently experienced an increased interest. One of the inevitable distortion signals of our cosmological concordance model is created by the cosmological recombination process, just a little before photons last scatter at redshift z ≃ 1100. These cosmological recombination lines, emitted by the hydrogen and helium plasma, should still be observable as tiny deviation from the CMB blackbody spectrum in the cm-dm spectral bands. In this paper, we present a forecast for the detectability of the recombination signal with future satellite experiments. We argue that serious consideration for future CMB experiments in space should be given to probing spectral distortions and, in particular, the recombination line signals. The cosmological recombination radiation not only allows determination of standard cosmological parameters, but also provides a direct observational confirmation for one of the key ingredients of our cosmological model: the cosmological recombination history. We show that, with present technology, such experiments are futuristic but feasible. The potential rewards won by opening this new window to the very early universe could be considerable.

  19. Recombination hotspots: Models and tools for detection.

    PubMed

    Paul, Prosenjit; Nag, Debjyoti; Chakraborty, Supriyo

    2016-04-01

    Recombination hotspots are the regions within the genome where the rate, and the frequency of recombination are optimum with a size varying from 1 to 2kb. The recombination event is mediated by the double-stranded break formation, guided by the combined enzymatic action of DNA topoisomerase and Spo 11 endonuclease. These regions are distributed non-uniformly throughout the human genome and cause distortions in the genetic map. Numerous lines of evidence suggest that the number of hotspots known in humans has increased manifold in recent years. A few facts about the hotspot evolutions were also put forward, indicating the differences in the hotspot position between chimpanzees and humans. In mice, recombination hot spots were found to be clustered within the major histocompatibility complex (MHC) region. Several models, that help explain meiotic recombination has been proposed. Moreover, scientists also developed some computational tools to locate the hotspot position and estimate their recombination rate in humans is of great interest to population and medical geneticists. Here we reviewed the molecular mechanisms, models and in silico prediction techniques of hot spot residues. PMID:26991854

  20. Biochemistry of homologous recombination in Escherichia coli.

    PubMed Central

    Kowalczykowski, S C; Dixon, D A; Eggleston, A K; Lauder, S D; Rehrauer, W M

    1994-01-01

    Homologous recombination is a fundamental biological process. Biochemical understanding of this process is most advanced for Escherichia coli. At least 25 gene products are involved in promoting genetic exchange. At present, this includes the RecA, RecBCD (exonuclease V), RecE (exonuclease VIII), RecF, RecG, RecJ, RecN, RecOR, RecQ, RecT, RuvAB, RuvC, SbcCD, and SSB proteins, as well as DNA polymerase I, DNA gyrase, DNA topoisomerase I, DNA ligase, and DNA helicases. The activities displayed by these enzymes include homologous DNA pairing and strand exchange, helicase, branch migration, Holliday junction binding and cleavage, nuclease, ATPase, topoisomerase, DNA binding, ATP binding, polymerase, and ligase, and, collectively, they define biochemical events that are essential for efficient recombination. In addition to these needed proteins, a cis-acting recombination hot spot known as Chi (chi: 5'-GCTGGTGG-3') plays a crucial regulatory function. The biochemical steps that comprise homologous recombination can be formally divided into four parts: (i) processing of DNA molecules into suitable recombination substrates, (ii) homologous pairing of the DNA partners and the exchange of DNA strands, (iii) extension of the nascent DNA heteroduplex; and (iv) resolution of the resulting crossover structure. This review focuses on the biochemical mechanisms underlying these steps, with particular emphases on the activities of the proteins involved and on the integration of these activities into likely biochemical pathways for recombination. Images PMID:7968921

  1. Multiplex PCR Method for Identifying Recombinant Vaccine-Related Polioviruses

    PubMed Central

    Kilpatrick, David R.; Ching, Karen; Iber, Jane; Campagnoli, Ray; Freeman, Christopher J.; Mishrik, Nada; Liu, Hong-Mei; Pallansch, Mark A.; Kew, Olen M.

    2004-01-01

    The recent discovery of recombinant circulating vaccine-derived poliovirus (recombinant cVDPV) has highlighted the need for enhanced global poliovirus surveillance to assure timely detection of any future cVDPV outbreaks. Six pairs of Sabin strain-specific recombinant primers were designed to permit rapid screening for VDPV recombinants by PCR. PMID:15365031

  2. Oligonucleotide recombination enabled site-specific mutagenesis in bacteria

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Recombineering refers to a strategy for engineering DNA sequences using a specialized mode of homologous recombination. This technology can be used for rapidly constructing precise changes in bacterial genome sequences in vivo. Oligo recombination is one type of recombineering that uses ssDNA olig...

  3. The Saccharomyces cerevisiae recombination enhancer biases recombination during interchromosomal mating-type switching but not in interchromosomal homologous recombination.

    PubMed Central

    Houston, Peter; Simon, Peter J; Broach, James R

    2004-01-01

    Haploid Saccharomyces can change mating type through HO-endonuclease cleavage of an expressor locus, MAT, followed by gene conversion using one of two repository loci, HML or HMR, as donor. The mating type of a cell dictates which repository locus is used as donor, with a cells using HML and alpha cells using HMR. This preference is established in part by RE, a locus on the left arm of chromosome III that activates the surrounding region, including HML, for recombination in a cells, an activity suppressed by alpha 2 protein in alpha cells. We have examined the ability of RE to stimulate different forms of interchromosomal recombination. We found that RE exerted an effect on interchromosomal mating-type switching and on intrachromosomal homologous recombination but not on interchromosomal homologous recombination. Also, even in the absence of RE, MAT alpha still influenced donor preference in interchromosomal mating-type switching, supporting a role of alpha 2 in donor preference independent of RE. These results suggest a model in which RE affects competition between productive and nonproductive recombination outcomes. In interchromosome gene conversion, RE enhances both productive and nonproductive pathways, whereas in intrachromosomal gene conversion and mating-type switching, RE enhances only the productive pathway. PMID:15082540

  4. DNA RECOMBINATION IN EUCARYOTIC CELLS BY THE BACTERIOPHAGE PHIC31 RECOMBINATION SYSTEM",

    Technology Transfer Automated Retrieval System (TEKTRAN)

    This invention provides methods for obtaining specific and stable integration of nucleic acids into eukaryotic cells. The invention makes use of site-specific recombination systems that use prokaryotic recombinase polypeptides, such as the ph:C31 integrase, that can mediate recombination between th...

  5. Perinatal induction of Cre recombination with tamoxifen.

    PubMed

    Lizen, Benoit; Claus, Melissa; Jeannotte, Lucie; Rijli, Filippo M; Gofflot, Françoise

    2015-12-01

    Temporal control of site-specific recombination is commonly achieved by using a tamoxifen-inducible form of Cre or Flp recombinases. Although powerful protocols of induction have been developed for gene inactivation at adult stages or during embryonic development, induction of recombination at late gestational or early postnatal stages is still difficult to achieve. In this context, using the ubiquitous CMV-CreER(T2) transgenic mice, we have tested and validated two procedures to achieve recombination just before and just after birth. The efficiency of recombination was evaluated in the brain, which is known to be more problematic to target. For the late gestation treatment with tamoxifen, different protocols of complementary administration of progesterone and estrogen were tested. However, delayed delivery and/or mortality of pups due to difficult delivery were always observed. To circumvent this problem, pups were collected from tamoxifen-treated pregnant dams by caesarian section at E18.5 and given to foster mothers. For postnatal treatment, different dosages of tamoxifen were administered by intragastric injection to the pups during 3 or 4 days after birth. The efficiency of these treatments was analyzed at P7 using a transgenic reporter line. They were also validated with the Hoxa5 conditional allele. In conclusion, we have developed efficient procedures that allow achieving efficient recombination of floxed alleles at perinatal stages. These protocols will allow investigating the late/adult functions of many developmental genes, whose characterization has been so far restricted to embryonic development. PMID:26395370

  6. Human recombinant lysosomal enzymes produced in microorganisms.

    PubMed

    Espejo-Mojica, Ángela J; Alméciga-Díaz, Carlos J; Rodríguez, Alexander; Mosquera, Ángela; Díaz, Dennis; Beltrán, Laura; Díaz, Sergio; Pimentel, Natalia; Moreno, Jefferson; Sánchez, Jhonnathan; Sánchez, Oscar F; Córdoba, Henry; Poutou-Piñales, Raúl A; Barrera, Luis A

    2015-01-01

    Lysosomal storage diseases (LSDs) are caused by accumulation of partially degraded substrates within the lysosome, as a result of a function loss of a lysosomal protein. Recombinant lysosomal proteins are usually produced in mammalian cells, based on their capacity to carry out post-translational modifications similar to those observed in human native proteins. However, during the last years, a growing number of studies have shown the possibility to produce active forms of lysosomal proteins in other expression systems, such as plants and microorganisms. In this paper, we review the production and characterization of human lysosomal proteins, deficient in several LSDs, which have been produced in microorganisms. For this purpose, Escherichia coli, Saccharomyces cerevisiae, Pichia pastoris, Yarrowia lipolytica, and Ogataea minuta have been used as expression systems. The recombinant lysosomal proteins expressed in these hosts have shown similar substrate specificities, and temperature and pH stability profiles to those produced in mammalian cells. In addition, pre-clinical results have shown that recombinant lysosomal enzymes produced in microorganisms can be taken-up by cells and reduce the substrate accumulated within the lysosome. Recently, metabolic engineering in yeasts has allowed the production of lysosomal enzymes with tailored N-glycosylations, while progresses in E. coli N-glycosylations offer a potential platform to improve the production of these recombinant lysosomal enzymes. In summary, microorganisms represent convenient platform for the production of recombinant lysosomal proteins for biochemical and physicochemical characterization, as well as for the development of ERT for LSD. PMID:26071627

  7. Graded recombination layers for multijunction photovoltaics.

    PubMed

    Koleilat, Ghada I; Wang, Xihua; Sargent, Edward H

    2012-06-13

    Multijunction devices consist of a stack of semiconductor junctions having bandgaps tuned across a broad spectrum. In solar cells this concept is used to increase the efficiency of photovoltaic harvesting, while light emitters and detectors use it to achieve multicolor and spectrally tunable behavior. In series-connected current-matched multijunction devices, the recombination layers must allow the hole current from one cell to recombine, with high efficiency and low voltage loss, with the electron current from the next cell. We recently reported a tandem solar cell in which the recombination layer was implemented using a progression of n-type oxides whose doping densities and work functions serve to connect, with negligible resistive loss at solar current densities, the constituent cells. Here we present the generalized conditions for design of efficient graded recombination layer solar devices. We report the number of interlayers and the requirements on work function and doping of each interlayer, to bridge an work function difference as high as 1.6 eV. We also find solutions that minimize the doping required of the interlayers in order to minimize optical absorption due to free carriers in the graded recombination layer (GRL). We demonstrate a family of new GRL designs experimentally and highlight the benefits of the progression of dopings and work functions in the interlayers. PMID:22554234

  8. Recombination in Eukaryotic Single Stranded DNA Viruses

    PubMed Central

    Martin, Darren P.; Biagini, Philippe; Lefeuvre, Pierre; Golden, Michael; Roumagnac, Philippe; Varsani, Arvind

    2011-01-01

    Although single stranded (ss) DNA viruses that infect humans and their domesticated animals do not generally cause major diseases, the arthropod borne ssDNA viruses of plants do, and as a result seriously constrain food production in most temperate regions of the world. Besides the well known plant and animal-infecting ssDNA viruses, it has recently become apparent through metagenomic surveys of ssDNA molecules that there also exist large numbers of other diverse ssDNA viruses within almost all terrestrial and aquatic environments. The host ranges of these viruses probably span the tree of life and they are likely to be important components of global ecosystems. Various lines of evidence suggest that a pivotal evolutionary process during the generation of this global ssDNA virus diversity has probably been genetic recombination. High rates of homologous recombination, non-homologous recombination and genome component reassortment are known to occur within and between various different ssDNA virus species and we look here at the various roles that these different types of recombination may play, both in the day-to-day biology, and in the longer term evolution, of these viruses. We specifically focus on the ecological, biochemical and selective factors underlying patterns of genetic exchange detectable amongst the ssDNA viruses and discuss how these should all be considered when assessing the adaptive value of recombination during ssDNA virus evolution. PMID:21994803

  9. Recombination and collisionally excited Balmer lines

    NASA Astrophysics Data System (ADS)

    Raga, A. C.; Castellanos-Ramírez, A.; Esquivel, A.; Rodríguez-González, A.; Velázquez, P. F.

    2015-10-01

    We present a model for the statistical equilibrium of the levels of H, considering recombinations to excited levels, collisional excitations up from the ground state and spontaneous radiative transitions. This problem has a simple "cascade matrix" solution, describing a cascade of downwards spontaneous transitions fed by both recombinations and collisional excitations. The resulting predicted Balmer line ratios show a transition between a low temperature and a high temperature regime (dominated by recombinations and by collisional excitations, respectively), both with only a weak line ratio vs. temperature dependence. This clear characteristic allows a direct observational identification of regions in which the Balmer lines are either recombination or collisionally excited transitions. We find that for a gas in coronal ionization equilibrium the Halpha and Hbeta lines are collisionally excited for all temperatures. In order to have recombination Halpha and Hbeta it is necessary to have higher ionization fractions of H than the ones obtained from coronal equilibrium (e.g., such as the ones found in a photoionized gas).

  10. Meiotic recombination and genome evolution in plants.

    PubMed

    Melamed-Bessudo, Cathy; Shilo, Shay; Levy, Avraham A

    2016-04-01

    Homologous recombination affects genome evolution through crossover, gene conversion and point mutations. Whole genome sequencing together with a detailed epigenome analysis have shed new light on our understanding of how meiotic recombination shapes plant genes and genome structure. Crossover events are associated with DNA sequence motifs, together with an open chromatin signature (hypomethylated CpGs, low nucleosome occupancy or specific histone modifications). The crossover landscape may differ between male and female meiocytes and between species. At the gene level, crossovers occur preferentially in promoter regions in Arabidopsis. In recent years, there is rising support suggesting that biased mismatch repair during meiotic recombination may increase GC content genome-wide and may be responsible for the GC content gradient found in many plant genes. PMID:26939088

  11. Dielectronic recombination of xenonlike tungsten ions

    SciTech Connect

    Schippers, S.; Bernhardt, D.; Mueller, A.; Krantz, C.; Grieser, M.; Repnow, R.; Wolf, A.; Lestinsky, M.; Hahn, M.; Novotny, O.; Savin, D. W.

    2011-01-15

    Dielectronic recombination (DR) of xenonlike W{sup 20+} forming W{sup 19+} has been studied experimentally at a heavy-ion storage ring. A merged-beams method has been employed for obtaining absolute rate coefficients for electron-ion recombination in the collision-energy range 0-140 eV. The measured rate coefficient is dominated by strong DR resonances even at the lowest experimental energies. At plasma temperatures where the fractional abundance of W{sup 20+} is expected to peak in a fusion plasma, the experimentally derived plasma recombination rate coefficient is over a factor of 4 larger than the theoretically calculated rate coefficient which is currently used in fusion plasma modeling. The largest part of this discrepancy stems most probably from the neglect in the theoretical calculations of DR associated with fine-structure excitations of the W{sup 20+}([Kr]4d{sup 10} 4f{sup 8}) ion core.

  12. Transposon-specified site-specific recombination.

    PubMed Central

    Kitts, P; Symington, L; Burke, M; Reed, R; Sherratt, D

    1982-01-01

    Cointegrate DNA molecules containing two copies of a transposable element appear to be intermediates in the transposition process. These structures are resolved by site-specific recombination to yield the normal end products of transposition. The transposable element gamma delta (Tn1000) synthesizes a product interchangeable with the Tn1/3tnpR protein in promoting Tn1/3 site-specific recombination. These data support the hypothesis that cointegrates containing directly repeated copies of Tn1/3 are obligatory intermediates in interreplicon transposition of Tn1/3. In addition, we show here that the reaction is independent of the element-encoded tnpA gene product. Tn501, which specifies mercury resistance, also produces cointegrates as intermediates in interreplicon transposition. The appearance of Tn501-specified recombination activity that can act on these cointegrates requires growth of cells in the presence of Hg2+. Images PMID:6275390

  13. Generation of active immunotoxins containing recombinant restrictocin.

    PubMed

    Rathore, D; Batra, J K

    1996-05-01

    Restrictocin, a toxin produced by the fungus Aspergillus restrictus, is a potent inhibitor of eukaryotic protein synthesis. Recombinant restrictocin was made in Escherichia coli and purified to homogeneity in large amounts. The recombinant protein was found to be poorly immunogenic in mice with low toxicity, when injected intraperitoneally. Two immunotoxins were constructed by coupling the recombinant restrictocin to an antibody to the human transferrin receptor, using a cleavable and a stable linkage. The immunotoxins so generated showed specific cytotoxic activity toward receptor bearing cells in tissue culture. Immunotoxin with a cleavable linkage, however, was more active than that containing a stable linkage. Restrictocin appears to be a promising candidate to be developed as a chimeric toxin for targeted therapy. PMID:8630074

  14. Transcription and Recombination: When RNA Meets DNA

    PubMed Central

    Aguilera, Andrés; Gaillard, Hélène

    2014-01-01

    A particularly relevant phenomenon in cell physiology and proliferation is the fact that spontaneous mitotic recombination is strongly enhanced by transcription. The most accepted view is that transcription increases the occurrence of double-strand breaks and/or single-stranded DNA gaps that are repaired by recombination. Most breaks would arise as a consequence of the impact that transcription has on replication fork progression, provoking its stalling and/or breakage. Here, we discuss the mechanisms responsible for the cross talk between transcription and recombination, with emphasis on (1) the transcription–replication conflicts as the main source of recombinogenic DNA breaks, and (2) the formation of cotranscriptional R-loops as a major cause of such breaks. The new emerging questions and perspectives are discussed on the basis of the interference between transcription and replication, as well as the way RNA influences genome dynamics. PMID:25085910

  15. Recombination and DNA Repair in Helicobacter pylori

    PubMed Central

    Dorer, Marion S.; Sessler, Tate H.; Salama, Nina R.

    2013-01-01

    All organisms have pathways that repair the genome, ensuring their survival and that of their progeny. But these pathways also serve to diversify the genome, causing changes on the level of nucleotide, whole gene, and genome structure. Sequencing of bacteria has revealed wide allelic diversity and differences in gene content within the same species, highlighting the importance of understanding pathways of recombination and DNA repair. The human stomach pathogen Helicobacter pylori is an excellent model system for studying these pathways. H. pylori harbors major recombination and repair pathways and is naturally competent, facilitating its ability to diversify its genome. Elucidation of DNA recombination, repair, and diversification programs in this pathogen will reveal connections between these pathways and their importance to infection. PMID:21682641

  16. Jet fragmentation via recombination of parton showers

    NASA Astrophysics Data System (ADS)

    Han, Kyong Chol; Fries, Rainer J.; Ko, Che Ming

    2016-04-01

    We propose to model hadronization of parton showers in QCD jets through a hybrid approach involving quark recombination and string fragmentation. This is achieved by allowing gluons at the end of the perturbative shower evolution to undergo a nonperturbative splitting into quark and antiquark pairs, then applying a Monte Carlo version of instantaneous quark recombination, and finally subjecting remnant quarks (those which have not found a recombination partner) to Lund string fragmentation. When applied to parton showers from the pythia Monte Carlo event generator, the final hadron spectra from our calculation compare quite well to pythia jets that have been hadronized with the default Lund string fragmentation. Our new approach opens up the possibility to generalize hadronization to jets embedded in a quark gluon plasma.

  17. CosmoRec: Cosmological Recombination code

    NASA Astrophysics Data System (ADS)

    Chluba, Jens; Thomas, Rajat Mani

    2013-04-01

    CosmoRec solves the recombination problem including recombinations to highly excited states, corrections to the 2s-1s two-photon channel, HI Lyn-feedback, n>2 two-photon profile corrections, and n≥2 Raman-processes. The code can solve the radiative transfer equation of the Lyman-series photon field to obtain the required modifications to the rate equations of the resolved levels, and handles electron scattering, the effect of HeI intercombination transitions, and absorption of helium photons by hydrogen. It also allows accounting for dark matter annihilation and optionally includes detailed helium radiative transfer effects.

  18. Graph Model of Coalescence with Recombinations

    NASA Astrophysics Data System (ADS)

    Parida, Laxmi

    One of the primary genetic events shaping an autosomal chromosome is recombination. This is a process that occurs during meiosis, in eukaryotes, that results in the offsprings having different combinations of (homologous) genes, or chromosomal segments, of the two parents. The presence of these recombination events in the evolutionary history of each chromosome complicates the genetic landscape of a population, and understanding the manifestations of these genetic exchanges in the chromosome sequences has been a subject of intense curiosity (see [Hud83, Gri99, HSW05] and citations therein).

  19. Recent Theoretical Studies On Excitation and Recombination

    NASA Technical Reports Server (NTRS)

    Pradhan, Anil K.

    2000-01-01

    New advances in the theoretical treatment of atomic processes in plasmas are described. These enable not only an integrated, unified, and self-consistent treatment of important radiative and collisional processes, but also large-scale computation of atomic data with high accuracy. An extension of the R-matrix work, from excitation and photoionization to electron-ion recombination, includes a unified method that subsumes both the radiative and the di-electronic recombination processes in an ab initio manner. The extensive collisional calculations for iron and iron-peak elements under the Iron Project are also discussed.

  20. Lectin glycoprofiling of recombinant therapeutic interleukin-7.

    PubMed

    Landemarre, Ludovic; Duverger, Eric

    2013-01-01

    Lectins array is a powerfull and complementary method of glycans analysis allowing fast identification of specific motifs on molecules or cells. This technology is of increased interest for the development of therapeutic recombinant glycoproteins and particularly relevant for a first study of lot-to-lot comparison, or detection of unwanted glycans. In this chapter, we describe a lectin array-type method specifically designed for the study of recombinant therapeutic interleukin-7 (rhIL-7). This specific method allows the analysis of the glycans motifs, the distribution of the glycoforms population, and the detection of potential immunogen glycans in rhIL-7 purified CHO-produced batches. PMID:23475723

  1. The mismatch repair system reduces meiotic homeologous recombination and stimulates recombination-dependent chromosome loss.

    PubMed Central

    Chambers, S R; Hunter, N; Louis, E J; Borts, R H

    1996-01-01

    Efficient genetic recombination requires near-perfect homology between participating molecules. Sequence divergence reduces the frequency of recombination, a process that is dependent on the activity of the mismatch repair system. The effects of chromosomal divergence in diploids of Saccharomyces cerevisiae in which one copy of chromosome III is derived from a closely related species, Saccharomyces paradoxus, have been examined. Meiotic recombination between the diverged chromosomes is decreased by 25-fold. Spore viability is reduced with an observable increase in the number of tetrads with only two or three viable spores. Asci with only two viable spores are disomic for chromosome III, consistent with meiosis I nondisjunction of the homeologs. Asci with three viable spores are highly enriched for recombinants relative to tetrads with four viable spores. In 96% of the class with three viable spores, only one spore possesses a recombinant chromosome III, suggesting that the recombination process itself contributes to meiotic death. This phenomenon is dependent on the activities of the mismatch repair genes PMS1 and MSH2. A model of mismatch-stimulated chromosome loss is proposed to account for this observation. As expected, crossing over is increased in pms1 and msh2 mutants. Furthermore, genetic exchange in pms1 msh2 double mutants is affected to a greater extent than in either mutant alone, suggesting that the two proteins act independently to inhibit homeologous recombination. All mismatch repair-deficient strains exhibited reductions in the rate of chromosome III nondisjunction. PMID:8887641

  2. Recombinant microorganisms for increased production of organic acids

    DOEpatents

    Yi, Jian; Kleff, Susanne; Guettler, Michael V.

    2012-02-21

    Disclosed are recombinant microorganisms for producing organic acids. The recombinant microorganisms express a polypeptide that has the enzymatic activity of an enzyme that is utilized in the pentose phosphate cycle. The recombinant microorganism may include recombinant Actinobacillus succinogenes that has been transformed to express a Zwischenferment (Zwf) gene. The recombinant microorganisms may be useful in fermentation processes for producing organic acids such as succinic acid and lactic acid. Also disclosed are novel plasmids that are useful for transforming microorganisms to produce recombinant microorganisms that express enzymes such as Zwf.

  3. Recombinant microorganisms for increased production of organic acids

    DOEpatents

    Yi, Jian; Kleff, Susanne; Guettler, Michael V

    2013-04-30

    Disclosed are recombinant microorganisms for producing organic acids. The recombinant microorganisms express a polypeptide that has the enzymatic activity of an enzyme that is utilized in the pentose phosphate cycle. The recombinant microorganism may include recombinant Actinobacillus succinogenes that has been transformed to express a Zwischenferment (Zwf) gene. The recombinant microorganisms may be useful in fermentation processes for producing organic acids such as succinic acid and lactic acid. Also disclosed are novel plasmids that are useful for transforming microorganisms to produce recombinant microorganisms that express enzymes such as Zwf.

  4. Recombinant Bovine Growth Hormone Criticism Grows.

    ERIC Educational Resources Information Center

    Gaard, Greta

    1995-01-01

    Discusses concerns related to the use of recombinant bovine growth hormone in the United States and other countries. Analyses the issue from the perspectives of animal rights, human health, world hunger, concerns of small and organic farmers, costs to the taxpayer, and environmental questions. A sidebar discusses Canadian review of the hormone.…

  5. Science: The Recombinant DNA Advisory Committee.

    ERIC Educational Resources Information Center

    Wright, Susan

    1979-01-01

    Reports on the status of the Recombinant DNA Advisory Committee (RAC) and attempts to rationalize Suburban Highway Policy. Effective communication among members of the RAC is a current problem facing the committee. A federal transportation priority spending policy is suggested during these times of money and fuel shortages. (MA)

  6. Vaccine development using recombinant DNA technology

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Vaccines induce an immune response in the host that subsequently recognizes infectious agents and helps fight off the disease; vaccines must do this without causing the disease. This paper reviews the development of recombinant DNA technologies as a means of providing new ways for attenuating diseas...

  7. Recombinant protein blends: silk beyond natural design.

    PubMed

    Dinjaski, Nina; Kaplan, David L

    2016-06-01

    Recombinant DNA technology and new material concepts are shaping future directions in biomaterial science for the design and production of the next-generation biomaterial platforms. Aside from conventionally used synthetic polymers, numerous natural biopolymers (e.g., silk, elastin, collagen, gelatin, alginate, cellulose, keratin, chitin, polyhydroxyalkanoates) have been investigated for properties and manipulation via bioengineering. Genetic engineering provides a path to increase structural and functional complexity of these biopolymers, and thereby expand the catalog of available biomaterials beyond that which exists in nature. In addition, the integration of experimental approaches with computational modeling to analyze sequence-structure-function relationships is starting to have an impact in the field by establishing predictive frameworks for determining material properties. Herein, we review advances in recombinant DNA-mediated protein production and functionalization approaches, with a focus on hybrids or combinations of proteins; recombinant protein blends or 'recombinamers'. We highlight the potential biomedical applications of fibrous protein recombinamers, such as Silk-Elastin Like Polypeptides (SELPs) and Silk-Bacterial Collagens (SBCs). We also discuss the possibility for the rationale design of fibrous proteins to build smart, stimuli-responsive biomaterials for diverse applications. We underline current limitations with production systems for these proteins and discuss the main trends in systems/synthetic biology that may improve recombinant fibrous protein design and production. PMID:26686863

  8. CATALYTIC RECOMBINER FOR A NUCLEAR REACTOR

    DOEpatents

    King, L.D.P.

    1960-07-01

    A hydrogen-oxygen recombiner is described for use with water-boiler type reactors. The catalyst used is the wellknown platinized alumina, and the novelty lies in the structural arrangement used to prevent flashback through the gas input system. The recombiner is cylindrical, the gases at the input end being deflected by a baffle plate through a first flashback shield of steel shot into an annular passage adjacent to and extending the full length of the housing. Below the baffle plate the gases flow first through an outer annular array of alumina pellets which serve as a second flashback shield, a means of distributing the flowing gases evenly and as a means of reducing radiation losses to the walls. Thereafter the gases flow inio the centrally disposed catalyst bed where recombination is effected. The steam and uncombined gases flow into a centrally disposed cylindrical passage inside the catalyst bod and thereafter out through the exit port. A high rate of recombination is effected.

  9. Gas recombination assembly for electrochemical cells

    DOEpatents

    Levy, Isaac; Charkey, Allen

    1989-01-01

    An assembly for recombining gases generated in electrochemical cells wherein a catalyst strip is enveloped within a hydrophobic, gas-porous film which, in turn, is encased between gas-porous, metallic layers. The sandwich construction of metallic layers and film is formed into a spiral with a tab for connection to the cell.

  10. Recombinant DNA: Scientific and Social Perspectives.

    ERIC Educational Resources Information Center

    Vandegrift, Vaughn

    1979-01-01

    This article is designed to inform chemical educators not engaged in this technology as to the nature and methods used in the technology, the reasons for scientific and social concern, and the attempts made to assuage concerns involving recombinant DNA research. (author/BB)

  11. Recombinant therapeutic proteins: production platforms and challenges.

    PubMed

    Dingermann, Theo

    2008-01-01

    Since the approval of insulin in 1982, more than 120 recombinant drug substances have been approved and become available as extremely valuable therapeutic options. Exact copying of the most common human form is no longer a value per se, as challenges, primarily related to the pharmacokinetics of artificial recombinant drugs, can be overcome by diverging from the original. However, relatively minor changes in manufacturing or packaging may impact safety of therapeutic proteins. A major achievement is the development of recombinant proteins capable of entering a cell. Such drugs open up completely new opportunities by targeting intracellular mechanisms or by substituting intracellularly operating enzymes. Concerns that protein variants would cause an intolerable immune response turned out to be exaggerated. Although most recombinant drugs provoke some immune response, they are still well tolerated. This knowledge might result in a change in attitude towards antibody formation, i.e., neutralizing antibody activity (in vitro) may be overcome by dosing consistently on the basis of antibody titers and not only on body weight. As with other drugs, efficacy and safety of therapeutic proteins have to be demonstrated in clinical studies, and superiority over available products has to be proven instead of just claimed. PMID:18041103

  12. Precise genotyping and recombination detection of Enterovirus

    PubMed Central

    2015-01-01

    Enteroviruses (EV) with different genotypes cause diverse infectious diseases in humans and mammals. A correct EV typing result is crucial for effective medical treatment and disease control; however, the emergence of novel viral strains has impaired the performance of available diagnostic tools. Here, we present a web-based tool, named EVIDENCE (EnteroVirus In DEep conception, http://symbiont.iis.sinica.edu.tw/evidence), for EV genotyping and recombination detection. We introduce the idea of using mixed-ranking scores to evaluate the fitness of prototypes based on relatedness and on the genome regions of interest. Using phylogenetic methods, the most possible genotype is determined based on the closest neighbor among the selected references. To detect possible recombination events, EVIDENCE calculates the sequence distance and phylogenetic relationship among sequences of all sliding windows scanning over the whole genome. Detected recombination events are plotted in an interactive figure for viewing of fine details. In addition, all EV sequences available in GenBank were collected and revised using the latest classification and nomenclature of EV in EVIDENCE. These sequences are built into the database and are retrieved in an indexed catalog, or can be searched for by keywords or by sequence similarity. EVIDENCE is the first web-based tool containing pipelines for genotyping and recombination detection, with updated, built-in, and complete reference sequences to improve sensitivity and specificity. The use of EVIDENCE can accelerate genotype identification, aiding clinical diagnosis and enhancing our understanding of EV evolution. PMID:26678286

  13. DNA Sequence Alignment during Homologous Recombination.

    PubMed

    Greene, Eric C

    2016-05-27

    Homologous recombination allows for the regulated exchange of genetic information between two different DNA molecules of identical or nearly identical sequence composition, and is a major pathway for the repair of double-stranded DNA breaks. A key facet of homologous recombination is the ability of recombination proteins to perfectly align the damaged DNA with homologous sequence located elsewhere in the genome. This reaction is referred to as the homology search and is akin to the target searches conducted by many different DNA-binding proteins. Here I briefly highlight early investigations into the homology search mechanism, and then describe more recent research. Based on these studies, I summarize a model that includes a combination of intersegmental transfer, short-distance one-dimensional sliding, and length-specific microhomology recognition to efficiently align DNA sequences during the homology search. I also suggest some future directions to help further our understanding of the homology search. Where appropriate, I direct the reader to other recent reviews describing various issues related to homologous recombination. PMID:27129270

  14. Precise genotyping and recombination detection of Enterovirus.

    PubMed

    Lin, Chieh-Hua; Wang, Yu-Bin; Chen, Shu-Hwa; Hsiung, Chao Agnes; Lin, Chung-Yen

    2015-01-01

    Enteroviruses (EV) with different genotypes cause diverse infectious diseases in humans and mammals. A correct EV typing result is crucial for effective medical treatment and disease control; however, the emergence of novel viral strains has impaired the performance of available diagnostic tools. Here, we present a web-based tool, named EVIDENCE (EnteroVirus In DEep conception, http://symbiont.iis.sinica.edu.tw/evidence), for EV genotyping and recombination detection. We introduce the idea of using mixed-ranking scores to evaluate the fitness of prototypes based on relatedness and on the genome regions of interest. Using phylogenetic methods, the most possible genotype is determined based on the closest neighbor among the selected references. To detect possible recombination events, EVIDENCE calculates the sequence distance and phylogenetic relationship among sequences of all sliding windows scanning over the whole genome. Detected recombination events are plotted in an interactive figure for viewing of fine details. In addition, all EV sequences available in GenBank were collected and revised using the latest classification and nomenclature of EV in EVIDENCE. These sequences are built into the database and are retrieved in an indexed catalog, or can be searched for by keywords or by sequence similarity. EVIDENCE is the first web-based tool containing pipelines for genotyping and recombination detection, with updated, built-in, and complete reference sequences to improve sensitivity and specificity. The use of EVIDENCE can accelerate genotype identification, aiding clinical diagnosis and enhancing our understanding of EV evolution. PMID:26678286

  15. Recombination phenomena in high efficiency silicon solar cells

    NASA Technical Reports Server (NTRS)

    Sah, C. T.

    1985-01-01

    The dominant recombination phenomena which limit the highest efficiency attainable in silicon solar cells under terrestrial sunlight are reviewed. The ultimate achievable efficiency is limited by the two intrinsic recombination mechanisms, the interband Auger recombination and interband Radiative recombination, both of which occur in the entire cell body but principally in the base layer. It is suggested that an optimum (26%) cell design is one with lowly doped 50 to 100 micron thick base, a perfect BSF, and zero extrinsic recombination such as the thermal mechanism at recombination centers the Shockley-Read-Hall process (SRH) in the bulk, on the surface and at the interfaces. The importance of recombination at the interfaces of a high-efficiency cell is demonstrated by the ohmic contact on the back surface whose interface recombination velocity is infinite. The importance of surface and interface recombination is demonstrated by representing the auger and radiative recombination losses by effective recombination velocities. It is demonstrated that the three highest efficiency cells may all be limited by the SRH recombination losses at recombination centers in the base layer.

  16. Radiative transfer effects in primordial hydrogen recombination

    SciTech Connect

    Ali-Haiemoud, Yacine; Hirata, Christopher M.; Grin, Daniel

    2010-12-15

    The calculation of a highly accurate cosmological recombination history has been the object of particular attention recently, as it constitutes the major theoretical uncertainty when predicting the angular power spectrum of cosmic microwave background anisotropies. Lyman transitions, in particular the Lyman-{alpha} line, have long been recognized as one of the bottlenecks of recombination, due to their very low escape probabilities. The Sobolev approximation does not describe radiative transfer in the vicinity of Lyman lines to a sufficient degree of accuracy, and several corrections have already been computed in other works. In this paper, we compute the impact of some radiative transfer effects that were previously ignored, or for which previous treatments were incomplete. First, the effect of Thomson scattering in the vicinity of the Lyman-{alpha} line is evaluated, using a full redistribution kernel incorporated into a radiative transfer code. The effect of feedback of distortions generated by the optically thick deuterium Lyman-{alpha} line blueward of the hydrogen line is investigated with an analytic approximation. It is shown that both effects are negligible during cosmological hydrogen recombination. Second, the importance of high-lying, nonoverlapping Lyman transitions is assessed. It is shown that escape from lines above Ly{gamma} and frequency diffusion in Ly{beta} and higher lines can be neglected without loss of accuracy. Third, a formalism generalizing the Sobolev approximation is developed to account for the overlap of the high-lying Lyman lines, which is shown to lead to negligible changes to the recombination history. Finally, the possibility of a cosmological hydrogen recombination maser is investigated. It is shown that there is no such maser in the purely radiative treatment presented here.

  17. Ancestries of a recombining diploid population.

    PubMed

    Sainudiin, R; Thatte, B; Véber, A

    2016-01-01

    We derive the exact one-step transition probabilities of the number of lineages that are ancestral to a random sample from the current generation of a bi-parental population that is evolving under the discrete Wright-Fisher model with n diploid individuals. Our model allows for a per-generation recombination probability of r . When r = 1, our model is equivalent to Chang's (Adv Appl Probab 31:1002-1038, 1999) model for the karyotic pedigree. When r = 0, our model is equivalent to Kingman's (Stoch Process Appl 13:235-248, 1982) discrete coalescent model for the cytoplasmic tree or sub-karyotic tree containing a DNA locus that is free of intra-locus recombination. When 0 < r < 1 our model can be thought to track a sub-karyotic ancestral graph containing a DNA sequence from an autosomal chromosome that has an intra-locus recombination probability r . Thus, our family of models indexed by r ∈ [0, 1] connects Kingman's discrete coalescent to Chang's pedigree in a continuous way as r goes from 0 to 1. For large populations, we also study three properties of the ancestral process corresponding to a given r ∈ (0, 1): the time Tn to a most recent common ancestor (MRCA) of the population, the time Un at which all individuals are either common ancestors of all present day individuals or ancestral to none of them, and the fraction of individuals that are common ancestors at time Un. These results generalize the three main results of Chang's (Adv Appl Probab 31:1002-1038, 1999). When we appropriately rescale time and recombination probability by the population size, our model leads to the continuous time Markov chain called the ancestral recombination graph of Hudson (Theor Popul Biol 23:183-201, 1983) and Griffiths (The two-locus ancestral graph, Institute of Mathematical Statistics 100-117, 1991). PMID:25925241

  18. Extrachromosomal recombination substrates recapitulate beyond 12/23 restricted VDJ recombination in nonlymphoid cells.

    PubMed

    Jung, David; Bassing, Craig H; Fugmann, Sebastian D; Cheng, Hwei-Ling; Schatz, David G; Alt, Frederick W

    2003-01-01

    V(D)J recombination occurs efficiently only between gene segments flanked by recombination signals (RSs) containing 12 and 23 base pair spacers (the 12/23 rule). A further limitation "beyond the 12/23 rule" (B12/23) exists at the TCRbeta locus and ensures Dbeta usage. Herein, we show that extrachromosomal V(D)J recombination substrates recapitulate B12/23 restriction in nonlymphoid cells. We further demonstrate that the Vbeta coding flank, the 12-RS heptamer/nonamer, and the 23-RS spacer each can significantly influence B12/23 restriction. Finally, purified core RAG1 and RAG2 proteins (together with HMG2) also reproduce B12/23 restriction in a cell-free system. Our findings indicate that B12/23 restriction of V(D)J recombination is cemented at the level of interactions between the RAG proteins and TCRbeta RS sequences. PMID:12530976

  19. Bacteriophage T4 DNA Topoisomerase Mediates Illegitimate Recombination in vitro

    NASA Astrophysics Data System (ADS)

    Ikeda, Hideo

    1986-02-01

    We have found that purified T4 DNA topoisomerase promotes recombination between two phage λ DNA molecules in an in vitro system. In this cross, T4 DNA topoisomerase alone is able to catalyze the recombination and produce a linear monomer recombinant DNA that can be packaged in vitro. ATP is not required for this recombination, while oxolinic acid stimulates it. The recombinant DNA molecules contain duplications or deletions, and the crossovers take place between nonhomologous and nonspecific sequences of λ DNA. Therefore, the recombination mediated by the T4 DNA topoisomerase is an illegitimate recombination that is similar to that mediated by Escherichia coli DNA gyrase. A model was proposed previously in which DNA gyrase molecules that are bound to DNA associate with each other and lead to the exchange of DNA strands through the exchange of DNA gyrase subunits. This model is also applicable to the recombination mediated by T4 DNA topoisomerase.

  20. Serial Recombination during Circulation of Type 1 Wild-Vaccine Recombinant Polioviruses in China

    PubMed Central

    Liu, Hong-Mei; Zheng, Du-Ping; Zhang, Li-Bi; Oberste, M. Steven; Kew, Olen M.; Pallansch, Mark A.

    2003-01-01

    Type 1 wild-vaccine recombinant polioviruses sharing a 367-nucleotide (nt) block of Sabin 1-derived sequence spanning the VP1 and 2A genes circulated widely in China from 1991 to 1993. We surveyed the sequence relationships among 34 wild-vaccine recombinants by comparing six genomic intervals: the conserved 5′-untranslated region (5′-UTR) (nt 186 to 639), the hypervariable portion of the 5′-UTR (nt 640 to 742), the VP4 and partial VP2 genes (nt 743 to 1176), the VP1 gene (nt 2480 to 3385), the 2A gene (nt 3386 to 3832), and the partial 3D gene (nt 6011 to 6544). The 5′-UTR, capsid (VP4-VP2 and VP1), and 2A sequence intervals had similar phylogenies. By contrast, the partial 3D sequences could be distributed into five divergent genetic classes. Most (25 of 34) of the wild-vaccine recombinant isolates showed no evidence of additional recombination beyond the initial wild-Sabin recombination event. Eight isolates from 1992 to 1993, however, appear to be derived from three independent additional recombination events, and one 1993 isolate was derived from two consecutive events. Complete genomic sequences of a representative isolate for each 3D sequence class demonstrated that these exchanges had occurred in the 2B, 2C, and 3D genes. The 3D gene sequences were not closely related to those of the Sabin strains or 53 diverse contemporary wild poliovirus isolates from China, but all were related to the 3D genes of species C enteroviruses. The appearance within approximately 2.5 years of five recombinant classes derived from a single ancestral infection illustrates the rapid emergence of new recombinants among circulating wild polioviruses. PMID:14512548

  1. Production and characterization of recombinant lignin peroxidase isozyme H2 from Phanerochaete chrysosporium using recombinant baculovirus.

    PubMed

    Johnson, T M; Pease, E A; Li, J K; Tien, M

    1992-08-01

    Recombinant Phanerochaete chrysosporium lignin peroxidase isozyme H2 (pI 4.4) was produced in insect cells infected with a genetically engineered baculovirus containing a copy of the cDNA clone lambda ML-6. The recombinant enzyme was purified to near homogeneity and is capable of oxidizing veratryl alcohol, iodide, and, to a lesser extent, guaiacol. The Km of the recombinant enzyme for veratryl alcohol and H2O2 is similar to that of the fungal enzyme. The guaiacol oxidation activity or any other activity is not dependent upon Mn2+. The purified recombinant peroxidase is glycosylated with N-linked carbohydrate(s). The recombinant lignin peroxidase eluted from an anion exchange resin similar to that of native isozyme H1 rather than H2. However, the pI of the recombinant enzymes is different from both H1 and H2 isozymes. Further characterization of native isozymes H1 and H2 from the fungal cultures revealed identical N-terminus residues. This indicates that isozymes H1 and H2 differ in post-translational modification. PMID:1632652

  2. An improved recombineering approach by adding RecA to lambda Red recombination.

    PubMed

    Wang, Junping; Sarov, Mihail; Rientjes, Jeanette; Fu, Jun; Hollak, Heike; Kranz, Harald; Xie, Wei; Stewart, A Francis; Zhang, Youming

    2006-01-01

    Recombineering is the use of homologous recombination in Escherichia coli for DNA engineering. Of several approaches, use of the lambda phage Red operon is emerging as the most reliable and flexible. The Red operon includes three components: Redalpha, a 5' to 3' exonuclease, Redbeta, an annealing protein, and Redgamma, an inhibitor of the major E. coli exonuclease and recombination complex, RecBCD. Most E. coli cloning hosts are recA deficient to eliminate recombination and therefore enhance the stability of cloned DNAs. However, loss of RecA also impairs general cellular integrity. Here we report that transient RecA co-expression enhances the total number of successful recombinations in bacterial artificial chromosomes (BACs), mostly because the E. coli host is more able to survive the stresses of DNA transformation procedures. We combined this practical improvement with the advantages of a temperature-sensitive version of the low copy pSC101 plasmid to develop a protocol that is convenient and more efficient than any recombineering procedure, for use of either double- or single-stranded DNA, published to date. PMID:16382181

  3. Molecular hydrogen in the cosmic recombination epoch

    SciTech Connect

    Alizadeh, Esfandiar; Hirata, Christopher M.

    2011-10-15

    The advent of precise measurements of the CMB anisotropies has motivated correspondingly precise calculations of the cosmic recombination history. Cosmic recombination proceeds far out of equilibrium because of a ''bottleneck'' at the n=2 level of hydrogen: atoms can only reach the ground state via slow processes--two-photon decay or Lyman-{alpha} resonance escape. However, even a small primordial abundance of molecules could have a large effect on the interline opacity in the recombination epoch and lead to an additional route for hydrogen recombination. Therefore, this paper computes the abundance of the H{sub 2} molecule during the cosmic recombination epoch. Hydrogen molecules in the ground electronic levels X{sup 1}{Sigma}{sub g}{sup +} can either form from the excited H{sub 2} electronic levels B{sup 1}{Sigma}{sub u}{sup +} and C{sup 1}{Pi}{sub u} or through the charged particles H{sub 2}{sup +}, HeH{sup +}, and H{sup -}. We follow the transitions among all of these species, resolving the rotational and vibrational sublevels. Since the energies of the X{sup 1}{Sigma}{sub g}{sup +}-B{sup 1}{Sigma}{sub u}{sup +} (Lyman band) and X{sup 1}{Sigma}{sub g}{sup +}-C{sup 1}{Pi}{sub u} (Werner band) transitions are near the Lyman-{alpha} energy, the distortion of the CMB spectrum caused by escaped H Lyman-line photons accelerates both the formation and the destruction of H{sub 2} due to this channel relative to the thermal rates. This causes the populations of H{sub 2} molecules in X{sup 1}{Sigma}{sub g}{sup +} energy levels to deviate from their thermal equilibrium abundances. We find that the resulting H{sub 2} abundance is 10{sup -17} at z=1200 and 10{sup -13} at z=800, which is too small to have any significant influence on the recombination history.

  4. The Unconventional Xer Recombination Machinery of Streptococci/Lactococci

    PubMed Central

    Le Bourgeois, Pascal; Bugarel, Marie; Campo, Nathalie; Daveran-Mingot, Marie-Line; Labonté, Jessica; Lanfranchi, Daniel; Lautier, Thomas; Pagès, Carine; Ritzenthaler, Paul

    2007-01-01

    Homologous recombination between circular sister chromosomes during DNA replication in bacteria can generate chromosome dimers that must be resolved into monomers prior to cell division. In Escherichia coli, dimer resolution is achieved by site-specific recombination, Xer recombination, involving two paralogous tyrosine recombinases, XerC and XerD, and a 28-bp recombination site (dif) located at the junction of the two replication arms. Xer recombination is tightly controlled by the septal protein FtsK. XerCD recombinases and FtsK are found on most sequenced eubacterial genomes, suggesting that the Xer recombination system as described in E. coli is highly conserved among prokaryotes. We show here that Streptococci and Lactococci carry an alternative Xer recombination machinery, organized in a single recombination module. This corresponds to an atypical 31-bp recombination site (difSL) associated with a dedicated tyrosine recombinase (XerS). In contrast to the E. coli Xer system, only a single recombinase is required to recombine difSL, suggesting a different mechanism in the recombination process. Despite this important difference, XerS can only perform efficient recombination when difSL sites are located on chromosome dimers. Moreover, the XerS/difSL recombination requires the streptococcal protein FtsKSL, probably without the need for direct protein-protein interaction, which we demonstrated to be located at the division septum of Lactococcus lactis. Acquisition of the XerS recombination module can be considered as a landmark of the separation of Streptococci/Lactococci from other firmicutes and support the view that Xer recombination is a conserved cellular function in bacteria, but that can be achieved by functional analogs. PMID:17630835

  5. Using Crossover Breakpoints in Recombinant Inbred Lines to Identify Quantitative Trait Loci Controlling the Global Recombination Frequency

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Recombination is a crucial component of evolution and breeding, producing new genetic combinations on which selection can act. Rates of recombination vary tremendously, not only between species but also within species and for specific chromosomal segments. In this study, by examining recombination...

  6. Recombination energy in double white dwarf formation

    NASA Astrophysics Data System (ADS)

    Nandez, J. L. A.; Ivanova, N.; Lombardi, J. C.

    2015-06-01

    In this Letter, we investigate the role of recombination energy during a common envelope event. We confirm that taking this energy into account helps to avoid the formation of the circumbinary envelope commonly found in previous studies. For the first time, we can model a complete common envelope event, with a clean compact double white dwarf binary system formed at the end. The resulting binary orbit is almost perfectly circular. In addition to considering recombination energy, we also show that between 1/4 and 1/2 of the released orbital energy is taken away by the ejected material. We apply this new method to the case of the double white dwarf system WD 1101+364, and we find that the progenitor system at the start of the common envelope event consisted of an ˜1.5 M⊙ red giant star in an ˜30 d orbit with a white dwarf companion.

  7. The evolution of sex dimorphism in recombination.

    PubMed Central

    Lenormand, Thomas

    2003-01-01

    Sex dimorphism in recombination is widespread on both sex chromosomes and autosomes. Various hypotheses have been proposed to explain these dimorphisms. Yet no theoretical model has been explored to determine how heterochiasmy--the autosomal dimorphism--could evolve. The model presented here shows three circumstances in which heterochiasmy is likely to evolve: (i) a male-female difference in haploid epistasis, (ii) a male-female difference in cis-epistasis minus trans-epistasis in diploids, or (iii) a difference in epistasis between combinations of genes inherited maternally or paternally. These results hold even if sources of linkage disequilibria besides epistasis, such as migration or Hill-Robertson interference, are considered and shed light on previous verbal models of sex dimorphism in recombination rates. Intriguingly, these results may also explain why imprinted regions on the autosomes of humans or sheep are particularly heterochiasmate. PMID:12618416

  8. Recombinant laccase: I. Enzyme cloning and characterization.

    PubMed

    Nicolini, Claudio; Bruzzese, Debora; Cambria, Maria Teresa; Bragazzi, Nicola Luigi; Pechkova, Eugenia

    2013-03-01

    We obtained structural and functional characterization of a recombinant Laccase from Rigidoporus lignosus (formerly Rigidoporus microporus), a white-rot basidiomycete, by means of circular dichroism (CD) spectra, cyclic voltammetry (CV) and biochemical assays. Here we report the optimization of expression and purification procedures of a recombinant Laccase expressed in supercompetent Escherichia coli cells. We amplified the coding sequence of Laccase using PCR from cDNA and cloned into a bacterial expression system. The resulting expression plasmid, pET-28b, was under a strong T7/Lac promoter induced by IPTG (isopropyl-β-d-thiogalactoipyranoside). We obtained purification by fast protein liquid chromatography (FPLC) method. We recorded the variation of the current of a solution containing purified Laccase with increasing Syringaldazine (SGZ) concentration using a potentiometer as proof of principle, showing its compatibility with the development of a new enzymatic biosensor for medical purposes, as described in Part II. PMID:22991171

  9. Recombination clumping factor during cosmic reionization

    SciTech Connect

    Kaurov, Alexander A.; Gnedin, Nickolay Y. E-mail: gnedin@fnal.gov

    2014-06-01

    We discuss the role of recombinations in the intergalactic medium, and the related concept of the clumping factor, during cosmic reionization. The clumping factor is, in general, a local quantity that depends on both the local overdensity and the scale below which the baryon density field can be assumed smooth. That scale, called the filtering scale, depends on over-density and local thermal history. We present a method for building a self-consistent analytical model of inhomogeneous reionization, assuming the linear growth rate of the density fluctuation, which simultaneously accounts for these effects. We show that taking into account the local clumping factor introduces significant corrections to the total recombination rate, compared to the model with a globally uniform clumping factor.

  10. Recombinant organisms for production of industrial products

    PubMed Central

    Adrio, Jose-Luis

    2010-01-01

    A revolution in industrial microbiology was sparked by the discoveries of ther double-stranded structure of DNA and the development of recombinant DNA technology. Traditional industrial microbiology was merged with molecular biology to yield improved recombinant processes for the industrial production of primary and secondary metabolites, protein biopharmaceuticals and industrial enzymes. Novel genetic techniques such as metabolic engineering, combinatorial biosynthesis and molecular breeding techniques and their modifications are contributing greatly to the development of improved industrial processes. In addition, functional genomics, proteomics and metabolomics are being exploited for the discovery of novel valuable small molecules for medicine as well as enzymes for catalysis. The sequencing of industrial microbal genomes is being carried out which bodes well for future process improvement and discovery of new industrial products. PMID:21326937

  11. Recombineering BAC transgenes for protein tagging.

    PubMed

    Ciotta, Giovanni; Hofemeister, Helmut; Maresca, Marcello; Fu, Jun; Sarov, Mihail; Anastassiadis, Konstantinos; Stewart, A Francis

    2011-02-01

    Protein tagging offers many advantages for proteomic and regulomic research, particularly due to the use of generic and highly sensitive methods that can be applied with reasonable throughput. Ideally, protein tagging is equivalent to having a high affinity antibody for every chosen protein. However, these advantages are compromised if the tagged protein is overexpressed, which is usually the case from cDNA expression vectors. BAC (bacterial artificial chromosome) transgenes present a way to express a chosen protein at physiological levels with all regulatory elements in their native configurations, including cell cycle, alternative splicing and microRNA regulation. Recombineering has become the method of choice for modifying large constructs like BACs. Here, we present a method for protein tagging by recombineering BACs, transfecting cells and evaluating tagged protein expression. PMID:20868752

  12. Hα diagnostic in a recombining plasma

    NASA Astrophysics Data System (ADS)

    Wenzel, U.; Goto, M.

    2016-05-01

    In fusion devices the hydrogen Balmer lines are used to measure the neutral flux from the walls into the plasma using the atomic physics factor S/XB. This is a standard diagnostic which can be applied in ionizing plasma using {{H}α} , {{H}β} or {{H}γ} without knowledge of the electron density. We will extend this method to a recombining plasma in front of a surface. {{H}α} can be used in an analogous way to measure the plasma flow to this surface which can be e.g. a divertor target. The other Balmer lines are not suitable because the corresponding atomic physics factor R/YB depends on density due to three-body recombination. An application of this diagnostic method is provided.

  13. Dissipative Stern-Gerlach recombination experiment

    SciTech Connect

    Oliveira, Thiago R. de; Caldeira, A. O.

    2006-04-15

    The possibility of obtaining the initial pure state in a usual Stern-Gerlach experiment through the recombination of the two emerging beams is investigated. We have extended the previous work of Englert, Schwinger, and Scully [Found Phys. 18, 1045 (1988)] including the fluctuations of the magnetic field generated by a properly chosen magnet. As a result we obtained an attenuation factor to the possible revival of coherence when the beams are perfectly recombined. When the source of the magnetic field is a superconducting quantum interference device (SQUID) the attenuation factor can be controlled by external circuits and the spin decoherence directly measured. For the proposed SQUID with dimensions in the scale of microns the attenuation factor has been shown unimportant when compared with the interaction time of the spin with the magnet.

  14. Designing recombinant Pseudomonas strains to enhance biodesulfurization.

    PubMed Central

    Gallardo, M E; Ferrández, A; De Lorenzo, V; García, J L; Díaz, E

    1997-01-01

    The dsz biodesulfurization cluster from Rhodococcus erythropolis IGTS8 has been engineered under the control of heterologous broad-host-range regulatory signals to alleviate the mechanism of sulfur repression, and it was stably inserted into the chromosomes of different Pseudomonas strains. The recombinant bacteria were able to desulfurize dibenzothiophene more efficiently than the native host. Furthermore, these new biocatalysts combine relevant industrial and environmental traits, such as production of biosurfactants, with the enhanced biodesulfurization phenotype. PMID:9371464

  15. Recombinant organisms capable of fermenting cellobiose

    DOEpatents

    Ingram, Lonnie O.; Lai, Xiaokuang; Moniruzzaman, Mohammed; York, Sean W.

    2000-01-01

    This invention relates to a recombinant microorganism which expresses pyruvate decarboxylase, alcohol dehydrogenase, Klebsiella phospho-.beta.-glucosidase and Klebsiella (phosphoenolpyruvate-dependent phosphotransferase system) cellobiose-utilizing Enzyme II, wherein said phospho-.beta.-glucosidase and said (phosphoenolpyruvate-dependent phosphotransferase) cellobiose-utilizing Enzyme II are heterologous to said microorganism and wherein said microorganism is capable of utilizing both hemicellulose and cellulose, including cellobiose, in the production of ethanol.

  16. Nonradiative Auger recombination in semiconductor nanocrystals.

    PubMed

    Vaxenburg, Roman; Rodina, Anna; Shabaev, Andrew; Lifshitz, Efrat; Efros, Alexander L

    2015-03-11

    We calculate the rate of nonradiative Auger recombination in negatively charged CdSe nanocrystals (NCs). The rate is nonmonotonic, strongly oscillating with NC size, and sensitive to the NC surface. The oscillations result in nonexponential decay of carriers in NC ensembles. Using a standard single-exponential approximation of the decay dynamics, we determine the apparent size dependence of the Auger rate in an ensemble and derive CdSe surface parameters consistent with the experimental dependence on size. PMID:25693512

  17. Mechanism and regulation of meiotic recombination initiation

    PubMed Central

    Lam, Isabel; Keeney, Scott

    2015-01-01

    Meiotic recombination involves the formation and repair of programmed DNA double-strand breaks (DSBs) catalyzed by the conserved Spo11 protein. This review summarizes recent studies pertaining to the formation of meiotic DSBs, including the mechanism of DNA cleavage by Spo11, proteins required for break formation, and mechanisms that control the location, timing, and number of DSBs. Where appropriate, findings in different organisms are discussed to highlight evolutionary conservation or divergence. PMID:25324213

  18. Development of Mycoplasma hyopneumoniae Recombinant Vaccines.

    PubMed

    Marchioro, Silvana Beutinger; Simionatto, Simone; Dellagostin, Odir

    2016-01-01

    Mycoplasma hyopneumoniae is the etiological agent of swine enzootic pneumonia (EP), a disease that affects swine production worldwide. Vaccination is the most cost-effective strategy for the control and prevention of the disease. Research using genome-based approach has the potential to elucidate the biology and pathogenesis of M. hyopneumoniae and contribute to the development of more effective vaccines. Here, we describe the protocol for developing M. hyopneumoniae recombinant vaccines using reverse vaccinology approaches. PMID:27076288

  19. Cultivating Insect Cells To Produce Recombinant Proteins

    NASA Technical Reports Server (NTRS)

    Spaulding, Glenn; Goodwin, Thomas; Prewett, Tacey; Andrews, Angela; Francis, Karen; O'Connor, Kim

    1996-01-01

    Method of producing recombinant proteins involves growth of insect cells in nutrient solution in cylindrical bioreactor rotating about cylindrical axis, oriented horizontally and infecting cells with viruses into which genes of selected type cloned. Genes in question those encoding production of desired proteins. Horizontal rotating bioreactor preferred for use in method, denoted by acronym "HARV", described in "High-Aspect-Ratio Rotating Cell-Culture Vessel" (MSC-21662).

  20. Three-Body Recombination of Oxygen Atoms

    NASA Astrophysics Data System (ADS)

    Huestis, D. L.; Kalogerakis, K. S.

    2002-05-01

    Dayside photodissociation of O2 and CO2 in the atmospheres of Earth, Venus, and Mars produces oxygen atoms that eventually undergo three-body recombination O + O + M -> O2* + M The variety of electronic states produced is observable as nightglow emissions, which have been the subject of many laboratory and interpretive investigations. Here we review the current understanding of the overall temperature-dependent rate coefficient for three-body recombination of oxygen atoms and describe a strategy for its measurement. The most recent measurement [1] is almost 30 years old. The most comprehensive review [2] is more than 25 years old and shows that the absolute rate coefficients for recombination and the reverse process, collision-induced dissociation, as well as the dependence on temperature and collider, are poorly determined, in spite of the relatively narrow error bars reported in the various studies. The most recent high-temperature dissociation study [3] actually increases the divergence. We plan experiments with a commercial F2 laser, providing roughly 50 mJ of 157 nm radiation in a 3-4 mm beam, to achieve greater than 80% dissociation of molecular oxygen, in the range from 0.5 to 5 torr. In a high-pressure N2 background (30-200 torr) the oxygen atoms will recombine in a time scale from 0.1 to 10 ms, as monitored by 845 nm fluorescence excited by two photons at 226 nm. [1] I. M. Campbell and C. N. Gray, Chem. Phys. Lett. 18, 607 (1973). [2] D. L. Baulch, D. D. Drysdale, J. Duxbury, and S. J. Grant, Evaluated Kinetic Data for High Temperature Reactions Vol. 3 ``Homogeneous Gas Phase Reactions of the O2--O3 System, the CO--O2--H2 System, and of Sulphur-Containing Species," (Butterworths, London, 1976). [3] V. Naudet, S. Abid, and C. E. Paillard, J. Chim. Phys. 96, 1123 (1999).

  1. Ion recombination in aircraft exhaust plumes

    NASA Astrophysics Data System (ADS)

    Sorokin, A.; Mirabel, P.

    In this article, a model which examines the evolution of ion concentrations in a hot aircraft exhaust plume on the ground is proposed. The model includes plume dilution and ion-ion recombination with coefficients which vary with temperature. A comparison of the model is made with the available ground-based experimental data obtained on the ATTAS research aircraft engines. From this comparison, an ion emission index of the order of 8 1016 ions/kg(fuel) inferred.

  2. [Homologous recombination among bacterial genomes: the measurement and identification].

    PubMed

    Xianwei, Yang; Ruifu, Yang; Yujun, Cui

    2016-02-01

    Homologous recombination is one of important sources in shaping the bacterial population diversity, which disrupts the clonal relationship among different lineages through horizontal transferring of DNA-segments. As consequence of blurring the vertical inheritance signals, the homologous recombination raises difficulties in phylogenetic analysis and reconstruction of population structure. Here we discuss the impacts of homologous recombination in inferring phylogenetic relationship among bacterial isolates, and summarize the tools and models separately used in recombination measurement and identification. We also highlight the merits and drawbacks of various approaches, aiming to assist in the practical application for the analysis of homologous recombination in bacterial evolution research. PMID:26907777

  3. RECOMBINATION RATE COEFFICIENTS OF Be-LIKE Si

    SciTech Connect

    Orban, I.; Boehm, S.; Schuch, R.; Loch, S. D.

    2010-10-01

    Recombination of Be-like Si{sup 10+} over the 0-43 eV electron-ion energy range is measured at the CRYRING electron cooler. In addition to radiative and dielectronic recombination, the recombination spectrum also shows strong contributions from trielectronic recombination. Below 100 meV, several very strong resonances associated with a spin-flip of the excited electron dominate the spectrum and also dominate the recombination in the photoionized plasma. The resonant plasma rate coefficients corrected for the experimental field ionization are in good agreement with calculated results by Gu and with AUTOSTRUCTURE calculations. All other calculations significantly underestimate the plasma rate coefficients at low temperatures.

  4. Radiative recombination of hot carriers in narrow-gap semiconductors

    SciTech Connect

    Pavlov, N. V.; Zegrya, G. G.

    2012-01-15

    The mechanism of the radiative recombination of hot carriers in narrow-gap semiconductors is analyzed using the example of indium antimonide. It is shown that the CHCC Auger recombination process may lead to pronounced carrier heating at high excitation levels. The distribution functions and concentrations of hot carriers are determined. The radiative recombination rate of hot carriers and the radiation gain coefficient are calculated in terms of the Kane model. It is demonstrated that the radiative recombination of hot carriers will make a substantial contribution to the total radiative recombination rate at high carrier concentrations.

  5. Recombination between linear double-stranded DNA substrates in vivo

    PubMed Central

    Narayanan, Kumaran; Sim, Edmund Ui-Hang; Ravin, Nikolai V.; Lee, Choon-Weng

    2009-01-01

    Recombineering technology in E. coli enables targeting of linear donor DNA to circular recipient DNA using short shared homology sequences. In this work, we demonstrate that recombineering is also able to support recombination between a pair of linear DNA substrates (linear/linear recombineering) in vivo in E. coli. Linear DNA up to 100 kb is accurately modified and remains intact without undergoing rearrangements after recombination. This system will be valuable for direct in vivo manipulation of large linear DNA including the N15 and PY54 prophages and linear animal viruses, and for assembly of linear constructs as artificial chromosome vectors. PMID:19454252

  6. Recombinant Origin of the Retrovirus XMRV

    PubMed Central

    Paprotka, Tobias; Delviks-Frankenberry, Krista A.; Cingöz, Oya; Martinez, Anthony; Kung, Hsing-Jien; Tepper, Clifford G.; Hu, Wei-Shau; Fivash, Matthew J.; Coffin, John M.; Pathak, Vinay K.

    2012-01-01

    The retrovirus XMRV (xenotropic murine leukemia virus-related virus) has been detected in human prostate tumors and in blood samples from patients with chronic fatigue syndrome, but these findings have not been replicated. We hypothesized that an understanding of when and how XMRV first arose might help explain the discrepant results. We studied human prostate cancer cell lines CWR22Rv1 and CWR-R1, which produce XMRV virtually identical to the viruses recently found in patient samples, as well as their progenitor human prostate tumor xenograft (CWR22) that had been passaged in mice. We detected XMRV infection in the two cell lines and in the later passage xenografts, but not in the early passages. Importantly, we found that the host mice contained two proviruses, PreXMRV-1 and PreXMRV-2, which share 99.92% identity with XMRV over >3.2-kb stretches of their genomes. We conclude that XMRV was not present in the original CWR22 tumor but was generated by recombination of two proviruses during tumor passaging in mice. The probability that an identical recombinant was generated independently is negligible (~10-12); our results suggest that the association of XMRV with human disease is due to contamination of human samples with virus originating from this recombination event. PMID:21628392

  7. Production of recombinant allergens in plants

    PubMed Central

    2010-01-01

    A large percentage of allergenic proteins are of plant origin. Hence, plant-based expression systems are considered ideal for the recombinant production of certain allergens. First attempts to establish production of plant-derived allergens in plants focused on transient expression in Nicotiana benthamiana infected with recombinant viral vectors. Accordingly, allergens from birch and mugwort pollen, as well as from apple have been expressed in plants. Production of house dust mite allergens has been achieved by Agrobacterium-mediated transformation of tobacco plants. Beside the use of plants as production systems, other approaches have focused on the development of edible vaccines expressing allergens or epitopes thereof, which bypasses the need of allergen purification. The potential of this approach has been convincingly demonstrated for transgenic rice seeds expressing seven dominant human T cell epitopes derived from Japanese cedar pollen allergens. Parallel to efforts in developing recombinant-based diagnostic and therapeutic reagents, different gene-silencing approaches have been used to decrease the expression of allergenic proteins in allergen sources. In this way hypoallergenic ryegrass, soybean, rice, apple, and tomato were developed. PMID:21258627

  8. Dissociative recombination of molecular ions with electrons

    NASA Technical Reports Server (NTRS)

    Johnsen, Rainer

    1990-01-01

    An overview is presented for the present state of the art of laboratory measurements of the dissociative recombination of molecular ions with electrons. Most work has focussed on obtaining rates and their temperature dependence, as these are of primary interest for model calculations of ionospheres. A comparison of data obtained using the microwave afterglow method, the flowing afterglow technique, and the merged beam technique shows that generally the agreement is quite good, but there are some serious discrepancies, especially in the case of H(3+) recombination, that need to be resolved. Results of some earlier experimental work need to be reexamined in the light of more recent developments. Such cases are pointed out and a compilation of rate coefficients that have withstood scrutiny is presented. Recent advances in experimental methods, such as the use of laser-in-duced fluorescence, make it possible to identify some neutral products of dissociative recombination. What has been done so far and what results one might expect from future work are briefly reviewed.

  9. Recombinant vaccine for canine parvovirus in dogs.

    PubMed

    López de Turiso, J A; Cortés, E; Martínez, C; Ruiz de Ybáñez, R; Simarro, I; Vela, C; Casal, I

    1992-05-01

    VP2 is the major component of canine parvovirus (CPV) capsids. The VP2-coding gene was engineered to be expressed by a recombinant baculovirus under the control of the polyhedrin promoter. A transfer vector that contains the lacZ gene under the control of the p10 promoter was used in order to facilitate the selection of recombinants. The expressed VP2 was found to be structurally and immunologically indistinguishable from authentic VP2. The recombinant VP2 shows also the capability to self-assemble, forming viruslike particles similar in size and appearance to CPV virions. These viruslike particles have been used to immunize dogs in different doses and combinations of adjuvants, and the anti-CPV responses have been measured by enzyme-linked immunosorbent assay, monolayer protection assays, and an assay for the inhibition of hemagglutination. A dose of ca. 10 micrograms of VP2 was able to elicit a good protective response, higher than that obtained with a commercially available, inactivated vaccine. The results indicate that these viruslike particles can be used to protect dogs from CPV infection. PMID:1313899

  10. FASEB Summer Research Conference. Genetic Recombination and Chromosome Rearrangements

    SciTech Connect

    Jinks-Robertson, Sue

    2002-02-01

    The 2001 meeting entitled ''Genetic Recombination and Genome Rearrangements'' was held July 21-26 in Snowmass, Colorado. The goal of the meeting was to bring together scientists using diverse approaches to study all aspects of genetic recombination. This goal was achieved by integrating talks covering the genetics, biochemistry and structural biology of homologous recombination, site-specific recombination, and nonhomologous recombination. The format of the meeting consisted of a keynote address on the opening evening, two formal plenary sessions on each of the four full meeting days, a single afternoon workshop consisting of short talks chosen from among submitted abstracts, and afternoon poster sessions on each of the four full meeting days. The eight plenary session were entitled: (1) Recombination Mechanisms, (2) Prokaryotic Recombination, (3) Repair and Recombination, (4) Site-specific Recombination and Transposition, (5) Eukaryotic Recombination I, (6) Genome Rearrangements, (7) Meiosis, and (8) Eukaryotic Recombination II. Each session included a mix of genetic, biochemical and structural talks; talks were limited to 20 minutes, followed by 10 minutes of very lively, general discussion. Much of the data presented in the plenary sessions was unpublished, thus providing attendees with the most up-to-date knowledge of this rapidly-moving field.

  11. Recombination Pattern Reanalysis of Some HIV-1 Circulating Recombination Forms Suggest the Necessity and Difficulty of Revision

    PubMed Central

    Jia, Lei; Li, Lin; Li, Hanping; Liu, Siyang; Wang, Xiaolin; Bao, Zuoyi; Li, Tianyi; Zhuang, Daomin; Liu, Yongjian; Li, Jingyun

    2014-01-01

    Background Recombination is one of the major mechanisms underlying the generation of HIV-1 variability. Currently 61 circulating recombinant forms of HIV-1 have been identified. With the development of recombination detection techniques and accumulation of HIV-1 reference stains, more accurate mosaic structures of circulating recombinant forms (CRFs), like CRF04 and CRF06, have undergone repeated analysis and upgrades. Such revisions may also be necessary for other CRFs. Unlike previous studies, whose results are based primarily on a single recombination detection program, the current study was based on multiple recombination analysis, which may have produced more impartial results. Methods Representative references of 3 categories of intersubtype recombinants were selected, including BC recombinants (CRF07 and CRF08), BG recombinants (CRF23 and CRF24), and BF recombinants (CRF38 and CRF44). They were reanalyzed in detail using both the jumping profile hidden Markov model and RDP3. Results The results indicate that revisions and upgrades are very necessary and the entire re-analysis suggested 2 types of revision: (i) length of inserted fragments; and (ii) number of inserted fragments. The reanalysis also indicated that determination of small regions of about 200 bases or fewer should be performed with more caution. Conclusion Results indicated that the involvement of multiple recombination detection programs is very necessary. Additionally, results suggested two major challenges, one involving the difficulty of accurately determining the locations of breakpoints and the second involving identification of small regions of about 200 bases or fewer with greater caution. Both indicate the complexity of HIV-1 recombination. The resolution would depend critically on development of a recombination analysis algorithm, accumulation of HIV-1 stains, and a higher sequencing quality. With the changes in recombination pattern, phylogenetic relationships of some CRFs may also

  12. Mechanisms and Factors that Influence High Frequency Retroviral Recombination

    PubMed Central

    Delviks-Frankenberry, Krista; Galli, Andrea; Nikolaitchik, Olga; Mens, Helene; Pathak, Vinay K.; Hu, Wei-Shau

    2011-01-01

    With constantly changing environmental selection pressures, retroviruses rely upon recombination to reassort polymorphisms in their genomes and increase genetic diversity, which improves the chances for the survival of their population. Recombination occurs during DNA synthesis, whereby reverse transcriptase undergoes template switching events between the two copackaged RNAs, resulting in a viral recombinant with portions of the genetic information from each parental RNA. This review summarizes our current understanding of the factors and mechanisms influencing retroviral recombination, fidelity of the recombination process, and evaluates the subsequent viral diversity and fitness of the progeny recombinant. Specifically, the high mutation rates and high recombination frequencies of HIV-1 will be analyzed for their roles in influencing HIV-1 global diversity, as well as HIV-1 diagnosis, drug treatment, and vaccine development. PMID:21994801

  13. Evolution of recombination in eutherian mammals: insights into mechanisms that affect recombination rates and crossover interference

    PubMed Central

    Segura, Joana; Ferretti, Luca; Ramos-Onsins, Sebastián; Capilla, Laia; Farré, Marta; Reis, Fernanda; Oliver-Bonet, Maria; Fernández-Bellón, Hugo; Garcia, Francisca; Garcia-Caldés, Montserrat; Robinson, Terence J.; Ruiz-Herrera, Aurora

    2013-01-01

    Recombination allows faithful chromosomal segregation during meiosis and contributes to the production of new heritable allelic variants that are essential for the maintenance of genetic diversity. Therefore, an appreciation of how this variation is created and maintained is of critical importance to our understanding of biodiversity and evolutionary change. Here, we analysed the recombination features from species representing the major eutherian taxonomic groups Afrotheria, Rodentia, Primates and Carnivora to better understand the dynamics of mammalian recombination. Our results suggest a phylogenetic component in recombination rates (RRs), which appears to be directional, strongly punctuated and subject to selection. Species that diversified earlier in the evolutionary tree have lower RRs than those from more derived phylogenetic branches. Furthermore, chromosome-specific recombination maps in distantly related taxa show that crossover interference is especially weak in the species with highest RRs detected thus far, the tiger. This is the first example of a mammalian species exhibiting such low levels of crossover interference, highlighting the uniqueness of this species and its relevance for the study of the mechanisms controlling crossover formation, distribution and resolution. PMID:24068360

  14. Recombinant pinoresinol/lariciresinol reductase, recombinant dirigent protein, and methods of use

    DOEpatents

    Lewis, Norman G.; Davin, Laurence B.; Dinkova-Kostova, Albena T.; Fujita, Masayuki; Gang, David R.; Sarkanen, Simo; Ford, Joshua D.

    2001-04-03

    Dirigent proteins and pinoresinol/lariciresinol reductases have been isolated, together with cDNAs encoding dirigent proteins and pinoresinol/lariciresinol reductases. Accordingly, isolated DNA sequences are provided which code for the expression of dirigent proteins and pinoresinol/lariciresinol reductases. In other aspects, replicable recombinant cloning vehicles are provided which code for dirigent proteins or pinoresinol/lariciresinol reductases or for a base sequence sufficiently complementary to at least a portion of dirigent protein or pinoresinol/lariciresinol reductase DNA or RNA to enable hybridization therewith. In yet other aspects, modified host cells are provided that have been transformed, transfected, infected and/or injected with a recombinant cloning vehicle and/or DNA sequence encoding dirigent protein or pinoresinol/lariciresinol reductase. Thus, systems and methods are provided for the recombinant expression of dirigent proteins and/or pinoresinol/lariciresinol reductases.

  15. The imperfect ancestral recombination graph reconstruction problem: upper bounds for recombination and homoplasy.

    PubMed

    Lam, Fumei; Tarpine, Ryan; Istrail, Sorin

    2010-06-01

    One of the central problems in computational biology is the reconstruction of evolutionary histories. While models incorporating recombination and homoplasy have been studied separately, a missing component in the theory is a robust and flexible unifying model which incorporates both of these major biological events shaping genetic diversity. In this article, we introduce the first such unifying model and develop algorithms to find the optimal ancestral recombination graph incorporating recombinations and homoplasy events. The power of our framework is the connection between our formulation and the Directed Steiner Arborescence Problem in combinatorial optimization. We implement linear programming techniques as well as heuristics for the Directed Steiner Arborescence Problem, and use our methods to construct evolutionary histories for both simulated and real data sets. PMID:20583925

  16. Late replicating domains are highly recombining in females but have low male recombination rates: implications for isochore evolution.

    PubMed

    Pink, Catherine J; Hurst, Laurence D

    2011-01-01

    In mammals sequences that are either late replicating or highly recombining have high rates of evolution at putatively neutral sites. As early replicating domains and highly recombining domains both tend to be GC rich we a priori expect these two variables to covary. If so, the relative contribution of either of these variables to the local neutral substitution rate might have been wrongly estimated owing to covariance with the other. Against our expectations, we find that sex-averaged recombination rates show little or no correlation with replication timing, suggesting that they are independent determinants of substitution rates. However, this result masks significant sex-specific complexity: late replicating domains tend to have high recombination rates in females but low recombination rates in males. That these trends are antagonistic explains why sex-averaged recombination is not correlated with replication timing. This unexpected result has several important implications. First, although both male and female recombination rates covary significantly with intronic substitution rates, the magnitude of this correlation is moderately underestimated for male recombination and slightly overestimated for female recombination, owing to covariance with replicating timing. Second, the result could explain why male recombination is strongly correlated with GC content but female recombination is not. If to explain the correlation between GC content and replication timing we suppose that late replication forces reduced GC content, then GC promotion by biased gene conversion during female recombination is partly countered by the antagonistic effect of later replicating sequence tending increase AT content. Indeed, the strength of the correlation between female recombination rate and local GC content is more than doubled by control for replication timing. Our results underpin the need to consider sex-specific recombination rates and potential covariates in analysis of GC

  17. Same-period emission and recombination in nonsequential double-recombination high-order-harmonic generation

    NASA Astrophysics Data System (ADS)

    Hansen, Kenneth K.; Madsen, Lars Bojer

    2016-05-01

    Nonsequential double-recombination (NSDR) high-order-harmonic generation (HHG) is studied in a molecular model system. We observe a unique molecular two-electron effect with a characteristic cutoff in the HHG spectrum at higher energies than what was previously seen for NSDR HHG in atoms. The effect is corroborated with a classical model where it is found that the effect is sensitive to the molecular potential and originates from same-period emission and recombination (SPEAR) of two electrons. The effect persists for intermediate nuclear distances of R ≳8.0 a.u.

  18. Live-Cell Imaging of Vaccinia Virus Recombination

    PubMed Central

    Paszkowski, Patrick; Noyce, Ryan S.; Evans, David H.

    2016-01-01

    Recombination between co-infecting poxviruses provides an important mechanism for generating the genetic diversity that underpins evolution. However, poxviruses replicate in membrane-bound cytoplasmic structures known as factories or virosomes. These are enclosed structures that could impede DNA mixing between co-infecting viruses, and mixing would seem to be essential for this process. We hypothesize that virosome fusion events would be a prerequisite for recombination between co-infecting poxviruses, and this requirement could delay or limit viral recombination. We have engineered vaccinia virus (VACV) to express overlapping portions of mCherry fluorescent protein fused to a cro DNA-binding element. In cells also expressing an EGFP-cro fusion protein, this permits live tracking of virus DNA and genetic recombination using confocal microscopy. Our studies show that different types of recombination events exhibit different timing patterns, depending upon the relative locations of the recombining elements. Recombination between partly duplicated sequences is detected soon after post-replicative genes are expressed, as long as the reporter gene sequences are located in cis within an infecting genome. The same kinetics are also observed when the recombining elements are divided between VACV and transfected DNA. In contrast, recombination is delayed when the recombining sequences are located on different co-infecting viruses, and mature recombinants aren’t detected until well after late gene expression is well established. The delay supports the hypothesis that factories impede inter-viral recombination, but even after factories merge there remain further constraints limiting virus DNA mixing and recombinant gene assembly. This delay could be related to the continued presence of ER-derived membranes within the fused virosomes, membranes that may once have wrapped individual factories. PMID:27525721

  19. An introduction to recombination and linkage analysis

    SciTech Connect

    Mcpeek, M.S.

    1996-12-31

    With a garden as his laboratory, Mendel was able to discern basic probabilistic laws of heredity. Although it first appeared as a baffling exception to one of Mendel`s principles, the phenomenon of variable linkage between characters was soon recognized to be a powerful tool in the process of chromosome mapping and location of genes of interest. In this introduction, we first describe Mendel`s work and the subsequent discovery of linkage. Next we describe the apparent cause of variable linkage, namely recombination, and we introduce linkage analysis. 33 refs., 1 fig., 2 tabs.

  20. Cooling and recombination processes in cometary plasma

    NASA Technical Reports Server (NTRS)

    Wallis, M. K.; Ong, R. S. B.

    1976-01-01

    The ion electron plasma in comets is examined for cooling processes which result from its interactions with the neutral coma. A cometary coma model is formulated that is composed predominantly of H2O and its decomposition products where electrons are cooled in a variety of processes at rates varying with energy. It is shown that solar plasma plus accumulated cometary ions and electrons is affected very strongly as it flows into the coma. The electrons are rapidly cooled and all but some 10% of the ions undergo charge exchange. Photodissociation of H2O is assumed where ion electron recombination is the dominant loss process.

  1. Nanobodies and recombinant binders in cell biology

    PubMed Central

    Helma, Jonas; Cardoso, M. Cristina; Muyldermans, Serge

    2015-01-01

    Antibodies are key reagents to investigate cellular processes. The development of recombinant antibodies and binders derived from natural protein scaffolds has expanded traditional applications, such as immunofluorescence, binding arrays, and immunoprecipitation. In addition, their small size and high stability in ectopic environments have enabled their use in all areas of cell research, including structural biology, advanced microscopy, and intracellular expression. Understanding these novel reagents as genetic modules that can be integrated into cellular pathways opens up a broad experimental spectrum to monitor and manipulate cellular processes. PMID:26056137

  2. Dissociation and recombination in an inhomogeneous gas

    NASA Astrophysics Data System (ADS)

    Kuščer, Ivan

    1991-09-01

    The system of Boltzmann equations of Ludwig and Heil for a dissociating gas mixture is reviewed and reformulated in terms of differential cross sections. For the purpose of Monte Carlo simulations, collision models of the type of Borgnakke and Larsen are proposed. In case of short-lived transient flows of molecular gases and at not too high temperatures, simulation of scattering collisions is expected to suffice; the distribution function of the dissociated particles can be evaluated afterwards by integration, and recombination can be ignored.

  3. Engineering thermoacidophilic archaea using linear DNA recombination.

    PubMed

    Maezato, Yukari; Dana, Karl; Blum, Paul

    2011-01-01

    Thermoacidophilic archaea comprise one of the major classes of extremophiles. Most belong to the family Sulfolobales within the phylum Crenarchaeota. They are of applied interest as sources of hyperstable enzymes, for biomining of base and precious metals, and for evolutionary studies because of their use of eukaryotic-like subcellular mechanisms. Genetic methods are available for several species particularly Sulfolobus solfataricus. This organism has a considerable number of methods available for the construction of novel cell lines with unique functions. This chapter presents recent developments in the use of homologous recombination and linear DNA for the engineering of site-specific changes in the genome of S. solfataricus. PMID:21815108

  4. Hydrogen recombiner catalyst test supporting data

    SciTech Connect

    Britton, M.D.

    1995-01-19

    This is a data package supporting the Hydrogen Recombiner Catalyst Performance and Carbon Monoxide Sorption Capacity Test Report, WHC-SD-WM-TRP-211, Rev 0. This report contains 10 appendices which consist of the following: Mass spectrometer analysis reports: HRC samples 93-001 through 93-157; Gas spectrometry analysis reports: HRC samples 93-141 through 93-658; Mass spectrometer procedure PNL-MA-299 ALO-284; Alternate analytical method for ammonia and water vapor; Sample log sheets; Job Safety analysis; Certificate of mixture analysis for feed gases; Flow controller calibration check; Westinghouse Standards Laboratory report on Bois flow calibrator; and Sorption capacity test data, tables, and graphs.

  5. Population inversion in a stationary recombining plasma

    SciTech Connect

    Otsuka, M.

    1980-12-01

    Population inversion, which occurs in a recombining plasma when a stationary He plasma is brought into contact with a neutral gas, is examined. With hydrogen as a contact gas, noticeable inversion between low-lying levels of H as been found. The overpopulation density is of the order of 10/sup 8/ cm/sup -3/, which is much higher then that (approx. =10/sup 5/ cm/sup -3/) obtained previously with He as a contact gas. Relations between these experimental results and the conditions for population inversion are discussed with the CR model.

  6. Patterns of recombination on human chromosome 22

    SciTech Connect

    Schlumpf, K.S.; Kim, D.; Haines, J.L.

    1994-09-01

    Virtually all genetic linkage maps generated to date are gross averages across individuals, ages, and (often) sexes. In addition, although some level of positive interference has been assumed, until recently little evidence to support this in humans has been available. The major stumbling block has been the quality of the data available, since even a few genotypic errors can have drastic effects on both the map length and the number of apparent recombinants. In addition, variation in recombination by factors other than sex have pretty much been ignored. To explore recombination in more detail, we have generated a microsatellite marker map of human chromosome 22. This map includes 32 markers genotyped through 46 sibships of the Venezuelan Reference Pedigree (VRP). Extensive error checking and regenotyping was performed to remove as many genotypic errors as possible, but no genotypes were removed simply because they created unlikely events. The following 1000:1 odds map has been obtained: cen--F8VWFP1--11--S264--3-S311--4--S257--2--TOP1P2--3--S156--1--CRYB2--1--S258--2--S310--6--S193--1--S275--3--S268--1--S280--4--S304--3--S283--2--LiR1--3--IL2RB--3--S299--1--S302--1--S537--2--S270--4--PDGF--8--S274--qter. The female map (91 cM) is twice as long as the male map (46 cM) and the log-likelihood difference in the maps (22.3) is highly significant (P=0.001, df=22) and appears constant across the chromosome. Analysis of recombination with age showed no particular trends for either males or females when chromosomes were grouped into three categories (20, 20-30, 30+) by parental age at birth of child. Positive interference was found in maternally derived chromosomes ({chi}{sup 2}=30.5 (4), p<0.005), but not in paternally derived chromosomes ({chi}{sup 2}=6.24 (3), P=0.10). This contrasts to data from chromosomes 9 and 21 where positive interference was found for both sexes. More detailed analyses are in progress.

  7. Cutaneous allergy to human (recombinant DNA) insulin.

    PubMed

    Grammer, L C; Metzger, B E; Patterson, R

    1984-03-16

    p6 report two cases of cutaneous allergy to human (recombinant DNA) insulin. Each patient had a history of systemic allergic reactions to porcine insulin and was at least as reactive to human as to porcine insulin by end-point cutaneous titration. Both patients' insulin allergy was managed with animal insulins and both have done well. Our experience with these two patients indicates that human insulin (rDNA) should not be expected to be efficacious in all patients with systemic allergy to insulin. PMID:6366262

  8. Metal binding proteins, recombinant host cells and methods

    DOEpatents

    Summers, Anne O.; Caguiat, Jonathan J.

    2004-06-15

    The present disclosure provides artificial heavy metal binding proteins termed chelons by the inventors. These chelons bind cadmium and/or mercuric ions with relatively high affinity. Also disclosed are coding sequences, recombinant DNA molecules and recombinant host cells comprising those recombinant DNA molecules for expression of the chelon proteins. In the recombinant host cells or transgenic plants, the chelons can be used to bind heavy metals taken up from contaminated soil, groundwater or irrigation water and to concentrate and sequester those ions. Recombinant enteric bacteria can be used within the gastrointestinal tracts of animals or humans exposed to toxic metal ions such as mercury and/or cadmium, where the chelon recombinantly expressed in chosen in accordance with the ion to be rededicated. Alternatively, the chelons can be immobilized to solid supports to bind and concentrate heavy metals from a contaminated aqueous medium including biological fluids.

  9. Recombinant DNA products: Insulin, interferon and growth hormone

    SciTech Connect

    Bollon, A.P.

    1984-01-01

    This book provides the discussion of products of biotechnology of recombinant DNA. The contents include: Recombinant DNA techniques; isolation, cloning, and expression of genes; from somatostatin to human insulin; yeast; an alternative organism for foreign protein production; background in human interferon; preclinical assessment of biological properties of recombinant DNA derived human interferons; human clinical trials of bacteria-derived human ..cap alpha.. interferon.f large scale production of human alpha interferon from bacteria; direct expression of human growth hormone in escherichia coli with the lipoprotein promoter; biological actions in humans of recombinant DNA synthesized human growth hormone; NIH guidelines for research involving recombinant DNA molecules; appendix; viral vectors and the NHY guidelines; FDA's role in approval and regulation of recombinant DNA drugs; and index.

  10. The recombination mediator RAD51D promotes geminiviral infection.

    PubMed

    Richter, Kathrin S; Serra, Heϊdi; White, Charles I; Jeske, Holger

    2016-06-01

    To study a possible role for homologous recombination in geminivirus replication, we challenged Arabidopsis recombination gene knockouts by Euphorbia yellow mosaic virus infection. Our results show that the RAD51 paralog RAD51D, rather than RAD51 itself, promotes viral replication at early stages of infection. Blot hybridization analyses of replicative intermediates using one- and two-dimensional gels and deep sequencing point to an unexpected facet of recombination-dependent replication, the repair by single-strand annealing (SSA) during complementary strand replication. A significant decrease of both intramolecular, yielding defective DNAs and intermolecular recombinant molecules between the two geminiviral DNA components (A, B) were observed in the absence of RAD51D. By contrast, DNA A and B reacted differentially with the generation of inversions. A model to implicate single-strand annealing recombination in geminiviral recombination-dependent replication is proposed. PMID:27018825

  11. The Many Landscapes of Recombination in Drosophila melanogaster

    PubMed Central

    Comeron, Josep M.; Ratnappan, Ramesh; Bailin, Samuel

    2012-01-01

    Recombination is a fundamental biological process with profound evolutionary implications. Theory predicts that recombination increases the effectiveness of selection in natural populations. Yet, direct tests of this prediction have been restricted to qualitative trends due to the lack of detailed characterization of recombination rate variation across genomes and within species. The use of imprecise recombination rates can also skew population genetic analyses designed to assess the presence and mode of selection across genomes. Here we report the first integrated high-resolution description of genomic and population variation in recombination, which also distinguishes between the two outcomes of meiotic recombination: crossing over (CO) and gene conversion (GC). We characterized the products of 5,860 female meioses in Drosophila melanogaster by genotyping a total of 139 million informative SNPs and mapped 106,964 recombination events at a resolution down to 2 kilobases. This approach allowed us to generate whole-genome CO and GC maps as well as a detailed description of variation in recombination among individuals of this species. We describe many levels of variation in recombination rates. At a large-scale (100 kb), CO rates exhibit extreme and highly punctuated variation along chromosomes, with hot and coldspots. We also show extensive intra-specific variation in CO landscapes that is associated with hotspots at low frequency in our sample. GC rates are more uniformly distributed across the genome than CO rates and detectable in regions with reduced or absent CO. At a local scale, recombination events are associated with numerous sequence motifs and tend to occur within transcript regions, thus suggesting that chromatin accessibility favors double-strand breaks. All these non-independent layers of variation in recombination across genomes and among individuals need to be taken into account in order to obtain relevant estimates of recombination rates, and should

  12. Dissociative recombination of N2H+

    NASA Astrophysics Data System (ADS)

    dos Santos, S. Fonseca; Ngassam, V.; Orel, A. E.; Larson, Å.

    2016-08-01

    The direct and indirect mechanisms of dissociative recombination of N2H+ are theoretically studied. At low energies, the electron capture is found to be driven by recombination into bound Rydberg states, while at collision energies above 0.1 eV, the direct capture and dissociation along electronic resonant states becomes important. Electron-scattering calculations using the complex Kohn variational method are performed to obtain the scattering matrix as well as energy positions and autoionization widths of resonant states. Potential-energy surfaces of electronic bound states of N2H and N2H+ are computed using structure calculations with the multireference configuration interaction method. The cross section for the indirect mechanism is calculated using a vibrational frame transformation of the elements of the scattering matrix at energies just above the ionization threshold. Here vibrational excitations of the ionic core from v =0 to v =1 and v =2 for all three normal modes are considered and autoionization is neglected. The cross section for the direct dissociation along electronic resonant states is computed with wave-packet calculations using the multiconfiguration time-dependent Hartree method, where all three internal degrees of freedom are considered. The calculated cross sections are compared to measurements.

  13. Photoexcited charge pair escape and recombination

    SciTech Connect

    Braun, C.L.

    1991-11-15

    We report photocurrent transients arising from the pulsed laser excitation of the dipolar first excited singlet sate S{sub 1} of trans 4-dimethyl-amino-4{prime}-nitrostilbene (DMANS) in toluene solution. The currents arise from rotational reorientation of DMANS dipoles with respect to the axis of an applied electric field. The method appears to offer a simple and general approach to the measurement of the change in dipole moment upon electronic excitation of a molecule. In another experiment, durene (1,2,4,5-tetramethylbenzene) dissolved in n-hexane was photoionized by 35 psec pulses at 266 nm. Transient absorption at 1064 nm arising chiefly from geminate electrons was detected and used to monitor the recombination of the electron-cation pairs produced by two-photon ionization. An excellent fit to the recombination kinetics was obtained by assuming that the distribution of initial electron-cation separations was of the form r{sup 2}EXP = r{sup 2}/(2L{sup 3})exp({minus}r/L) with a mean radius 3L = 5.7 nm.

  14. The landscape of recombination in African Americans.

    PubMed

    Hinch, Anjali G; Tandon, Arti; Patterson, Nick; Song, Yunli; Rohland, Nadin; Palmer, Cameron D; Chen, Gary K; Wang, Kai; Buxbaum, Sarah G; Akylbekova, Ermeg L; Aldrich, Melinda C; Ambrosone, Christine B; Amos, Christopher; Bandera, Elisa V; Berndt, Sonja I; Bernstein, Leslie; Blot, William J; Bock, Cathryn H; Boerwinkle, Eric; Cai, Qiuyin; Caporaso, Neil; Casey, Graham; Cupples, L Adrienne; Deming, Sandra L; Diver, W Ryan; Divers, Jasmin; Fornage, Myriam; Gillanders, Elizabeth M; Glessner, Joseph; Harris, Curtis C; Hu, Jennifer J; Ingles, Sue A; Isaacs, William; John, Esther M; Kao, W H Linda; Keating, Brendan; Kittles, Rick A; Kolonel, Laurence N; Larkin, Emma; Le Marchand, Loic; McNeill, Lorna H; Millikan, Robert C; Murphy, Adam; Musani, Solomon; Neslund-Dudas, Christine; Nyante, Sarah; Papanicolaou, George J; Press, Michael F; Psaty, Bruce M; Reiner, Alex P; Rich, Stephen S; Rodriguez-Gil, Jorge L; Rotter, Jerome I; Rybicki, Benjamin A; Schwartz, Ann G; Signorello, Lisa B; Spitz, Margaret; Strom, Sara S; Thun, Michael J; Tucker, Margaret A; Wang, Zhaoming; Wiencke, John K; Witte, John S; Wrensch, Margaret; Wu, Xifeng; Yamamura, Yuko; Zanetti, Krista A; Zheng, Wei; Ziegler, Regina G; Zhu, Xiaofeng; Redline, Susan; Hirschhorn, Joel N; Henderson, Brian E; Taylor, Herman A; Price, Alkes L; Hakonarson, Hakon; Chanock, Stephen J; Haiman, Christopher A; Wilson, James G; Reich, David; Myers, Simon R

    2011-08-11

    Recombination, together with mutation, gives rise to genetic variation in populations. Here we leverage the recent mixture of people of African and European ancestry in the Americas to build a genetic map measuring the probability of crossing over at each position in the genome, based on about 2.1 million crossovers in 30,000 unrelated African Americans. At intervals of more than three megabases it is nearly identical to a map built in Europeans. At finer scales it differs significantly, and we identify about 2,500 recombination hotspots that are active in people of West African ancestry but nearly inactive in Europeans. The probability of a crossover at these hotspots is almost fully controlled by the alleles an individual carries at PRDM9 (P value < 10(-245)). We identify a 17-base-pair DNA sequence motif that is enriched in these hotspots, and is an excellent match to the predicted binding target of PRDM9 alleles common in West Africans and rare in Europeans. Sites of this motif are predicted to be risk loci for disease-causing genomic rearrangements in individuals carrying these alleles. More generally, this map provides a resource for research in human genetic variation and evolution. PMID:21775986

  15. Recombination-aware alignment of diploid individuals

    PubMed Central

    2014-01-01

    Background Traditionally biological similarity search has been studied under the abstraction of a single string to represent each genome. The more realistic representation of diploid genomes, with two strings defining the genome, has so far been largely omitted in this context. With the development of sequencing techniques and better phasing routines through haplotype assembly algorithms, we are not far from the situation when individual diploid genomes could be represented in their full complexity with a pair-wise alignment defining the genome. Results We propose a generalization of global alignment that is designed to measure similarity between phased predictions of individual diploid genomes. This generalization takes into account that individual diploid genomes evolve through a mutation and recombination process, and that predictions may be erroneous in both dimensions. Even though our model is generic, we focus on the case where one wants to measure only the similarity of genome content allowing free recombination. This results into efficient algorithms for direct application in (i) evaluation of variation calling predictions and (ii) progressive multiple alignments based on labeled directed acyclic graphs (DAGs) to represent profiles. The latter may be of more general interest, in connection to covering alignment of DAGs. Extensions of our model and algorithms can be foreseen to have applications in evaluating phasing algorithms, as well as more fundamental role in phasing child genome based on parent genomes. PMID:25572943

  16. Modified Fragmentation Function from Quark Recombination

    SciTech Connect

    Majumder, A.; Wang, Enke; Wang, Xin-Nian

    2005-07-26

    Within the framework of the constituent quark model, it isshown that the single hadron fragmentation function of a parton can beexpressed as a convolution of shower diquark or triquark distributionfunction and quark recombination probability, if the interference betweenamplitudes of quark recombination with different momenta is neglected.Therecombination probability is determined by the hadron's wavefunction inthe constituent quark model. The shower diquark or triquark distributionfunctions of a fragmenting jet are defined in terms of overlappingmatrices of constituent quarks and parton field operators. They aresimilar in form to dihadron or trihadron fragmentation functions in termsof parton operator and hadron states. Extending the formalism to thefield theory at finite temperature, we automatically derive contributionsto the effective single hadron fragmentation function from therecombination of shower and thermal constituent quarks. Suchcontributions involve single or diquark distribution functions which inturn can be related to diquark or triquark distribution functions via sumrules. We also derive QCD evolution equations for quark distributionfunctions that in turn determine the evolution of the effective jetfragmentation functions in a thermal medium.

  17. Lipopolysaccharide induced conversion of recombinant prion protein

    PubMed Central

    Saleem, Fozia; Bjorndahl, Trent C; Ladner, Carol L; Perez-Pineiro, Rolando; Ametaj, Burim N; Wishart, David S

    2014-01-01

    The conformational conversion of the cellular prion protein (PrPC) to the β-rich infectious isoform PrPSc is considered a critical and central feature in prion pathology. Although PrPSc is the critical component of the infectious agent, as proposed in the “protein-only” prion hypothesis, cellular components have been identified as important cofactors in triggering and enhancing the conversion of PrPC to proteinase K resistant PrPSc. A number of in vitro systems using various chemical and/or physical agents such as guanidine hydrochloride, urea, SDS, high temperature, and low pH, have been developed that cause PrPC conversion, their amplification, and amyloid fibril formation often under non-physiological conditions. In our ongoing efforts to look for endogenous and exogenous chemical mediators that might initiate, influence, or result in the natural conversion of PrPC to PrPSc, we discovered that lipopolysaccharide (LPS), a component of gram-negative bacterial membranes interacts with recombinant prion proteins and induces conversion to an isoform richer in β sheet at near physiological conditions as long as the LPS concentration remains above the critical micelle concentration (CMC). More significant was the LPS mediated conversion that was observed even at sub-molar ratios of LPS to recombinant ShPrP (90–232). PMID:24819168

  18. Ethanol precipitation for purification of recombinant antibodies.

    PubMed

    Tscheliessnig, Anne; Satzer, Peter; Hammerschmidt, Nikolaus; Schulz, Henk; Helk, Bernhard; Jungbauer, Alois

    2014-10-20

    Currently, the golden standard for the purification of recombinant humanized antibodies (rhAbs) from CHO cell culture is protein A chromatography. However, due to increasing rhAbs titers alternative methods have come into focus. A new strategy for purification of recombinant human antibodies from CHO cell culture supernatant based on cold ethanol precipitation (CEP) and CaCl2 precipitation has been developed. This method is based on the cold ethanol precipitation, the process used for purification of antibodies and other components from blood plasma. We proof the applicability of the developed process for four different antibodies resulting in similar yield and purity as a protein A chromatography based process. This process can be further improved using an anion-exchange chromatography in flowthrough mode e.g. a monolith as last step so that residual host cell protein is reduced to a minimum. Beside the ethanol based process, our data also suggest that ethanol could be replaced with methanol or isopropanol. The process is suited for continuous operation. PMID:25087738

  19. Recombinant synthesis of hyaluronan by Agrobacterium sp.

    PubMed

    Mao, Zichao; Chen, Rachel Ruizhen

    2007-01-01

    Hyaluronan (HA) is a sugar polymer of a repeating disaccharide, beta1-3 D-N-acetylglucosamine (GlcNAc) beta1-4 D-glucuronic acid (GlcA). It finds applications in numerous biomedical procedures such as ophthalmic surgery and osteoarthritis treatment. Until recently, the only commercial sources were extraction of rooster combs and from fermentation of pathogenic Streptococcus. In this work, we demonstrate that metabolic engineering strategies enable the recombinant synthesis of hyaluronan in a safe microorganism. Agrobacterium sp. ATCC 31749 is a commercial production strain for a food polymer, Curdlan. A broad host range expression vector was successfully developed to express the 3 kb HA synthase gene from Pasteurella multocida, along with a kfiD gene encoding UDP-glucose dehydrogenase from Escherichia coli K5 strain. Coexpression of these two heterologous enzymes enables Agrobacterium to produce HA. Hyaluronan was accumulated up to 0.3 g/L in shaker flask cultivation. The molecular weight of the polymer from various Agrobacterium strains is in the range of 0.7-2 MD. To our knowledge, this is the first successful recombinant hyaluronan synthesis in a Gram-negative bacterium that naturally produces a food product. The ease of genetic modifications provides future opportunities to tailor properties of polymers for specific applications. PMID:17705506

  20. Recombinant viral vaccines for enzootic bovine leucosis.

    PubMed

    Daniel, R C; Gatei, M H; Good, M F; Boyle, D B; Lavin, M F

    1993-10-01

    Recently published studies on the development and use of recombinant vaccinia virus (VV) vaccines incorporating either the complete envelope (env) gene or only a fragment of the env gene consisting of the coding sequence for the env glycoprotein 51 (gp51) and part of gp30 of the bovine leukaemia virus (BLV) are described. It has been reported that vaccination of sheep with recombinant VV vaccines containing the complete env gene appears to protect sheep against challenge infection with BLV. The evidence for this protection is based on the lack of persistence of high titres of anti-gp51 antibodies compared with unvaccinated BLV infected controls, on the enhanced CD4 proliferative responses to specific BLV gp51 synthetic peptides in the vaccinated sheep, and on the inability to detect BLV pro-virus by polymerase chain reaction in the vaccinated sheep after 4 months following challenge infection compared with continual detection in unvaccinated sheep over a 16 month trial period. It has been suggested that cell-mediated immune responses may be an important aspect of protective immunity against BLV infection and it has been reported that large tracts of amino acid sequences within the env and pol genes are highly conserved in different isolates from different countries which is of importance in designing peptide derived vaccines. PMID:8270269

  1. Recombinant protein scaffolds for tissue engineering.

    PubMed

    Werkmeister, Jerome A; Ramshaw, John A M

    2012-02-01

    New biological materials for tissue engineering are now being developed using common genetic engineering capabilities to clone and express a variety of genetic elements that allow cost-effective purification and scaffold fabrication from these recombinant proteins, peptides or from chimeric combinations of these. The field is limitless as long as the gene sequences are known. The utility is dependent on the ease, product yield and adaptability of these protein products to the biomedical field. The development of recombinant proteins as scaffolds, while still an emerging technology with respect to commercial products, is scientifically superior to current use of natural materials or synthetic polymer scaffolds, in terms of designing specific structures with desired degrees of biological complexities and motifs. In the field of tissue engineering, next generation scaffolds will be the key to directing appropriate tissue regeneration. The initial period of biodegradable synthetic scaffolds that provided shape and mechanical integrity, but no biological information, is phasing out. The era of protein scaffolds offers distinct advantages, particularly with the combination of powerful tools of molecular biology. These include, for example, the production of human proteins of uniform quality that are free of infectious agents and the ability to make suitable quantities of proteins that are found in low quantity or are hard to isolate from tissue. For the particular needs of tissue engineering scaffolds, fibrous proteins like collagens, elastin, silks and combinations of these offer further advantages of natural well-defined structural scaffolds as well as endless possibilities of controlling functionality by genetic manipulation. PMID:22262725

  2. Geminate recombination of O2 and hemoglobin.

    PubMed Central

    Chernoff, D A; Hochstrasser, R M; Steele, A W

    1980-01-01

    The photolysis of HbO2 and HbCO has been studied by measuring transient absorption spectra in the Soret region after excitation with picosecond pulses at 530 nm. Dissociation occurred promptly in both cases, followed (for HbO2) by geminate recombination of ca. 40% of the photodissociated O2 with a lifetime of 200 +/- 70 psec (25 degrees C). No recombination of Hb + CO was observed up to 1200 psec after photolysis. The HbO2 and HbCO photoproduct spectra were broader, weaker, and red-shifted in comparison to the spectrum of stable Hb and Gibson's fast-reacting form, Hb. For HbO2 the spectrum was initially much broader to longer wavelengths but relaxed to a constant shape within 90 psec, whereas for HbCO there was no spectral evolution. The photophysics is analyzed by considering the effect of spin constraints as well as spin--orbit coupling and orbital correlation among the various electronic states of liganded and deoxy hemoglobins. The small quantum yield of HbO2 dissociation is not primarily due to rebinding but rather to electronic relaxation to nonreactive states. PMID:6932659

  3. [Telomere Recombination in Normal Mammalian Cells].

    PubMed

    Zhdanova, N S; Rubtsov, N B

    2016-01-01

    Two mechanisms of telomere length maintenance are known to date. The first includes the use of a special enzymatic telomerase complex to solve the problems that arise during the replication of linear DNA in a normal diploid and part of tumor cells. Alternative lengthening of telomeres (ALT), which is based on the homologous recombination of telomere DNA, represents the second mechanism. Until recently, ALT was assumed to be expressed only in 15-20% of tumors lacking active telomerase and, together with telomerase reactivation represented one of two possibilities to overcome the replicative senescence observed in somatic mammalian cells due to aging or during cell culturing in vitro. Previously described sporadic cases of combinations of the two mechanisms of telomere length maintenance in several cell lines in vitro were attributed to the experimental design rather than to a real biological phenomenon, since active cellular division without active telomerase was considered to be the "gold standard" of ALT. The present review describes the morphological and functional reorganizations of mammalian telomeres observed with ALT activation, as well as recently observed,and well-documented cases of combinations between ALT-like and telomerase-dependent mechanisms in mammalian cells. The possible role of telomere recombination in telomerase-dependent cells is discussed. PMID:27183789

  4. The landscape of recombination in African Americans

    PubMed Central

    Hinch, Anjali G.; Tandon, Arti; Patterson, Nick; Song, Yunli; Rohland, Nadin; Palmer, Cameron D.; Chen, Gary K.; Wang, Kai; Buxbaum, Sarah G.; Akylbekova, Meggie; Aldrich, Melinda C.; Ambrosone, Christine B.; Amos, Christopher; Bandera, Elisa V.; Berndt, Sonja I.; Bernstein, Leslie; Blot, William J.; Bock, Cathryn H.; Boerwinkle, Eric; Cai, Qiuyin; Caporaso, Neil; Casey, Graham; Cupples, L. Adrienne; Deming, Sandra L.; Diver, W. Ryan; Divers, Jasmin; Fornage, Myriam; Gillanders, Elizabeth M.; Glessner, Joseph; Harris, Curtis C.; Hu, Jennifer J.; Ingles, Sue A.; Isaacs, Williams; John, Esther M.; Kao, W. H. Linda; Keating, Brendan; Kittles, Rick A.; Kolonel, Laurence N.; Larkin, Emma; Le Marchand, Loic; McNeill, Lorna H.; Millikan, Robert C.; Murphy, Adam; Musani, Solomon; Neslund-Dudas, Christine; Nyante, Sarah; Papanicolaou, George J.; Press, Michael F.; Psaty, Bruce M.; Reiner, Alex P.; Rich, Stephen S.; Rodriguez-Gil, Jorge L.; Rotter, Jerome I.; Rybicki, Benjamin A.; Schwartz, Ann G.; Signorello, Lisa B.; Spitz, Margaret; Strom, Sara S.; Thun, Michael J.; Tucker, Margaret A.; Wang, Zhaoming; Wiencke, John K.; Witte, John S.; Wrensch, Margaret; Wu, Xifeng; Yamamura, Yuko; Zanetti, Krista A.; Zheng, Wei; Ziegler, Regina G.; Zhu, Xiaofeng; Redline, Susan; Hirschhorn, Joel N.; Henderson, Brian E.; Taylor, Herman A.; Price, Alkes L.; Hakonarson, Hakon; Chanock, Stephen J.; Haiman, Christopher A.; Wilson, James G.; Reich, David; Myers, Simon R.

    2011-01-01

    Recombination, together with mutation, is the ultimate source of genetic variation in populations. We leverage the recent mixture of people of African and European ancestry in the Americas to build a genetic map measuring the probability of crossing-over at each position in the genome, based on about 2.1 million crossovers in 30,000 unrelated African Americans. At intervals of more than three megabases it is nearly identical to a map built in Europeans. At finer scales it differs significantly, and we identify about 2,500 recombination hotspots that are active in people of West African ancestry but nearly inactive in Europeans. The probability of a crossover at these hotspots is almost fully controlled by the alleles an individual carries at PRDM9 (P<10−245). We identify a 17 base pair DNA sequence motif that is enriched in these hotspots, and is an excellent match to the predicted binding target of African-enriched alleles of PRDM9. PMID:21775986

  5. Monitoring recombinant human erythropoietin abuse among athletes.

    PubMed

    Citartan, Marimuthu; Gopinath, Subash C B; Chen, Yeng; Lakshmipriya, Thangavel; Tang, Thean-Hock

    2015-01-15

    The illegal administration of recombinant human erythropoietin (rHuEPO) among athletes is largely preferred over blood doping to enhance stamina. The advent of recombinant DNA technology allowed the expression of EPO-encoding genes in several eukaryotic hosts to produce rHuEPO, and today these performance-enhancing drugs are readily available. As a mimetic of endogenous EPO (eEPO), rHuEPO augments the oxygen carrying capacity of blood. Thus, monitoring the illicit use of rHuEPO among athletes is crucial in ensuring an even playing field and maintaining the welfare of athletes. A number of rHuEPO detection methods currently exist, including measurement of hematologic parameters, gene-based detection methods, glycomics, use of peptide markers, electrophoresis, isoelectric focusing (IEF)-double immunoblotting, aptamer/antibody-based methods, and lateral flow tests. This review gleans these different strategies and highlights the leading molecular recognition elements that have potential roles in rHuEPO doping detection. PMID:25058943

  6. Kinetics of radiative recombination in quantum wells

    NASA Astrophysics Data System (ADS)

    Ridley, B. K.

    1990-06-01

    A theory of radiative-recombination kinetics which treats free carriers, excitons, and photon recycling in a quantum-well system is presented. An expression for the temporal decay of excess carriers which encompasses large- and small-signal regimes is derived. When excitons are present the decay can be approximated by two exponentials in general, and in the large-signal regime the photoluminescence time constant is half as long as that associated with photoconductivity. Explicit expressions for the recombination coefficients are given and their magnitudes discussed for nondegenerate and degenerate populations in GaAs. Excitons are shown to enhance the temperature dependence. A simple model of exciton screening is used to illustrate the dependence of radiative time constants on background carrier density, which deviates significantly from the conventional free-carrier dependence. The magnitudes of radiative time constants in real systems depend, in addition to material characteristics, upon the details of exciton screening, the overlap of the electron and hole wave functions in the quantum well, and the probability of photon reabsorption, all of which are specimen specific. It is pointed out that the transition from a degenerate to a nondegenerate population may be misinterpreted in terms of Auger processes.

  7. Expression and purification of recombinant nattokinase in Spodoptera frugiperda cells.

    PubMed

    Li, Xiaoxiang; Wang, Xiaoli; Xiong, Shaoling; Zhang, Jing; Cai, Litao; Yang, Yanyan

    2007-10-01

    A recombinant baculovirus, rv-egfp-NK, containing a reporter gene encoding the enhanced green fluorescent protein (EGFP), was used to express nattokinase (NK), a fibrinolytic enzyme, in Spodoptera frugiperda (SF-9) cells. The recombinant protein also included a histidine tag for purification using Ni(2+) resins. The recombinant NK, approximately 30 kDa, retained fibrinolytic activity (60 U/ml). The integration of the EGFP expression cassette in the Bac-to-Bac system is thus an effective method for the expression and purification of recombinant NK protein in Spodoptera frugiperda insect cells. PMID:17581705

  8. The evolution of recombination in a heterogeneous environment.

    PubMed Central

    Lenormand, T; Otto, S P

    2000-01-01

    Most models describing the evolution of recombination have focused on the case of a single population, implicitly assuming that all individuals are equally likely to mate and that spatial heterogeneity in selection is absent. In these models, the evolution of recombination is driven by linkage disequilibria generated either by epistatic selection or drift. Models based on epistatic selection show that recombination can be favored if epistasis is negative and weak compared to directional selection and if the recombination modifier locus is tightly linked to the selected loci. In this article, we examine the joint effects of spatial heterogeneity in selection and epistasis on the evolution of recombination. In a model with two patches, each subject to different selection regimes, we consider the cases of mutation-selection and migration-selection balance as well as the spread of beneficial alleles. We find that including spatial heterogeneity extends the range of epistasis over which recombination can be favored. Indeed, recombination can be favored without epistasis, with negative and even with positive epistasis depending on environmental circumstances. The selection pressure acting on recombination-modifier loci is often much stronger with spatial heterogeneity, and even loosely linked modifiers and free linkage may evolve. In each case, predicting whether recombination is favored requires knowledge of both the type of environmental heterogeneity and epistasis, as none of these factors alone is sufficient to predict the outcome. PMID:10978305

  9. Charge recombination in disordered neat polymer films under imbalanced excitation conditions studied using the recombination time of flight technique

    NASA Astrophysics Data System (ADS)

    Ravia, Israel; Tessler, Nir

    2012-05-01

    It has recently been suggested that the charge recombination rate in amorphous polymers could be affected by the energy distribution of electrons and holes as well as that of the resulting excitons. To test this hypothesis, we developed a new method for measuring charge recombination under highly imbalanced conditions. We find that if the electron density is higher than that of the holes, increasing the electron density further results in reduction of the recombination coefficient. We attribute this to the very different energy distribution between low and high carrier densities, which is not accounted for in the Langevin recombination model.

  10. Photoionization and electron-ion recombination of Ti I

    NASA Astrophysics Data System (ADS)

    Nahar, Sultana N.

    2016-07-01

    Study of the inverse processes of photoionization and electron-ion recombination of (Ti I + h ν ⇋ Ti II + e) using the unified method is reported. The method, based on close coupling (CC) approximation and R-matrix method, subsumes both the radiative recombination (RR) and dielectronic recombination (DR) in a unified manner and provides state-specific and total electron-ion recombination rate coefficients which are self-consistent with the state-specific photoionization cross sections. The present results include state-specific electron-ion recombination rates (αRC(i))and partial photoionization cross sections (σPI(i)) leaving the ion in the ground state of 813 bound states with n ≤ 10 and l ≤ 9 of Ti I. Various features of state-specific and total electron-ion recombination with temperature, and the corresponding photoionization cross sections with energies are discussed with illustrations. Due to closely lying excited states near the ground state of the core, photoionization cross sections show presence of narrow Rydberg resonances in low energy region near the ionization threshold. Many excited states also show broad and enhanced Seaton resonances due to PEC (photo-excitation-of-core) which contribute to the high temperature recombination. The total recombination rate coefficient is found to show a low hump around temperature 280 K and a high dielectronic recombination peak at temperature 25,000 K. Total spectrum of recombination cross sections and rates with photoelectron energy are also presented for experimental observation. Calculations were carried out using a CC wave function expansion of 36 states of the core ion Ti II. The large set of data for recombination rates and partial photoionization cross sections with resonances should provide a complete and accurate modelings of plasmas.

  11. Recombination of Globally Circulating Varicella-Zoster Virus

    PubMed Central

    Depledge, Daniel P.; Kundu, Samit; Atkinson, Claire; Brown, Julianne; Haque, Tanzina; Hussaini, Yusuf; MacMahon, Eithne; Molyneaux, Pamela; Papaevangelou, Vassiliki; Sengupta, Nitu; Koay, Evelyn S. C.; Tang, Julian W.; Underhill, Gillian S.; Grahn, Anna; Studahl, Marie; Breuer, Judith; Bergström, Tomas

    2015-01-01

    ABSTRACT Varicella-zoster virus (VZV) is a human herpesvirus, which during primary infection typically causes varicella (chicken pox) and establishes lifelong latency in sensory and autonomic ganglia. Later in life, the virus may reactivate to cause herpes zoster (HZ; also known as shingles). To prevent these diseases, a live-attenuated heterogeneous vaccine preparation, vOka, is used routinely in many countries worldwide. Recent studies of another alphaherpesvirus, infectious laryngotracheitis virus, demonstrate that live-attenuated vaccine strains can recombine in vivo, creating virulent progeny. These findings raised concerns about using attenuated herpesvirus vaccines under conditions that favor recombination. To investigate whether VZV may undergo recombination, which is a prerequisite for VZV vaccination to create such conditions, we here analyzed 115 complete VZV genomes. Our results demonstrate that recombination occurs frequently for VZV. It thus seems that VZV is fully capable of recombination if given the opportunity, which may have important implications for continued VZV vaccination. Although no interclade vaccine-wild-type recombinant strains were found, intraclade recombinants were frequently detected in clade 2, which harbors the vaccine strains, suggesting that the vaccine strains have already been involved in recombination events, either in vivo or in vitro during passages in cell culture. Finally, previous partial and complete genomic studies have described strains that do not cluster phylogenetically to any of the five established clades. The additional VZV strains sequenced here, in combination with those previously published, have enabled us to formally define a novel sixth VZV clade. IMPORTANCE Although genetic recombination has been demonstrated to frequently occur for other human alphaherpesviruses, herpes simplex viruses 1 and 2, only a few ancient and isolated recent recombination events have hitherto been demonstrated for VZV. In the

  12. Visible charge exchange recombination spectroscopy on TFTR

    SciTech Connect

    Stratton, B.C.; Fonck, R.J.; Jaehnig, K.P.; Schechtman, N.; Synakowski, E.J.

    1991-03-01

    Visible charge exchange recombination spectroscopy is routinely used to measure the time evolution of the ion temperature (T{sub i}) and toroidal rotation velocity (v{sub {phi}}) profiles on TFTR. These measurements are made with the CHERS diagnostic, a fiber-optically coupled spectrometer equipped with a two-dimensional photodiode array detector which provides both spectral and spatial resolution. The instrumentation, data analysis techniques, and examples of T{sub i} and v{sub {phi}} measurements are described. Recently, CHERS has been used to perform impurity transport experiments: radial profiles of diffusivities and convective velocities for helium and iron have been deduced from measurements of the time evolutions of He{sup 2+} and Fe{sup 24+} profiles following impurity injection. Examples of these measurements are given. 12 refs., 8 figs.

  13. Recombinant hepatitis B triple antigen vaccine: Hepacare.

    PubMed

    Zuckerman, Jane N; Zuckerman, Arie J

    2002-08-01

    Infection with hepatitis B virus is a public health problem throughout the world. Hepatitis B vaccines are now included in national immunization programmes of infants and/or adolescents in 129 countries. Current single antigen vaccines, that are plasma-derived or produced by recombinant DNA technology are highly effective, but between 5-10% or more of healthy immunocompetent subjects do not mount an antihepatitis B surface antibody protective response and others respond poorly (hyporesponders). The inclusion of pre-S1 and -S2 components of hepatitis B surface antigen in addition to the single antigen (triple antigen) in a novel vaccine, Hepacare, Medeva Pharma Plc, Speke, UK, overcomes nonresponsiveness and hyporesponsiveness in a significant number of individuals. The triple antigen is indicated for vaccination of nonresponders (and hyporesponders) to the current single antigen vaccines and for persons who require rapid protection against hepatitis B infection. PMID:12901552

  14. Recombining overlapping BACs into single large BACs.

    PubMed

    Kotzamanis, George; Kotsinas, Athanassios

    2015-01-01

    BAC clones containing the entire genomic region of a gene including the long-range regulatory elements are very useful for gene functional analysis. However, large genes often span more than the insert of a BAC clone, and single BACs covering the entire region of interest are not available. Here, we describe a general system for linking two or more overlapping BACs into a single clone. Two rounds of homologous recombination are used. In the first, the BAC inserts are subcloned into the pBACLink vectors. In the second, the two BACs are combined together. Multiple BACs in a contig can be combined by alternating use of the pBACLInk vectors, resulting in several BAC clones containing as much of the genomic region of a gene as required. Such BACs can then be used in gene expression studies and/or gene therapy applications. PMID:25239744

  15. Epidemiology, Genetic Recombination, and Pathogenesis of Coronaviruses.

    PubMed

    Su, Shuo; Wong, Gary; Shi, Weifeng; Liu, Jun; Lai, Alexander C K; Zhou, Jiyong; Liu, Wenjun; Bi, Yuhai; Gao, George F

    2016-06-01

    Human coronaviruses (HCoVs) were first described in the 1960s for patients with the common cold. Since then, more HCoVs have been discovered, including those that cause severe acute respiratory syndrome (SARS) and Middle East respiratory syndrome (MERS), two pathogens that, upon infection, can cause fatal respiratory disease in humans. It was recently discovered that dromedary camels in Saudi Arabia harbor three different HCoV species, including a dominant MERS HCoV lineage that was responsible for the outbreaks in the Middle East and South Korea during 2015. In this review we aim to compare and contrast the different HCoVs with regard to epidemiology and pathogenesis, in addition to the virus evolution and recombination events which have, on occasion, resulted in outbreaks amongst humans. PMID:27012512

  16. Integrative mobile elements exploiting Xer recombination.

    PubMed

    Das, Bhabatosh; Martínez, Eriel; Midonet, Caroline; Barre, François-Xavier

    2013-01-01

    Integrative mobile genetic elements directly participate in the rapid response of bacteria to environmental challenges. They generally encode their own dedicated recombination machineries. CTXφ, a filamentous bacteriophage that harbors the genes encoding cholera toxin in Vibrio cholerae provided the first notable exception to this rule: it hijacks XerC and XerD, two chromosome-encoded tyrosine recombinases for lysogenic conversion. XerC and XerD are highly conserved in bacteria because of their role in the topological maintenance of circular chromosomes and, with the advent of high throughput sequencing, numerous other integrative mobile elements exploiting them have been discovered. Here, we review our understanding of the molecular mechanisms of integration of the different integrative mobile elements exploiting Xer (IMEXs) so far described. PMID:23127381

  17. Charge exchange recombination spectroscopy on fusion devices

    SciTech Connect

    Duval, B. P.

    2012-05-25

    For fusion, obtaining reliable measurements of basic plasma parameters like ion and electron densities and temperatures is a primary goal. For theory, measurements are needed as a function of time and space to understand plasma transport and confinement with the ultimate goal of achieving economic nuclear fusion power. Electron profile measurements and plasma spectroscopy for the plasma ions are introduced. With the advent of Neutral Beam auxiliary plasma heating, Charge Exchange Recombination Spectroscopy provides accurate and time resolved measurements of the ions in large volume fusion devices. In acknowledgement of Nicol Peacock's role in the development of these techniques, still at the forefront of plasma fusion research, this paper describes the evolution of this diagnostic method.

  18. Dissociative Recombination Dynamics of the Ozone Cation

    SciTech Connect

    Zhaunerchyk, Vitali; Geppert, W.; sterdahl, F.; Larsson, Mats; Thomas, R. D.; Bahati Musafiri, Eric; Bannister, Mark E; Fogle, Mark R.; Vane, C Randy

    2008-02-01

    The dissociative recombination of the ozone cation has been studied at the heavy-ion storage ring CRYRING. The total cross section and branching fractions have been measured. The cross section from {approx}0eV to 0.2 eV follows a nearly E{sup -1} dependence, which was theoretically predicted to be a characteristic of the direct dissociative recombination mechanism. The thermal rate coefficient has been deduced from the cross section to be 7.37x10{sup -7}(T/300){sup -0.55}cm{sup 3}s{sup -1}. The branching fraction analysis carried out at {approx}0eV interaction energy has shown a strong propensity (94%) to dissociate through the three-body channel. Due to the overwhelming dominance of this channel it has been investigated in more detail. Of the six energetically available three-body pathways only three are significantly populated, such that the production of O(S1) is highly unfavored and all atomic oxygen fragments are predominantly formed in P3 and D1 states. Analysis of the breakup geometries has been performed by means of the Dalitz plot. It is observed that the molecules dissociating through the O(P3)+O(P3)+O(P3) and O(P3)+O(P3)+O(D1) channels have an open linear geometry where the cleavage of two valence bonds occurs preferentially in unison, while the O(P3)+O(D1)+O(D1) breakup might proceed partly through a sequential mechanism.

  19. Functional, Responsive Materials Assembled from Recombinant Oleosin

    NASA Astrophysics Data System (ADS)

    Hammer, Daniel

    Biological cells are surrounded by a plasma membrane made primarily of phospholipids that form a bilayer. This membrane is permselective and compartmentalizes the cell. A simple form of artificial cell is the vesicle, in which a phospholipid bilayer membrane surrounds an aqueous solution. However, there is no a priori reason why a membrane needs to be made of phospholipids. It could be made of any surfactant that forms a bilayer. We have assembled membranes and other structures from the recombinant plant protein oleosin. The ability to assemble from a recombinant protein means that every molecule is identical, we have complete control over the sequence, and hence can build in designer functionality with high fidelity, including adhesion and enzymatic activity. Such incorporation is trivial using the tools of molecular biology. We find that while many variants of oleosin make membranes, others make micelles and sheets. We show how the type of supramolecular structure can be altered by the conditions of solvent, such as ionic strength, and the architecture of the surfactant itself. We show that protease cleavable domains can be incorporated within oleosin, and be engineered to protect other functional domains such as adhesive motifs, to make responsive materials whose activity and shape depend on the action of proteases. We will also present the idea of making ``Franken''-oleosins, where large domains of native oleosin are replaced with domains from other functional proteins, to make hybrids conferred by the donor protein. Thus, we can view oleosin as a template upon which a vast array of designer functionalities can be imparted..

  20. The Landscape of Realized Homologous Recombination in Pathogenic Bacteria

    PubMed Central

    Yahara, Koji; Didelot, Xavier; Jolley, Keith A.; Kobayashi, Ichizo; Maiden, Martin C.J.; Sheppard, Samuel K.; Falush, Daniel

    2016-01-01

    Recombination enhances the adaptive potential of organisms by allowing genetic variants to be tested on multiple genomic backgrounds. Its distribution in the genome can provide insight into the evolutionary forces that underlie traits, such as the emergence of pathogenicity. Here, we examined landscapes of realized homologous recombination of 500 genomes from ten bacterial species and found all species have “hot” regions with elevated rates relative to the genome average. We examined the size, gene content, and chromosomal features associated with these regions and the correlations between closely related species. The recombination landscape is variable and evolves rapidly. For example in Salmonella, only short regions of around 1 kb in length are hot whereas in the closely related species Escherichia coli, some hot regions exceed 100 kb, spanning many genes. Only Streptococcus pyogenes shows evidence for the positive correlation between GC content and recombination that has been reported for several eukaryotes. Genes with function related to the cell surface/membrane are often found in recombination hot regions but E. coli is the only species where genes annotated as “virulence associated” are consistently hotter. There is also evidence that some genes with “housekeeping” functions tend to be overrepresented in cold regions. For example, ribosomal proteins showed low recombination in all of the species. Among specific genes, transferrin-binding proteins are recombination hot in all three of the species in which they were found, and are subject to interspecies recombination. PMID:26516092

  1. Radiative recombination and excited-state photoionization of lithium

    SciTech Connect

    Lahiri, J. ); Manson, S.T. )

    1993-11-01

    The radiative-recombination rate coefficients for electrons impinging on Li[sup +], along with the associated excited-state photoionization cross sections for Li, are calculated in the low-energy region. In addition to the totals, the contribution of the recombination of individual excited states to the total is discussed.

  2. Inference of Ancestral Recombination Graphs through Topological Data Analysis

    PubMed Central

    Cámara, Pablo G.; Levine, Arnold J.; Rabadán, Raúl

    2016-01-01

    The recent explosion of genomic data has underscored the need for interpretable and comprehensive analyses that can capture complex phylogenetic relationships within and across species. Recombination, reassortment and horizontal gene transfer constitute examples of pervasive biological phenomena that cannot be captured by tree-like representations. Starting from hundreds of genomes, we are interested in the reconstruction of potential evolutionary histories leading to the observed data. Ancestral recombination graphs represent potential histories that explicitly accommodate recombination and mutation events across orthologous genomes. However, they are computationally costly to reconstruct, usually being infeasible for more than few tens of genomes. Recently, Topological Data Analysis (TDA) methods have been proposed as robust and scalable methods that can capture the genetic scale and frequency of recombination. We build upon previous TDA developments for detecting and quantifying recombination, and present a novel framework that can be applied to hundreds of genomes and can be interpreted in terms of minimal histories of mutation and recombination events, quantifying the scales and identifying the genomic locations of recombinations. We implement this framework in a software package, called TARGet, and apply it to several examples, including small migration between different populations, human recombination, and horizontal evolution in finches inhabiting the Galápagos Islands. PMID:27532298

  3. Not so fast on recombination analysis of Newcastle disease virus

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Regarding the letter published in the Journal of Virology Vol. 82 No. 13 p. 6782 indicating that “powerful evidence” of recombination is a call for caution in the use of Newcastle Disease Virus (NDV) based vaccines, I would like to suggest that evidence for recombination is still weak. The authors ...

  4. Termolecular ionic recombination at high ambient gas density

    NASA Technical Reports Server (NTRS)

    Bates, D. R.

    1983-01-01

    It is shown that the importance of diffusion relative to drift in ion-ion recombination is a decreasing function of the separation between the ions (contrary to a common supposition). Langevin's formula for the recombination coefficient is obtained very simply using a physical model which allows for both diffusion and drift.

  5. Experimental studies of the dissociative recombination of H3 +

    NASA Astrophysics Data System (ADS)

    Larsson, M.

    2000-09-01

    Despite many experimental efforts over several decades, no consensus has bee n reached concerning the rate of dissociative recombination of H3+. We review the current status concerning different experimental appro aches to the measurement of the cross-section, rate coefficient and branching r atios in electron recombination of H3+.

  6. Recombination in viruses: mechanisms, methods of study, and evolutionary consequences.

    PubMed

    Pérez-Losada, Marcos; Arenas, Miguel; Galán, Juan Carlos; Palero, Ferran; González-Candelas, Fernando

    2015-03-01

    Recombination is a pervasive process generating diversity in most viruses. It joins variants that arise independently within the same molecule, creating new opportunities for viruses to overcome selective pressures and to adapt to new environments and hosts. Consequently, the analysis of viral recombination attracts the interest of clinicians, epidemiologists, molecular biologists and evolutionary biologists. In this review we present an overview of three major areas related to viral recombination: (i) the molecular mechanisms that underlie recombination in model viruses, including DNA-viruses (Herpesvirus) and RNA-viruses (Human Influenza Virus and Human Immunodeficiency Virus), (ii) the analytical procedures to detect recombination in viral sequences and to determine the recombination breakpoints, along with the conceptual and methodological tools currently used and a brief overview of the impact of new sequencing technologies on the detection of recombination, and (iii) the major areas in the evolutionary analysis of viral populations on which recombination has an impact. These include the evaluation of selective pressures acting on viral populations, the application of evolutionary reconstructions in the characterization of centralized genes for vaccine design, and the evaluation of linkage disequilibrium and population structure. PMID:25541518

  7. Recombinant HT.sub.m4 gene, protein and assays

    DOEpatents

    Lim, Bing; Adra, Chaker N.; Lelias, Jean-Michel

    1996-01-01

    The invention relates to a recombinant DNA molecule which encodes a HT.sub.m4 protein, a transformed host cell which has been stably transfected with a DNA molecule which encodes a HT.sub.m4 protein and a recombinant HT.sub.m4 protein. The invention also relates to a method for detecting the presence of a hereditary atopy.

  8. Recombination in Avian Gamma-Coronavirus Infectious Bronchitis Virus

    PubMed Central

    Thor, Sharmi W.; Hilt, Deborah A.; Kissinger, Jessica C.; Paterson, Andrew H.; Jackwood, Mark W.

    2011-01-01

    Recombination in the family Coronaviridae has been well documented and is thought to be a contributing factor in the emergence and evolution of different coronaviral genotypes as well as different species of coronavirus. However, there are limited data available on the frequency and extent of recombination in coronaviruses in nature and particularly for the avian gamma-coronaviruses where only recently the emergence of a turkey coronavirus has been attributed solely to recombination. In this study, the full-length genomes of eight avian gamma-coronavirus infectious bronchitis virus (IBV) isolates were sequenced and along with other full-length IBV genomes available from GenBank were analyzed for recombination. Evidence of recombination was found in every sequence analyzed and was distributed throughout the entire genome. Areas that have the highest occurrence of recombination are located in regions of the genome that code for nonstructural proteins 2, 3 and 16, and the structural spike glycoprotein. The extent of the recombination observed, suggests that this may be one of the principal mechanisms for generating genetic and antigenic diversity within IBV. These data indicate that reticulate evolutionary change due to recombination in IBV, likely plays a major role in the origin and adaptation of the virus leading to new genetic types and strains of the virus. PMID:21994806

  9. Oxygen recombination in the sealed nickel-cadmium cell

    NASA Technical Reports Server (NTRS)

    Zimmerman, Al H.; Barrera, Tom P.

    1991-01-01

    The purpose of this study was to determine what parameters are most critical for controlling overcharge pressure in a sealed nickel cadmium cell. Topics covered in viewgraph format are: parameters examined; oxygen evolution and recombination; experimental test description; and effect of parameters on recombination rate.

  10. Short communication: A comparative analysis of recombinant chymosins.

    PubMed

    Vallejo, J A; Ageitos, J M; Poza, M; Villa, T G

    2012-02-01

    The first step in cheesemaking is the milk clotting process, in which κ-caseinolytic enzymes contribute to micelle precipitation. The best enzyme for this purpose is chymosin because of its high degree of specificity toward κ-casein. Although recombinant bovine chymosin is the most frequently used chymosin in the industry, new sources of recombinant chymosin, such as goat, camel, or buffalo, are now available. The present work represents a comparative study of 4 different recombinant chymosins (goat and buffalo chymosins expressed in Pichia pastoris, and bovine and camel chymosin expressed in Aspergillus niger). Recombinant goat chymosin exhibited the best catalytic efficiency compared with the buffalo, bovine, or camel recombinant enzymes. Moreover, recombinant goat chymosin exhibited the best specific proteolytic activity, a wider pH range of action, and a lower glycosylation degree than the other 3 enzymes. In conclusion, we propose that recombinant goat chymosin represents a serious alternative to recombinant bovine chymosin for use in the cheesemaking industry. PMID:22281325

  11. Heterogeneity in rates of recombination across the mouse genome

    SciTech Connect

    Nachman, M.W.; Churchill, G.A.

    1996-02-01

    If loci are randomly distributed on a physical map, the density of markers on a genetic map will be inversely proportional to recombination rate. First proposed by Mary Lyon, we have used this idea to estimate recombination rates from the Drosophila melanogaster linkage map. These results were compared with results of two other studies that estimated regional recombination rates in D. melanogaster using both physical and genetic maps. The three methods were largely concordant in identifying large-scale genomic patterns of recombination. The marker density method was then applied to the Mus musculus microsatellite linkage map. The distribution of microsatellites provided evidence for heterogeneity in recombination rates. Centromeric regions for several mouse chromosomes had significantly greater numbers of markers than expected, suggesting that recombination rates were lower in these regions. In contrast, most telomeric regions contained significantly fewer markers than expected. This indicates that recombination rates are elevated at the telomeres of many mouse chromosomes and is consistent with a comparison of the genetic and cytogenetic maps in these regions. The density of markers on a genetic map may provide a generally useful way to estimate regional recombination rates in species for which genetic, but not physical, maps are available. 44 refs., 5 figs., 4 tabs.

  12. A Heritable Recombination system for synthetic Darwinian evolution in yeast.

    PubMed

    Romanini, Dante W; Peralta-Yahya, Pamela; Mondol, Vanessa; Cornish, Virginia W

    2012-12-21

    Genetic recombination is central to the generation of molecular diversity and enhancement of evolutionary fitness in living systems. Methods such as DNA shuffling that recapitulate this diversity mechanism in vitro are powerful tools for engineering biomolecules with useful new functions by directed evolution. Synthetic biology now brings demand for analogous technologies that enable the controlled recombination of beneficial mutations in living cells. Thus, here we create a Heritable Recombination system centered around a library cassette plasmid that enables inducible mutagenesis via homologous recombination and subsequent combination of beneficial mutations through sexual reproduction in Saccharomyces cerevisiae. Using repair of nonsense codons in auxotrophic markers as a model, Heritable Recombination was optimized to give mutagenesis efficiencies of up to 6% and to allow successive repair of different markers through two cycles of sexual reproduction and recombination. Finally, Heritable Recombination was employed to change the substrate specificity of a biosynthetic enzyme, with beneficial mutations in three different active site loops crossed over three continuous rounds of mutation and selection to cover a total sequence diversity of 10(13). Heritable Recombination, while at an early stage of development, breaks the transformation barrier to library size and can be immediately applied to combinatorial crossing of beneficial mutations for cell engineering, adding important features to the growing arsenal of next generation molecular biology tools for synthetic biology. PMID:23412545

  13. Procedures for monitoring recombinant erythropoietin and analogs in doping.

    PubMed

    Lamon, Séverine; Robinson, Neil; Saugy, Martial

    2010-03-01

    Hemoglobin concentration is one of the principal factors of aerobic power and, consequently, of performance in many types of physical activities. The use of recombinant human erythropoietin is, therefore, particularly powerful for improving the physical performances of patients, and, more generally, improving their quality of life. This article discusses procedures for monitoring recombinant erythropoietin and its analogues in doping for athletic performance. PMID:20122455

  14. Inference of Ancestral Recombination Graphs through Topological Data Analysis.

    PubMed

    Cámara, Pablo G; Levine, Arnold J; Rabadán, Raúl

    2016-08-01

    The recent explosion of genomic data has underscored the need for interpretable and comprehensive analyses that can capture complex phylogenetic relationships within and across species. Recombination, reassortment and horizontal gene transfer constitute examples of pervasive biological phenomena that cannot be captured by tree-like representations. Starting from hundreds of genomes, we are interested in the reconstruction of potential evolutionary histories leading to the observed data. Ancestral recombination graphs represent potential histories that explicitly accommodate recombination and mutation events across orthologous genomes. However, they are computationally costly to reconstruct, usually being infeasible for more than few tens of genomes. Recently, Topological Data Analysis (TDA) methods have been proposed as robust and scalable methods that can capture the genetic scale and frequency of recombination. We build upon previous TDA developments for detecting and quantifying recombination, and present a novel framework that can be applied to hundreds of genomes and can be interpreted in terms of minimal histories of mutation and recombination events, quantifying the scales and identifying the genomic locations of recombinations. We implement this framework in a software package, called TARGet, and apply it to several examples, including small migration between different populations, human recombination, and horizontal evolution in finches inhabiting the Galápagos Islands. PMID:27532298

  15. Three Recombinant Engineered Antibodies against Recombinant Tags with High Affinity and Specificity

    PubMed Central

    Zhao, Hongyu; Shen, Ao; Xiang, Yang K.; Corey, David P.

    2016-01-01

    We describe three recombinant engineered antibodies against three recombinant epitope tags, constructed with divalent binding arms to recognize divalent epitopes and so achieve high affinity and specificity. In two versions, an epitope is inserted in tandem into a protein of interest, and a homodimeric antibody is constructed by fusing a high-affinity epitope-binding domain to a human or mouse Fc domain. In a third, a heterodimeric antibody is constructed by fusing two different epitope-binding domains which target two different binding sites in GFP, to polarized Fc fragments. These antibody/epitope pairs have affinities in the low picomolar range and are useful tools for many antibody-based applications. PMID:26943906

  16. Applications of recombinant DNA technology in the production of glycosylated recombinant human granulocyte colony stimulating factor.

    PubMed

    Holloway, C J

    1994-01-01

    Lenograstim has been developed by recombinant DNA technology and is expressed in large-scale mammalian cell culture. It has been shown that lenograstim is indistinguishable in its physicochemical, structural and biological properties with respect to native granulocyte colony stimulating factor isolated from a human cell line. In particular, both the recombinant and natural proteins have identical amino acid sequences, contain the same intra-polypeptide chain disulphide bridges and exhibit the same posttranslational carbohydrate structures. Lenograstim is manufactured by expanding inoculum from vials of the Manufacturer's Working Cell Bank (from molecular cloning) followed by culture in a large bioreactor. Purification of lenograstim involves a four-step chromatographic process. The active ingredient is monitored by in-process controls at all stages of manufacture and routinely as purified bulk. The finished product is formulated into excipients reflecting conditions close to the natural environment of the protein with respect to pH, osmolarity and the presence of human serum albumin. PMID:7535067

  17. Modulating Mek1 kinase alters outcomes of meiotic recombination and the stringency of the recombination checkpoint response

    PubMed Central

    Hsin-Yen, Wu; Hsuan-Chung, Ho; Burgess, Sean M.

    2010-01-01

    Summary Background During meiosis, recombination between homologous chromosomes promotes their proper segregation. In budding yeast, programmed double-strand breaks (DSBs) promote recombination between homologs versus sister chromatids by dimerizing and activating Mek1, a chromosome axis-associated kinase. Mek1 is also a proposed effector kinase in the recombination checkpoint that arrests exit from pachytene in response to aberrant DNA/axis structures. Elucidating a role for Mek1 in the recombination checkpoint has been difficult since in mek1 loss-of-function mutants DSBs are rapidly repaired using a sister chromatid thereby bypassing formation of checkpoint-activating lesions. Here we tested the hypothesis that a MEK1 gain-of-function allele would enhance interhomolog bias and the recombination checkpoint response. Results When Mek1 activation was artificially maintained through GST-mediated dimerization, there was an enhanced skew toward interhomolog recombination and reduction of intersister events including multi-chromatid joint molecules. Increased interhomolog events were specifically repaired as noncrossovers rather than crossovers. Ectopic Mek1 dimerization was also sufficient to impose interhomolog bias in the absence of recombination checkpoint functions, thereby uncoupling these two processes. Finally, the stringency of the recombination checkpoint was enhanced in weak meiotic recombination mutants by blocking prophase exit in a subset of cells where arrest is not absolute. Conclusions We propose that Mek1 plays dual roles during meiotic prophase I by phosphorylating targets directly involved in the recombination checkpoint as well as targets involved in sister chromatid recombination. We discuss how regulation of pachytene exit by Mek1 or similar kinases could influence checkpoint stringency, which may differ among species and between sexes. PMID:20888230

  18. Recombination in liquid xenon for low-energy recoils

    NASA Astrophysics Data System (ADS)

    Wang, Lu; Mei, Dongming; Cubed Collaboration

    2014-09-01

    Detector response to low-energy recoils in sub-keV region is critical to detection of low-mass dark matter particles-WIMPS (Weakly interacting massive particles). The role of electron-ion recombination is important to the interpretation of the relation between ionization yield and scintillation yield, which are in general anti-correlated. Recent experimental results show that ionization yield increases down to keV range. This phenomenon contradicts general understanding for low energy recoils in the keV range in which direct excitation dominates. The explanation is that recombination becomes much less efficient when the track length is smaller than the thermalization distance of electrons. However, recombination rate is also proportional to ionization density, which is very high for keV recoils. To understand how recombination rate behaves for keV recoils, we calculated both initial recombination rate and volume recombination rate for keV recoils in liquid xenon. In this paper, we show the results of the calculated recombination rate as a function of recoil energy for both electronic recoils and nuclear recoils. Detector response to low-energy recoils in sub-keV region is critical to detection of low-mass dark matter particles-WIMPS (Weakly interacting massive particles). The role of electron-ion recombination is important to the interpretation of the relation between ionization yield and scintillation yield, which are in general anti-correlated. Recent experimental results show that ionization yield increases down to keV range. This phenomenon contradicts general understanding for low energy recoils in the keV range in which direct excitation dominates. The explanation is that recombination becomes much less efficient when the track length is smaller than the thermalization distance of electrons. However, recombination rate is also proportional to ionization density, which is very high for keV recoils. To understand how recombination rate behaves for keV recoils

  19. Two dimensional Langevin recombination in regioregular poly(3-hexylthiophene)

    NASA Astrophysics Data System (ADS)

    Juška, Gytis; Genevičius, Kristijonas; Nekrašas, Nerijus; Sliaužys, Gytis; Österbacka, Ronald

    2009-07-01

    In this work, it is shown that recombination in regioregular poly(3-hexylthiophene):[6,6]-phenyl-C61-butyric acid methyl ester (RRP3HT:PCBM) bulk-heterojunction solar cells is caused by the two dimensional (2D) Langevin recombination in the lamellar structures of RRP3HT, which are formed after annealing process. Due to 2D Langevin process, bimolecular recombination coefficient is reduced in comparison with three dimensional Langevin case, and bimolecular recombination coefficient depends on the density of charge carriers n1/2. Data obtained from the different experimental techniques (charge extraction with linearly increasing voltage, integral time of flight, double injection current transients and transient absorption spectroscopy) confirms 2D Langevin recombination in RR3PHT.

  20. Charge carrier recombination dynamics in perovskite and polymer solar cells

    NASA Astrophysics Data System (ADS)

    Paulke, Andreas; Stranks, Samuel D.; Kniepert, Juliane; Kurpiers, Jona; Wolff, Christian M.; Schön, Natalie; Snaith, Henry J.; Brenner, Thomas J. K.; Neher, Dieter

    2016-03-01

    Time-delayed collection field experiments are applied to planar organometal halide perovskite (CH3NH3PbI3) based solar cells to investigate charge carrier recombination in a fully working solar cell at the nanosecond to microsecond time scale. Recombination of mobile (extractable) charges is shown to follow second-order recombination dynamics for all fluences and time scales tested. Most importantly, the bimolecular recombination coefficient is found to be time-dependent, with an initial value of ca. 10-9 cm3/s and a progressive reduction within the first tens of nanoseconds. Comparison to the prototypical organic bulk heterojunction device PTB7:PC71BM yields important differences with regard to the mechanism and time scale of free carrier recombination.

  1. Recombinant human bone morphogenetic protein induces bone formation.

    PubMed Central

    Wang, E A; Rosen, V; D'Alessandro, J S; Bauduy, M; Cordes, P; Harada, T; Israel, D I; Hewick, R M; Kerns, K M; LaPan, P

    1990-01-01

    We have purified and characterized active recombinant human bone morphogenetic protein (BMP) 2A. Implantation of the recombinant protein in rats showed that a single BMP can induce bone formation in vivo. A dose-response and time-course study using the rat ectopic bone formation assay revealed that implantation of 0.5-115 micrograms of partially purified recombinant human BMP-2A resulted in cartilage by day 7 and bone formation by day 14. The time at which bone formation occurred was dependent on the amount of BMP-2A implanted; at high doses bone formation could be observed at 5 days. The cartilage- and bone-inductive activity of the recombinant BMP-2A is histologically indistinguishable from that of bone extracts. Thus, recombinant BMP-2A has therapeutic potential to promote de novo bone formation in humans. Images PMID:2315314

  2. Sex recombination, and reproductive fitness: an experimental study using Paramecium

    SciTech Connect

    Nyberg, D.

    1982-08-01

    The effect of sex and recombination on reproductive fitness are measured using five wild stocks of Paramecium primaurelia. Among the wild stocks there were highly significant differences in growth rates. No hybrid had as low a fitness as the least fit parental stock. Recombination produced genotypes of higher fitness than those of either parent only in the cross between the two stocks of lowest fitness. The increase in variance of fitness as a result of recombination was almost exclusively attributable to the generation lines with low fitness. The fitness consequences of sexuality and mate choice were stock specific; some individuals leaving the most descendants by inbreeding, others by outcrossing. For most crosses the short-term advantage of sex, if any, accrue from the fusion of different gametes (hybrid vigor) and not from recombination. Since the homozygous genotype with the highest fitnes left the most progeny by inbreeding (no recombination), the persistence of conjugation in P. primaurelia is paradoxical. (JMT)

  3. Utilization of Site-Specific Recombination in Biopharmaceutical Production

    PubMed Central

    Ahmadi, Maryam; Damavandi, Narges; Akbari, Mohammad Reza; Davami, Fatemeh

    2016-01-01

    Mammalian expression systems, due to their capacity in post-translational modification, are preferred systems for biopharmaceutical protein production. Several recombinant protein systems have been introduced to the market, most of which are under clinical development. In spite of significant improvements such as cell line engineering, introducing novel expression methods, gene silencing and process development, expression level is unpredictable and unstable because of the random location of integration in the genome. Site-specific recombination techniques are capable of producing stable and high producer clonal cells; therefore, they are gaining more importance in the biopharmaceutical production. Site-specific recombination methods increase the recombinant protein production by specifically inserting a vector at a locus with specific expression trait. The present review focused on the latest developments in site-specific recombination techniques, their specific features and comparisons. PMID:26602035

  4. Production and secretion of recombinant proteins in Dictyostelium discoideum.

    PubMed

    Dittrich, W; Williams, K L; Slade, M B

    1994-06-01

    We have expressed useful amounts of three recombinant proteins in a new eukaryotic host/vector system. The cellular slime mold Dictyostelium discoideum efficiently secreted two recombinant products, a soluble form of the normally cell surface associated D. discoideum glycoprotein (PsA) and the heterologous protein glutathione-S-transferase (GST) from Schistosoma japonicum, while the enzyme beta-glucuronidase (GUS) from Escherichia coli was cell associated. Up to 20mg/l of recombinant PsA and 1mg/l of GST were obtained after purification from a standard, peptone based growth medium. The secretion signal peptide was correctly cleaved from the recombinant GST- and PsA-proteins and the expression of recombinant PsA was shown to be stable for at least one hundred generations in the absence of selection. PMID:7764951

  5. CRISPR-directed mitotic recombination enables genetic mapping without crosses.

    PubMed

    Sadhu, Meru J; Bloom, Joshua S; Day, Laura; Kruglyak, Leonid

    2016-05-27

    Linkage and association studies have mapped thousands of genomic regions that contribute to phenotypic variation, but narrowing these regions to the underlying causal genes and variants has proven much more challenging. Resolution of genetic mapping is limited by the recombination rate. We developed a method that uses CRISPR (clustered, regularly interspaced, short palindromic repeats) to build mapping panels with targeted recombination events. We tested the method by generating a panel with recombination events spaced along a yeast chromosome arm, mapping trait variation, and then targeting a high density of recombination events to the region of interest. Using this approach, we fine-mapped manganese sensitivity to a single polymorphism in the transporter Pmr1. Targeting recombination events to regions of interest allows us to rapidly and systematically identify causal variants underlying trait differences. PMID:27230379

  6. High cell density cultivation of recombinant Escherichia coli for prodrug of recombinant human GLPs production.

    PubMed

    Zhou, Ying; Ma, Xue; Hou, Zheng; Xue, Xiaoyan; Meng, Jingru; Li, Mingkai; Jia, Min; Luo, Xiaoxing

    2012-09-01

    Glucagon-like peptide-1 (GLP-1)(2) has been attracting increasing interest on account of its prominent benefits in type 2 diabetes. However, its clinical applications are limited by the short half-life in vivo. To overcome this limitation, a new polymer of GLP-1 was developed by prodrug strategy. In this study a recombinant protein, rhGLPs, was successfully constructed, cloned into plasmid pET30a (+) and expressed in Escherichia coli ArcticExpress(DE3)RP in the form of inclusion body. The recombinant fusion protein productivity could be enhanced by high cell density culture of the recombinant strain. As a result, about 40 g wet weight cells per liter were obtained. The protein was purified by size-exclusion chromatography on a Superdex 75 column and refolded using reverse dilution and dialysis methods. SDS-PAGE, HPLC and MALDI-TOF mass spectrometry were undertaken to determine the purity and molecular weight of rhGLPs. Bioactivity assay revealed that it had glucose-lowering and insulin-releasing action in vivo. PMID:22771632

  7. Recombinational Landscape and Population Genomics of Caenorhabditis elegans

    PubMed Central

    Rockman, Matthew V.; Kruglyak, Leonid

    2009-01-01

    Recombination rate and linkage disequilibrium, the latter a function of population genomic processes, are the critical parameters for mapping by linkage and association, and their patterns in Caenorhabditis elegans are poorly understood. We performed high-density SNP genotyping on a large panel of recombinant inbred advanced intercross lines (RIAILs) of C. elegans to characterize the landscape of recombination and, on a panel of wild strains, to characterize population genomic patterns. We confirmed that C. elegans autosomes exhibit discrete domains of nearly constant recombination rate, and we show, for the first time, that the pattern holds for the X chromosome as well. The terminal domains of each chromosome, spanning about 7% of the genome, exhibit effectively no recombination. The RIAILs exhibit a 5.3-fold expansion of the genetic map. With median marker spacing of 61 kb, they are a powerful resource for mapping quantitative trait loci in C. elegans. Among 125 wild isolates, we identified only 41 distinct haplotypes. The patterns of genotypic similarity suggest that some presumed wild strains are laboratory contaminants. The Hawaiian strain, CB4856, exhibits genetic isolation from the remainder of the global population, whose members exhibit ample evidence of intercrossing and recombining. The population effective recombination rate, estimated from the pattern of linkage disequilibrium, is correlated with the estimated meiotic recombination rate, but its magnitude implies that the effective rate of outcrossing is extremely low, corroborating reports of selection against recombinant genotypes. Despite the low population, effective recombination rate and extensive linkage disequilibrium among chromosomes, which are techniques that account for background levels of genomic similarity, permit association mapping in wild C. elegans strains. PMID:19283065

  8. Genetic recombination of bacterial plasmid DNA: effect of RecF pathway mutations on plasmid recombination in Escherichia coli.

    PubMed Central

    Kolodner, R; Fishel, R A; Howard, M

    1985-01-01

    Tn5 insertion mutations in the recN gene, and in what appears to be a new RecF pathway gene designated recO and mapping at approximately 55.4 min on the standard genetic map, were isolated by screening Tn5 insertion mutations that cotransduced with tyrA. The recO1504::Tn5 mutation decreased the frequency of recombination during Hfr-mediated crosses and increased the susceptibility to killing by UV irradiation and mitomycin C when present in a recB recC sbcB background, but only increased the sensitivity to killing by UV irradiation when present in an otherwise Rec+ background. The effects of these and other RecF pathway mutations on plasmid recombination were tested. Mutations in the recJ, recO, and ssb genes, when present in otherwise Rec+ E. coli strains, decreased the frequency of plasmid recombination, whereas the lexA3, recAo281, recN, and ruv mutations had no effect on plasmid recombination. Tn5 insertion mutations in the lexA gene increased the frequency of plasmid recombination. These data indicate that plasmid recombination events in wild-type Escherichia coli strains are catalyzed by a recombination pathway that is related to the RecF recombination pathway and that some component of this pathway besides the recA gene product is regulated by the lexA gene product. PMID:2993230

  9. Genome Sequence of Complex HIV-1 Unique Recombinant Forms Sharing a Common Recombination Breakpoint Identified in Malaysia

    PubMed Central

    Cheong, Hui Ting; Ng, Kim Tien; Ong, Lai Yee; Takebe, Yutaka; Chan, Kok Gan; Koh, Clayton; Al-Darraji, Haider Abdulrazzaq Abed; Kamarulzaman, Adeeba

    2015-01-01

    Three strains of HIV-1 unique recombinant forms (URFs) descended from subtypes B, B′, and CRF01_AE were identified among people who inject drugs in Kuala Lumpur, Malaysia. These three URFs shared a common recombination breakpoint in the reverse transcriptase region, indicating frequent linkage within the drug-injecting networks in Malaysia. PMID:26543107

  10. 75 FR 42114 - Office of Biotechnology Activities; Recombinant DNA Research: Proposed Action Under the NIH...

    Federal Register 2010, 2011, 2012, 2013, 2014

    2010-07-20

    ... HUMAN SERVICES National Institutes of Health Office of Biotechnology Activities; Recombinant DNA Research: Proposed Action Under the NIH Guidelines for Research Involving Recombinant DNA Molecules (NIH... transgenic rodents by recombinant DNA technology must be registered with the Institutional...

  11. Effect of sex, age, and breed on genetic recombination features in cattle

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Meiotic recombination is a fundamental biological process which generates genetic diversity, affects fertility, and influences evolvability. Here we investigate the roles of sex, age, and breed in cattle recombination features, including recombination rate, location and crossover interference. Usin...

  12. Auger recombination as the dominant recombination process in indium nitride at low temperatures during steady-state photoluminescence

    SciTech Connect

    Seetoh, I. P.; Soh, C. B.; Fitzgerald, E. A.; Chua, S. J.

    2013-03-11

    Auger recombination in InN films grown by metal-organic chemical vapor deposition was studied by steady-state photoluminescence at different laser excitation powers and sample temperatures. It was dominant over radiative recombination and Shockley-Read-Hall recombination at low temperatures, contributing to the sub-linear relationship between the integrated photoluminescence intensity and laser excitation power. Auger recombination rates increased gradually with temperature with an activation energy of 10-17 meV, in good agreement with values from transient photoluminescence reported in literature. As the Auger recombination rates were independent of material quality, they may form an upper limit to the luminous efficiency of InN.

  13. Auger recombination as the dominant recombination process in indium nitride at low temperatures during steady-state photoluminescence

    NASA Astrophysics Data System (ADS)

    Seetoh, I. P.; Soh, C. B.; Fitzgerald, E. A.; Chua, S. J.

    2013-03-01

    Auger recombination in InN films grown by metal-organic chemical vapor deposition was studied by steady-state photoluminescence at different laser excitation powers and sample temperatures. It was dominant over radiative recombination and Shockley-Read-Hall recombination at low temperatures, contributing to the sub-linear relationship between the integrated photoluminescence intensity and laser excitation power. Auger recombination rates increased gradually with temperature with an activation energy of 10-17 meV, in good agreement with values from transient photoluminescence reported in literature. As the Auger recombination rates were independent of material quality, they may form an upper limit to the luminous efficiency of InN.

  14. Retroviral Vectors for Analysis of Viral Mutagenesis and Recombination

    PubMed Central

    Rawson, Jonathan M.O.; Mansky, Louis M.

    2014-01-01

    Retrovirus population diversity within infected hosts is commonly high due in part to elevated rates of replication, mutation, and recombination. This high genetic diversity often complicates the development of effective diagnostics, vaccines, and antiviral drugs. This review highlights the diverse vectors and approaches that have been used to examine mutation and recombination in retroviruses. Retroviral vectors for these purposes can broadly be divided into two categories: those that utilize reporter genes as mutation or recombination targets and those that utilize viral genes as targets of mutation or recombination. Reporter gene vectors greatly facilitate the detection, quantification, and characterization of mutants and/or recombinants, but may not fully recapitulate the patterns of mutagenesis or recombination observed in native viral gene sequences. In contrast, the detection of mutations or recombination events directly in viral genes is more biologically relevant but also typically more challenging and inefficient. We will highlight the advantages and disadvantages of the various vectors and approaches used as well as propose ways in which they could be improved. PMID:25254386

  15. Calculations of effective recombination coefficients for nebular astrophysics

    NASA Astrophysics Data System (ADS)

    Fang, Xuan; Liu, Xiaowei; Storey, Pete J.

    2015-08-01

    In the seemingly well established field of nebular astrophysics, there has been a long-standing dichotomy in plasma diagnostics and heavy elemental abundance determinations using the traditional method based on collisionally excited lines on the one hand, and optical recombination lines/continua, on the other. Several mechanisms have been proposed to explain this fundamental problem. Deep spectroscopy and recombination line analysis of emission line nebulae (planetary nebulae and H II regions) in the past decade have pointed to the existence of another previously unknown component of cold, H-deficient material as the culprit. Better constraints are needed on the physical conditions (electron temperature and density), chemical composition, mass, and spatial distribution of the postulated H-deficient inclusions in order to unravel their astrophysical origins. This requires knowledge of the relevant atomic parameters, most importantly the effective recombination coefficients of abundant heavy element ions, such as C II, O II, N II, and Ne II, appropriate for the physical conditions prevailing in those cold inclusions (e.g., electron temperature Te < 1000 K).In this contribution, I will introduce the creation of new effective recombination coefficients for the heavy element optical recombination lines, and review the recent progress in nebular astrophysics since the availability of new and high-quality atomic data. I will also present our new calculations of the effective recombination coefficients for the Ne II recombination line spectrum.

  16. Phylogenetic and Recombination Analysis of Tomato Spotted Wilt Virus

    PubMed Central

    Yu, Jisuk; Kim, Mi-Kyeong; Choi, Hong-Soo; Kim, Kook-Hyung

    2013-01-01

    Tomato spotted wilt virus (TSWV) severely damages and reduces the yield of many economically important plants worldwide. In this study, we determined the whole-genome sequences of 10 TSWV isolates recently identified from various regions and hosts in Korea. Phylogenetic analysis of these 10 isolates as well as the three previously sequenced isolates indicated that the 13 Korean TSWV isolates could be divided into two groups reflecting either two different origins or divergences of Korean TSWV isolates. In addition, the complete nucleotide sequences for the 13 Korean TSWV isolates along with previously sequenced TSWV RNA segments from Korea and other countries were subjected to phylogenetic and recombination analysis. The phylogenetic analysis indicated that both the RNA L and RNA M segments of most Korean isolates might have originated in Western Europe and North America but that the RNA S segments for all Korean isolates might have originated in China and Japan. Recombination analysis identified a total of 12 recombination events among all isolates and segments and five recombination events among the 13 Korea isolates; among the five recombinants from Korea, three contained the whole RNA L segment, suggesting reassortment rather than recombination. Our analyses provide evidence that both recombination and reassortment have contributed to the molecular diversity of TSWV. PMID:23696821

  17. The influence of recombination on human genetic diversity.

    PubMed

    Spencer, Chris C A; Deloukas, Panos; Hunt, Sarah; Mullikin, Jim; Myers, Simon; Silverman, Bernard; Donnelly, Peter; Bentley, David; McVean, Gil

    2006-09-22

    In humans, the rate of recombination, as measured on the megabase scale, is positively associated with the level of genetic variation, as measured at the genic scale. Despite considerable debate, it is not clear whether these factors are causally linked or, if they are, whether this is driven by the repeated action of adaptive evolution or molecular processes such as double-strand break formation and mismatch repair. We introduce three innovations to the analysis of recombination and diversity: fine-scale genetic maps estimated from genotype experiments that identify recombination hotspots at the kilobase scale, analysis of an entire human chromosome, and the use of wavelet techniques to identify correlations acting at different scales. We show that recombination influences genetic diversity only at the level of recombination hotspots. Hotspots are also associated with local increases in GC content and the relative frequency of GC-increasing mutations but have no effect on substitution rates. Broad-scale association between recombination and diversity is explained through covariance of both factors with base composition. To our knowledge, these results are the first evidence of a direct and local influence of recombination hotspots on genetic variation and the fate of individual mutations. However, that hotspots have no influence on substitution rates suggests that they are too ephemeral on an evolutionary time scale to have a strong influence on broader scale patterns of base composition and long-term molecular evolution. PMID:17044736

  18. Mechanics and Single-Molecule Interrogation of DNA Recombination.

    PubMed

    Bell, Jason C; Kowalczykowski, Stephen C

    2016-06-01

    The repair of DNA by homologous recombination is an essential, efficient, and high-fidelity process that mends DNA lesions formed during cellular metabolism; these lesions include double-stranded DNA breaks, daughter-strand gaps, and DNA cross-links. Genetic defects in the homologous recombination pathway undermine genomic integrity and cause the accumulation of gross chromosomal abnormalities-including rearrangements, deletions, and aneuploidy-that contribute to cancer formation. Recombination proceeds through the formation of joint DNA molecules-homologously paired but metastable DNA intermediates that are processed by several alternative subpathways-making recombination a versatile and robust mechanism to repair damaged chromosomes. Modern biophysical methods make it possible to visualize, probe, and manipulate the individual molecules participating in the intermediate steps of recombination, revealing new details about the mechanics of genetic recombination. We review and discuss the individual stages of homologous recombination, focusing on common pathways in bacteria, yeast, and humans, and place particular emphasis on the molecular mechanisms illuminated by single-molecule methods. PMID:27088880

  19. Identification and Manipulation of the Molecular Determinants Influencing Poliovirus Recombination

    PubMed Central

    Runckel, Charles; Westesson, Oscar; Andino, Raul; DeRisi, Joseph L.

    2013-01-01

    The control and prevention of communicable disease is directly impacted by the genetic mutability of the underlying etiological agents. In the case of RNA viruses, genetic recombination may impact public health by facilitating the generation of new viral strains with altered phenotypes and by compromising the genetic stability of live attenuated vaccines. The landscape of homologous recombination within a given RNA viral genome is thought to be influenced by several factors; however, a complete understanding of the genetic determinants of recombination is lacking. Here, we utilize gene synthesis and deep sequencing to create a detailed recombination map of the poliovirus 1 coding region. We identified over 50 thousand breakpoints throughout the genome, and we show the majority of breakpoints to be concentrated in a small number of specific “hotspots,” including those associated with known or predicted RNA secondary structures. Nucleotide base composition was also found to be associated with recombination frequency, suggesting that recombination is modulated across the genome by predictable and alterable motifs. We tested the predictive utility of the nucleotide base composition association by generating an artificial hotspot in the poliovirus genome. Our results imply that modification of these motifs could be extended to whole genome re-designs for the development of recombination-deficient, genetically stable live vaccine strains. PMID:23408891

  20. A Glance at Recombination Hotspots in the Domestic Cat.

    PubMed

    Alhaddad, Hasan; Zhang, Chi; Rannala, Bruce; Lyons, Leslie A

    2016-01-01

    Recombination has essential roles in increasing genetic variability within a population and in ensuring successful meiotic events. The objective of this study is to (i) infer the population-scaled recombination rate (ρ), and (ii) identify and characterize regions of increased recombination rate for the domestic cat, Felis silvestris catus. SNPs (n = 701) were genotyped in twenty-two East Asian feral cats (random bred). The SNPs covered ten different chromosomal regions (A1, A2, B3, C2, D1, D2, D4, E2, F2, X) with an average region size of 850 Kb and an average SNP density of 70 SNPs/region. The Bayesian method in the program inferRho was used to infer regional population recombination rates and hotspots localities. The regions exhibited variable population recombination rates and four decisive recombination hotspots were identified on cat chromosome A2, D1, and E2 regions. As a description of the identified hotspots, no correlation was detected between the GC content and the locality of recombination spots, and the hotspots enclosed L2 LINE elements and MIR and tRNA-Lys SINE elements. PMID:26859385

  1. Monte Carlo Simulations of the Inside Intron Recombination

    NASA Astrophysics Data System (ADS)

    Cebrat, Stanisław; PȨKALSKI, Andrzej; Scharf, Fabian

    Biological genomes are divided into coding and non-coding regions. Introns are non-coding parts within genes, while the remaining non-coding parts are intergenic sequences. To study evolutionary significance of the inside intron recombination we have used two models based on the Monte Carlo method. In our computer simulations we have implemented the internal structure of genes by declaring the probability of recombination between exons. One situation when inside intron recombination is advantageous is recovering functional genes by combining proper exons dispersed in the genetic pool of the population after a long period without selection for the function of the gene. Populations have to pass through the bottleneck, then. These events are rather rare and we have expected that there should be other phenomena giving profits from the inside intron recombination. In fact we have found that inside intron recombination is advantageous only in the case when after recombination, besides the recombinant forms, parental haplotypes are available and selection is set already on gametes.

  2. Data Mining for Expressivity of Recombinant Protein Expression

    NASA Astrophysics Data System (ADS)

    Kira, Satoshi; Isoai, Atsushi; Yamamura, Masayuki

    We analyzed the expressivity of recombinant proteins by using data mining methods. The expression technique of recombinant protein is a key step towards elucidating the functions of genes discovered through genomic sequence projects. We have studied the productive efficiency of recombinant proteins in fission yeast, Schizosaccharomyces pombe (S.pombe), by mining the expression results. We gathered 57 proteins whose expression levels were known roughly in the host. Correlation analysis, principal component analysis and decision tree analysis were applied to these expression data. Analysis featuring codon usage and amino acid composition clarified that the amino acid composition affected to the expression levels of a recombinant protein strongly than the effect of codon usage. Furthermore, analysis of amino acid composition showed that protein solubility and the metabolism cost of amino acids correlated with a protein expressivity. Codon usage was often interesting in the field of recombinant expressions. However, our analysis found the weak correlation codon features with expressivities. These results indicated that ready-made indices of codon bias were irrelevant ones for modeling the expressivities of recombinant proteins. Our data driven approach was an easy and powerful method to improve recombinant protein expression, and this approach should be concentrated attention with the huge amount of expression data accumulating through the post-genome era.

  3. Recombination-mediated genetic engineering of large genomic DNA transgenes.

    PubMed

    Ejsmont, Radoslaw Kamil; Ahlfeld, Peter; Pozniakovsky, Andrei; Stewart, A Francis; Tomancak, Pavel; Sarov, Mihail

    2011-01-01

    Faithful gene activity reporters are a useful tool for evo-devo studies enabling selective introduction of specific loci between species and assaying the activity of large gene regulatory sequences. The use of large genomic constructs such as BACs and fosmids provides an efficient platform for exploration of gene function under endogenous regulatory control. Despite their large size they can be easily engineered using in vivo homologous recombination in Escherichia coli (recombineering). We have previously demonstrated that the efficiency and fidelity of recombineering are sufficient to allow high-throughput transgene engineering in liquid culture, and have successfully applied this approach in several model systems. Here, we present a detailed protocol for recombineering of BAC/fosmid transgenes for expression of fluorescent or affinity tagged proteins in Drosophila under endogenous in vivo regulatory control. The tag coding sequence is seamlessly recombineered into the genomic region contained in the BAC/fosmid clone, which is then integrated into the fly genome using ϕC31 recombination. This protocol can be easily adapted to other recombineering projects. PMID:22065454

  4. A Glance at Recombination Hotspots in the Domestic Cat

    PubMed Central

    Alhaddad, Hasan; Zhang, Chi; Rannala, Bruce; Lyons, Leslie A.

    2016-01-01

    Recombination has essential roles in increasing genetic variability within a population and in ensuring successful meiotic events. The objective of this study is to (i) infer the population-scaled recombination rate (ρ), and (ii) identify and characterize regions of increased recombination rate for the domestic cat, Felis silvestris catus. SNPs (n = 701) were genotyped in twenty-two East Asian feral cats (random bred). The SNPs covered ten different chromosomal regions (A1, A2, B3, C2, D1, D2, D4, E2, F2, X) with an average region size of 850 Kb and an average SNP density of 70 SNPs/region. The Bayesian method in the program inferRho was used to infer regional population recombination rates and hotspots localities. The regions exhibited variable population recombination rates and four decisive recombination hotspots were identified on cat chromosome A2, D1, and E2 regions. As a description of the identified hotspots, no correlation was detected between the GC content and the locality of recombination spots, and the hotspots enclosed L2 LINE elements and MIR and tRNA-Lys SINE elements. PMID:26859385

  5. The Contribution of Genetic Recombination to CRISPR Array Evolution

    PubMed Central

    Kupczok, Anne; Landan, Giddy; Dagan, Tal

    2015-01-01

    CRISPR (clustered regularly interspaced short palindromic repeats) is a microbial immune system against foreign DNA. Recognition sequences (spacers) encoded within the CRISPR array mediate the immune reaction in a sequence-specific manner. The known mechanisms for the evolution of CRISPR arrays include spacer acquisition from foreign DNA elements at the time of invasion and array erosion through spacer deletion. Here, we consider the contribution of genetic recombination between homologous CRISPR arrays to the evolution of spacer repertoire. Acquisition of spacers from exogenic arrays via recombination may confer the recipient with immunity against unencountered antagonists. For this purpose, we develop a novel method for the detection of recombination in CRISPR arrays by modeling the spacer order in arrays from multiple strains from the same species. Because the evolutionary signal of spacer recombination may be similar to that of pervasive spacer deletions or independent spacer acquisition, our method entails a robustness analysis of the recombination inference by a statistical comparison to resampled and perturbed data sets. We analyze CRISPR data sets from four bacterial species: two Gammaproteobacteria species harboring CRISPR type I and two Streptococcus species harboring CRISPR type II loci. We find that CRISPR array evolution in Escherichia coli and Streptococcus agalactiae can be explained solely by vertical inheritance and differential spacer deletion. In Pseudomonas aeruginosa, we find an excess of single spacers potentially incorporated into the CRISPR locus during independent acquisition events. In Streptococcus thermophilus, evidence for spacer acquisition by recombination is present in 5 out of 70 strains. Genetic recombination has been proposed to accelerate adaptation by combining beneficial mutations that arose in independent lineages. However, for most species under study, we find that CRISPR evolution is shaped mainly by spacer acquisition and

  6. Recombinant phage probes for Listeria monocytogenes

    NASA Astrophysics Data System (ADS)

    Carnazza, S.; Gioffrè, G.; Felici, F.; Guglielmino, S.

    2007-10-01

    Monitoring of food and environmental samples for biological threats, such as Listeria monocytogenes, requires probes that specifically bind biological agents and ensure their immediate and efficient detection. There is a need for robust and inexpensive affinity probes as an alternative to antibodies. These probes may be recruited from random peptide libraries displayed on filamentous phage. In this study, we selected from two phage peptide libraries phage clones displaying peptides capable of specific and strong binding to the L. monocytogenes cell surface. The ability of isolated phage clones to interact specifically with L. monocytogenes was demonstrated using enzyme-linked immunosorbent assay (ELISA) and confirmed by co-precipitation assay. We also assessed the sensitivity of phage-bacteria binding by PCR on phage-captured Listeria cells, which could be detected at a concentration of 104 cells ml-1. In addition, as proof-of-concept, we tested the possibility of immobilizing the affinity-selected phages to a putative biosensor surface. The quality of phage deposition was monitored by ELISA and fluorescent microscopy. Phage-bacterial binding was confirmed by high power optical phase contrast microscopy. Overall, the results of this work validate the concept of affinity-selected recombinant filamentous phages as probes for detecting and monitoring bacterial agents under any conditions that warrant their recognition, including in food products.

  7. Recombinant albumin monolayers on latex particles.

    PubMed

    Sofińska, Kamila; Adamczyk, Zbigniew; Kujda, Marta; Nattich-Rak, Małgorzata

    2014-01-14

    The adsorption of recombinant human serum albumin (rHSA) on negatively charged polystyrene latex micro-particles was studied at pH 3.5 and the NaCl concentration range of 10(-3) to 0.15 M. The electrophoretic mobility of latex monotonically increased with the albumin concentration in the suspension. The coverage of adsorbed albumin was quantitatively determined using the depletion method, where the residual protein concentration was determined by electrokinetic measurements and AFM imaging. It was shown that albumin adsorption was irreversible. Its maximum coverage on latex varied between 0.7 mg m(-2) for 10(-3) M NaCl to 1.3 mg m(-2) for 0.15 M NaCl. The latter value matches the maximum coverage previously determined for human serum albumin on mica using the streaming potential method. The increase in the maximum coverage was interpreted in terms of reduced electrostatic repulsion among adsorbed molecules. These facts confirm that albumin adsorption at pH 3.5 is governed by electrostatic interactions and proceeds analogously to colloid particle deposition. The stability of albumin monolayers was measured in additional experiments where changes in the latex electrophoretic mobility and the concentration of free albumin in solutions were monitored over prolonged time periods. Based on these experimental data, a robust procedure of preparing albumin monolayers on latex particles of well-controlled coverage and molecule distribution was proposed. PMID:24354916

  8. Developing recombinant antibodies for biomarker detection

    SciTech Connect

    Baird, Cheryl L.; Fischer, Christopher J.; Pefaur, Noah B.; Miller, Keith D.; Kagen, Jacob; Srivastava, Sudhir; Rodland, Karin D.

    2010-10-01

    Monoclonal antibodies (mAbs) have an essential role in biomarker validation and diagnostic assays. A barrier to pursuing these applications is the reliance on immunization and hybridomas to produce mAbs, which is time-consuming and may not yield the desired mAb. We recommend a process flow for affinity reagent production that utilizes combinatorial protein display systems (eg, yeast surface display or phage display) rather than hybridomas. These systems link a selectable phenotype-binding conferred by an antibody fragment-with a means for recovering the encoding gene. Recombinant libraries obtained from immunizations can produce high-affinity antibodies (<10 nM) more quickly than other methods. Non-immune libraries provide an alternate route when immunizations are not possible, or when suitable mAbs are not recovered from an immune library. Directed molecular evolution (DME) is an integral part of optimizing mAbs obtained from combinatorial protein display, but can also be used on hybridoma-derived mAbs. Variants can easily be obtained and screened to increase the affinity of the parent mAb (affinity maturation). We discuss examples where DME has been used to tailor affinity reagents to specific applications. Combinatorial protein display also provides an accessible method for identifying antibody pairs, which are necessary for sandwich-type diagnostic assays.

  9. Multivalent Recombinant Protein Vaccine against Coccidioidomycosis

    PubMed Central

    Tarcha, Eric J.; Basrur, Venkatesha; Hung, Chiung-Yu; Gardner, Malcolm J.; Cole, Garry T.

    2006-01-01

    Coccidioidomycosis is a human respiratory disease that is endemic to the southwestern United States and is caused by inhalation of the spores of a desert soilborne fungus. Efforts to develop a vaccine against this disease have focused on identification of T-cell-reactive antigens derived from the parasitic cell wall which can stimulate protective immunity against Coccidioides posadasii infection in mice. We previously described a productive immunoproteomic/bioinformatic approach to the discovery of vaccine candidates which makes use of the translated genome of C. posadasii and a computer-based method of scanning deduced sequences of seroreactive proteins for epitopes that are predicted to bind to human major histocompatibility (MHC) class II-restricted molecules. In this study we identified a set of putative cell wall proteins predicted to contain multiple, promiscuous MHC II binding epitopes. Three of these were expressed by Escherichia coli, combined in a vaccine, and tested for protective efficacy in C57BL/6 mice. Approximately 90% of the mice survived beyond 90 days after intranasal challenge, and the majority cleared the pathogen. We suggest that the multicomponent vaccine stimulates a broader range of T-cell clones than the single recombinant protein vaccines and thereby may be capable of inducing protection in an immunologically heterogeneous human population. PMID:16988258

  10. Dielectronic Recombination In Active Galactic Nuclei

    NASA Technical Reports Server (NTRS)

    Lukic, D. V.; Schnell, M.; Savin, D. W.; Altun, Z.; Badnell, N.; Brandau, C.; Schmidt, E. W.; Mueller, A.; Schippers, S.; Sprenger, F.; Lestinsky, M.; Wolf, A.

    2006-01-01

    XMM-Newton and Chandra observations of active galactic nuclei (AGN) show rich spectra of X-ray absorption lines. These observations have detected a broad unresolved transition array (UTA) between approx. 15-17 A. This is attributed to inner-shell photoexcitation of M-shell iron ions. Modeling these UTA features is currently limited by uncertainties in the low-temperature dielectronic recombination (DR) data for M-shell iron. In order to resolve this issue, and to provide reliable iron M-shell DR data for plasma modeling, we are carrying out a series of laboratory measurements using the heavy-ion Test Storage Ring (TSR) at the Max-Plank-Institute for Nuclear Physics in Heidelberg, Germany. Currently, laboratory measurements of low temperature DR can only be performed at storage rings. We use the DR data obtained at TSR, to calculate rate coefficients for plasma modeling and to benchmark theoretical DR calculations. Here we report our recent experimental results for DR of Fe XIV forming Fe XIII.

  11. Development of recombinant vaccines for botulinum neurotoxin.

    PubMed

    Smith, L A

    1998-11-01

    Synthetic genes encoding non-toxic, carboxyl-terminal regions (approximately 50 kDa) of botulinum neurotoxin (BoNT) serotypes A and B (referred to as fragment C or HC) were constructed and cloned into the methylotropic yeast, Pichia pastoris. Genes specifying BoNTA(HC) and BoNTB(HC) were expressed as both intracellular and secreted products. Recombinants, expressed intracellularly, yielded products with the expected molecular weight as judged by SDS PAGE and Western blot (immunoblot) analysis, while secreted products were larger due to glycosylation. Gene products were used to vaccinate mice and evaluated for their ability to elicit protective antibody titers in vivo. Mice given three intramuscular vaccinations with yeast supernatant containing glycosylated BoNTA(HC) were protected against an intraperitoneal challenge of 10(6) 50% mouse lethal doses (MLD50) of serotype A neurotoxin, a result not duplicated by its BoNTB(HC) counterpart. Vaccinating mice with cytoplasmically produced BoNTA(HC) and BoNTB(HC) protected animals from a challenge of 10(6) MLD50 of serotype A and B toxins, respectively. Because of the glycosylation encountered with secreted BoNT(HC), our efforts focused on the production and purification of products from intracellular expression. PMID:9792170

  12. HSV Recombinant Vectors for Gene Therapy

    PubMed Central

    Manservigi, Roberto; Argnani, Rafaela; Marconi, Peggy

    2010-01-01

    The very deep knowledge acquired on the genetics and molecular biology of herpes simplex virus (HSV), has allowed the development of potential replication-competent and replication-defective vectors for several applications in human healthcare. These include delivery and expression of human genes to cells of the nervous systems, selective destruction of cancer cells, prophylaxis against infection with HSV or other infectious diseases, and targeted infection to specific tissues or organs. Replication-defective recombinant vectors are non-toxic gene transfer tools that preserve most of the neurotropic features of wild type HSV-1, particularly the ability to express genes after having established latent infections, and are thus proficient candidates for therapeutic gene transfer settings in neurons. A replication-defective HSV vector for the treatment of pain has recently entered in phase 1 clinical trial. Replication-competent (oncolytic) vectors are becoming a suitable and powerful tool to eradicate brain tumours due to their ability to replicate and spread only within the tumour mass, and have reached phase II/III clinical trials in some cases. The progress in understanding the host immune response induced by the vector is also improving the use of HSV as a vaccine vector against both HSV infection and other pathogens. This review briefly summarizes the obstacle encountered in the delivery of HSV vectors and examines the various strategies developed or proposed to overcome such challenges. PMID:20835362

  13. Epigenetic Codes Programing Class Switch Recombination

    PubMed Central

    Vaidyanathan, Bharat; Chaudhuri, Jayanta

    2015-01-01

    Class switch recombination imparts B cells with a fitness-associated adaptive ­advantage during a humoral immune response by using a precision-tailored DNA excision and ligation process to swap the default constant region gene of the antibody with a new one that has unique effector functions. This secondary diversification of the antibody repertoire is a hallmark of the adaptability of B cells when confronted with environmental and pathogenic challenges. Given that the nucleotide sequence of genes during class switching remains unchanged (genetic constraints), it is logical and necessary therefore, to integrate the adaptability of B cells to an epigenetic state, which is dynamic and can be heritably modulated before, after, or even during an antibody-dependent immune response. Epigenetic regulation encompasses heritable changes that affect function (phenotype) without altering the sequence information embedded in a gene, and include histone, DNA and RNA modifications. Here, we review current literature on how B cells use an epigenetic code language as a means to ensure antibody plasticity in light of pathogenic insults. PMID:26441954

  14. Therapeutic Use of Native and Recombinant Enteroviruses.

    PubMed

    Ylä-Pelto, Jani; Tripathi, Lav; Susi, Petri

    2016-03-01

    Research on human enteroviruses has resulted in the identification of more than 100 enterovirus types, which use more than 10 protein receptors and/or attachment factors required in cell binding and initiation of the replication cycle. Many of these "viral" receptors are overexpressed in cancer cells. Receptor binding and the ability to replicate in specific target cells define the tropism and pathogenesis of enterovirus types, because cellular infection often results in cytolytic response, i.e., disruption of the cells. Viral tropism and cytolytic properties thus make native enteroviruses prime candidates for oncolytic virotherapy. Copy DNA cloning and modification of enterovirus genomes have resulted in the generation of enterovirus vectors with properties that are useful in therapy or in vaccine trials where foreign antigenic epitopes are expressed from or on the surface of the vector virus. The small genome size and compact particle structure, however, set limits to enterovirus genome modifications. This review focuses on the therapeutic use of native and recombinant enteroviruses and the methods that have been applied to modify enterovirus genomes for therapy. PMID:26907330

  15. Recombinant Canine Coronaviruses in Dogs, Europe

    PubMed Central

    Mari, Viviana; Elia, Gabriella; Addie, Diane D.; Camero, Michele; Lucente, Maria Stella; Martella, Vito; Buonavoglia, Canio

    2010-01-01

    Coronaviruses of potential recombinant origin with porcine transmissible gastroenteritis virus (TGEV), referred to as a new subtype (IIb) of canine coronavirus (CCoV), were recently identified in dogs in Europe. To assess the distribution of the TGEV-like CCoV subtype, during 2001–2008 we tested fecal samples from dogs with gastroenteritis. Of 1,172 samples, 493 (42.06%) were positive for CCoV. CCoV-II was found in 218 samples, and CCoV-I and CCoV-II genotypes were found in 182. Approximately 20% of the samples with CCoV-II had the TGEV-like subtype; detection rates varied according to geographic origin. The highest and lowest rates of prevalence for CCoV-II infection were found in samples from Hungary and Greece (96.87% and 3.45%, respectively). Sequence and phylogenetic analyses showed that the CCoV-IIb strains were related to prototype TGEV-like strains in the 5′ and the 3′ ends of the spike protein gene. PMID:20031041

  16. Therapeutic Use of Native and Recombinant Enteroviruses

    PubMed Central

    Ylä-Pelto, Jani; Tripathi, Lav; Susi, Petri

    2016-01-01

    Research on human enteroviruses has resulted in the identification of more than 100 enterovirus types, which use more than 10 protein receptors and/or attachment factors required in cell binding and initiation of the replication cycle. Many of these “viral” receptors are overexpressed in cancer cells. Receptor binding and the ability to replicate in specific target cells define the tropism and pathogenesis of enterovirus types, because cellular infection often results in cytolytic response, i.e., disruption of the cells. Viral tropism and cytolytic properties thus make native enteroviruses prime candidates for oncolytic virotherapy. Copy DNA cloning and modification of enterovirus genomes have resulted in the generation of enterovirus vectors with properties that are useful in therapy or in vaccine trials where foreign antigenic epitopes are expressed from or on the surface of the vector virus. The small genome size and compact particle structure, however, set limits to enterovirus genome modifications. This review focuses on the therapeutic use of native and recombinant enteroviruses and the methods that have been applied to modify enterovirus genomes for therapy. PMID:26907330

  17. Recombinant protein vaccines produced in insect cells.

    PubMed

    Cox, Manon M J

    2012-02-27

    The baculovirus-insect cell expression system is a well known tool for the production of complex proteins. The technology is also used for commercial manufacture of various veterinary and human vaccines. This review paper provides an overview of how this technology can be applied to produce a multitude of vaccine candidates. The key advantage of this recombinant protein manufacturing platform is that a universal "plug and play" process may be used for producing a broad range of protein-based prophylactic and therapeutic vaccines for both human and veterinary use while offering the potential for low manufacturing costs. Large scale mammalian cell culture facilities previously established for the manufacturing of monoclonal antibodies that have now become obsolete due to yield improvement could be deployed for the manufacturing of these vaccines. Alternatively, manufacturing capacity could be established in geographic regions that do not have any vaccine production capability. Dependent on health care priorities, different vaccines could be manufactured while maintaining the ability to rapidly convert to producing pandemic influenza vaccine when the need arises. PMID:22265860

  18. Homologous recombination deficiency and ovarian cancer.

    PubMed

    Ledermann, Jonathan A; Drew, Yvette; Kristeleit, Rebecca S

    2016-06-01

    The discovery that PARP inhibitors block an essential pathway of DNA repair in cells harbouring a BRCA mutation has opened up a new therapeutic avenue for high-grade ovarian cancers. BRCA1 and BRCA2 proteins are essential for high-fidelity repair of double-strand breaks of DNA through the homologous recombination repair (HRR) pathway. Deficiency in HRR (HRD) is a target for PARP inhibitors. The first PARP inhibitor, olaparib, has now been licensed for BRCA-mutated ovarian cancers. While mutated BRCA genes are individually most commonly associated with HRD other essential HRR proteins may be mutated or functionally deficient potentially widening the therapeutic opportunities for PARP inhibitors. HRD is the first phenotypically defined predictive marker for therapy with PARP inhibitors in ovarian cancer. Several different PARP inhibitors are being trialled in ovarian cancer and this class of drugs has been shown to be a new selective therapy for high-grade ovarian cancer. Around 20% of high-grade serous ovarian cancers harbour germline or somatic BRCA mutations and testing for BRCA mutations should be incorporated into routine clinical practice. The expanded use of PARP inhibitors in HRD deficient (non-BRCA mutant) tumours using a signature of HRD in clinical practice requires validation. PMID:27065456

  19. Recombinant Human Elastase Treatment of Cephalic Veins

    PubMed Central

    Wong, Marco D; Bingham, Karen; Moss, Emma; Warn, J Donald; Smirnov, Igor; Bland, Kimberly S; Starcher, Barry; Franano, F Nicholas; Burke, Steven K

    2016-01-01

    Background Vessel injury at the time of Arteriovenous Fistula (AVF) creation may lead to neointimal hyperplasia that impairs AVF maturation. Vonapanitase, a recombinant human chymotrypsin-like elastase family member 1, is an investigational drug under development to improve AVF maturation and patency. The current studies were designed to document vonapanitase effects in human cephalic veins that are used in AVF creation. Methods Human cephalic veins were mounted on a perfusion myograph. Vonapanitase 1.2, 4, 13.2, and 40 μg/ml or saline was applied drop wise on the vein followed by saline rinse. Vein segments were cut into rings for elastin content determination by desmosine radioimmunoassay and histology. Fluorescently-labelled vonapanitase was applied to veins and adventitial imaging was performed using laser scanning confocal microscopy. In vivo time course experiments were performed by treating rabbit jugular veins and harvesting 1 h and 4 h after vonapanitase treatment. Results / Conclusion Vonapanitase reduced desmosine content in a dose-related manner. Histology also confirmed a dose-related reduction in elastic fiber staining. Fluorescently-labelled vonapanitase persistently localized to elastic fibers in the vein adventitia. In vivo experiments showed a reduction in desmosine content in jugular veins from 1 h to 4 h following treatment. These data suggest that vonapanitase targets elastin in elastic fibers in a dose related manner and that elastase remains in the vessel wall and has catalytic activity for at least 1 h.

  20. Overview of the Purification of Recombinant Proteins

    PubMed Central

    Wingfield, Paul T.

    2015-01-01

    When the first version of this unit was written in 1995 protein purification of recombinant proteins was based on a variety of standard chromatographic methods and approaches many of which were described and mentioned in this unit and elsewhere in the book. In the interim there has been a shift towards an almost universal usage of the affinity or fusion tag. This may not be the case for biotechnology manufacture where affinity tags can complicate producing proteins under regulatory conditions. Regardless of the protein expression system, questions are asked as to which and how many affinity tags to use, where to attach them in the protein and whether to engineer a self cleavage system or simply leave them on. We will briefly address some of these issues. Also although this overview focuses on E.coli, protein expression and purification from the other commonly used expression systems are mentioned and apart from cell breakage methods, the protein purification methods and strategies are essentially the same. PMID:25829302

  1. Dissociative recombination of rotationally cold H+3

    NASA Astrophysics Data System (ADS)

    McCall, B. J.; Huneycutt, A. J.; Saykally, R. J.; Djuric, N.; Dunn, G. H.; Semaniak, J.; Novotny, O.; Al-Khalili, A.; Ehlerding, A.; Hellberg, F.; Kalhori, S.; Neau, A.; Thomas, R. D.; Paal, A.; Österdahl, F.; Larsson, M.

    2004-11-01

    This paper presents the first dissociative recombination (DR) measurement of electrons with rotationally and vibrationally cold H+3 ions. A dc discharge pinhole supersonic jet source was developed and characterized using infrared cavity ringdown spectroscopy before installation on the CRYRING ion storage ring for the DR measurements. Rotational state distributions ( Trot ˜30 K ) produced using the source were comparable to those in the diffuse interstellar medium. Our measurement of the electron energy dependence of the DR cross section showed resonances not clearly seen in experiments using rotationally hot ions, and allowed calculation of the thermal DR rate coefficient for ions at interstellar temperatures, αDR (23 K)=2.6× 10-7 cm3 s-1 . This value is in general agreement with recent theoretical predictions by

    Kokoouline and Greene [Phys. Rev. A 68, 012703 (2003)
    ]. The branching fractions of the two breakup channels, H+H+H and H+ H2 , have also been measured for rotationally and vibrationally cold H+3 .

  2. Dielectronic recombination studies based on EBIT

    SciTech Connect

    Xiao Jun; Han Chuan; Yao Ke; Shen Yang; Yang Yang; Wei Baoren; Fu Yunqing; Lu Di; Hutton, Roger; Zou Yaming

    2013-04-19

    Dielectronic recombination (DR) process plays an important role in high temperature plasmas, where DR can affect charge balance and level populations significantly, and can cause radiative energy loss. Resolvable DR sourced satellite lines are often used for plasma temperature diagnostics, while the un-resolvable ones disturb determining spectral line shape, line intensity, and line position. Data of DR resonant strength is vital for accurate modeling of high temperature plasmas. DR studies are also important for testing atomic structure and atomic collision theories, since they carry information on quantum electrodynamics, relativistic effects, electron correlations and so on. Electron beam ion trap (EBIT) is an accelerator type device, which is capable of acting as both ion sources and light sources. EBIT can produce a special sort of plasma, in which electron energy is tunable and has a very narrow distribution. This made it possible for disentanglement studies on electron ion collision processes in plasmas. In this paper, experimental studies of DR processes based on electron beam ion traps (EBIT) will be discussed.

  3. Dissociative Recombination of Molecular Ions for Astrochemistry

    NASA Astrophysics Data System (ADS)

    Savin, Daniel W.; Novotný, O.; Becker, A.; Buhr, H.; Geppert, W.; Hamberg, M.; Krantz, C.; Kreckel, H.; Schwalm, D.; Spruck, K.; Stützel, J.; Wolf, A.; Yang, B.

    2013-06-01

    Dissociative recombination (DR) of molecular ions is a key chemical process in the cold interstellar medium (ISM). DR affects the composition, charge state, and energy balance of such environments. Astrochemical models of the ISM require reliable total DR cross sections as well as knowledge of the chemical composition and excitation states of the neutral DR products. Theory cannot reliably provide these data. We have systematically measured DR for many astrophysically relevant molecular ions utilizing the TSR storage ring at the Max-Planck-Institute for Nuclear Physics in Heidelberg, Germany. We used the merged ion-electron beam technique combined with an energy- and position-sensitive imaging detector and are able to study DR down to plasma temperatures of 10 K. The DR count rate is used to obtain absolute DR rate coefficient. Additionally we determine the masses of the DR products by measuring their kinetic energy. This allows us to assign particular DR fragmentation channels and to obtain their branching ratios. Moreover, the distribution of detected fragment distances provides information on the kinetic energy released in DR and thus also on the internal excitation of the DR products. All this information is particularly important for understanding DR of heteronuclear polyatomic ions. We will present DR results for several ions recently investigated at TSR. This work is supported in part by NASA and the NSF.

  4. Functional divergence of gene duplicates through ectopic recombination

    PubMed Central

    Christiaens, Joaquin F; Van Mulders, Sebastiaan E; Duitama, Jorge; Brown, Chris A; Ghequire, Maarten G; De Meester, Luc; Michiels, Jan; Wenseleers, Tom; Voordeckers, Karin; Verstrepen, Kevin J

    2012-01-01

    Gene duplication stimulates evolutionary innovation as the resulting paralogs acquire mutations that lead to sub- or neofunctionalization. A comprehensive in silico analysis of paralogs in Saccharomyces cerevisiae reveals that duplicates of cell-surface and subtelomeric genes also undergo ectopic recombination, which leads to new chimaeric alleles. Mimicking such intergenic recombination events in the FLO (flocculation) family of cell-surface genes shows that chimaeric FLO alleles confer different adhesion phenotypes than the parental genes. Our results indicate that intergenic recombination between paralogs can generate a large set of new alleles, thereby providing the raw material for evolutionary adaptation and innovation. PMID:23070367

  5. Resonant phenomena in laser-assisted radiative attachment or recombination

    NASA Astrophysics Data System (ADS)

    Zheltukhin, A. N.; Flegel, A. V.; Frolov, M. V.; Manakov, N. L.; Starace, Anthony F.

    2012-04-01

    Resonant enhancements are predicted in cross sections σn for laser-assisted radiative attachment or electron-ion recombination accompanied by absorption of n laser photons. These enhancements occur for incoming electron energies at which the electron can be attached or recombined by emitting μ laser photons followed by emission of a spontaneous photon upon absorbing n + μ laser photons. The close similarity between rescattering plateaus in spectra of resonant attachment/recombination and of high-order harmonic generation is shown based on a general parametrization for σn and on numerical results for e - H attachment.

  6. General features of the dissociative recombination of polyatomic molecules

    DOE PAGESBeta

    Pratt, S. T.; Jungen, Ch.; Schneider, I. F.; Dulieu, O.; Robert, J.

    2015-01-29

    We discuss some aspects of a simple expression for the low-energy dissociative recombination cross section that applies when the recombination process is dominated by the indirect mechanism. In most previous applications, this expression has been applied to capture into vibrationally excited Rydberg states with the assumption that capture is always followed by prompt dissociation. Here we consider the dissociative recombination of larger polyatomic ions and electrons. More specifically, we consider capture into electronically core-excited Rydberg states, and begin to assess its potential importance for larger systems.

  7. Plasmodium knowlesi Sporozoite Antigen: Expression by Infectious Recombinant Vaccinia Virus

    NASA Astrophysics Data System (ADS)

    Smith, Geoffrey L.; Godson, G. Nigel; Nussenzweig, Victor; Nussenzweig, Ruth S.; Barnwell, John; Moss, Bernard

    1984-04-01

    The gene coding for the circumsporozoite antigen of the malaria parasite Plasmodium knowlesi was inserted into the vaccinia virus genome under the control of a defined vaccinia virus promoter. Cells infected with the recombinant virus synthesized polypeptides of 53,000 to 56,000 daltons that reacted with monoclonal antibody against the repeating epitope of the malaria protein. Furthermore, rabbits vaccinated with the recombinant virus produced antibodies that bound specifically to sporozoites. These data provide evidence for expression of a cloned malaria gene in mammalian cells and illustrate the potential of vaccinia virus recombinants as live malaria vaccines.

  8. Dynamics of carrier recombination in a semiconductor laser structure

    SciTech Connect

    Dzhioev, R. I. Kavokin, K. V.; Kusrayev, Yu. G.; Poletaev, N. K.

    2015-11-15

    Carrier-recombination dynamics is studied by the method of optical orientation at room temperature in the active layer of a laser diode structure. The dependence of the degree of electron-spin orientation on the excitation density is attributed to saturation of the nonradiative-recombination channel. The time of electron capture at recombination centers is determined to be τ{sub e} = 5 × 10{sup –9} s. The temperature of nonequilibrium electrons heated by a He–Ne laser is estimated.

  9. Minimizing recombinations in consensus networks for phylogeographic studies

    PubMed Central

    Parida, Laxmi; Javed, Asif; Melé, Marta; Calafell, Francesc; Bertranpetit, Jaume

    2009-01-01

    Background We address the problem of studying recombinational variations in (human) populations. In this paper, our focus is on one computational aspect of the general task: Given two networks G1 and G2, with both mutation and recombination events, defined on overlapping sets of extant units the objective is to compute a consensus network G3 with minimum number of additional recombinations. We describe a polynomial time algorithm with a guarantee that the number of computed new recombination events is within ϵ = sz(G1, G2) (function sz is a well-behaved function of the sizes and topologies of G1 and G2) of the optimal number of recombinations. To date, this is the best known result for a network consensus problem. Results Although the network consensus problem can be applied to a variety of domains, here we focus on structure of human populations. With our preliminary analysis on a segment of the human Chromosome X data we are able to infer ancient recombinations, population-specific recombinations and more, which also support the widely accepted 'Out of Africa' model. These results have been verified independently using traditional manual procedures. To the best of our knowledge, this is the first recombinations-based characterization of human populations. Conclusion We show that our mathematical model identifies recombination spots in the individual haplotypes; the aggregate of these spots over a set of haplotypes defines a recombinational landscape that has enough signal to detect continental as well as population divide based on a short segment of Chromosome X. In particular, we are able to infer ancient recombinations, population-specific recombinations and more, which also support the widely accepted 'Out of Africa' model. The agreement with mutation-based analysis can be viewed as an indirect validation of our results and the model. Since the model in principle gives us more information embedded in the networks, in our future work, we plan to investigate

  10. Determination of surface recombination velocity in heavily doped silicon

    NASA Technical Reports Server (NTRS)

    Watanabe, M.; Gatos, H. C.; Actor, G.

    1976-01-01

    A method was developed and successfully tested for the determination of the effective surface recombination velocity of silicon layers doped by diffusion of phosphorus to a level of 10 to the 19th to 10 to the 21st per cu cm. The effective recombination velocity was obtained from the dependence of the electron-beam-induced current on the penetration of the electron beam of a scanning electron microscope. A special silicon diode was constructed which permitted the collection at the p-n junction of the carriers excited by the electron beam. This diode also permitted the study of the effects of surface preparation on the effective surface recombination velocity.

  11. Further corrections to the theory of cosmological recombination

    NASA Technical Reports Server (NTRS)

    Krolik, Julian H.

    1990-01-01

    Krolik (1989) pointed out that frequency redistribution due to scattering is more important than cosmological expansion in determining the Ly-alpha frequency profile during cosmological recombination, and that its effects substantially modify the rate of recombination. Although the first statement is true, the second statement is not: a basic symmetry of photon scattering leads to identical cancellations which almost completely erase the effects of both coherent and incoherent scattering. Only a small correction due to atomic recoil alters the line profile from the prediction of pure cosmological expansion, so that the pace of cosmological recombination can be well approximated by ignoring Ly-alpha scattering.

  12. Clinical experience with recombinant molecules for allergy vaccination.

    PubMed

    Cromwell, Oliver; Niederberger, Verena; Horak, Friedrich; Fiebig, Helmut

    2011-01-01

    Numerous allergens have been cloned and produced by the use of recombinant DNA technology. In several cases recombinant variants with reduced IgE-reactivity have also been developed as candidates for allergen specific immunotherapy. Only very few of these proteins have as yet been tested in the clinic, and the major focus has been on birch and grass pollen, two of the most common causes of IgE-mediated allergic disease. This article serves to justify the rational for using recombinant products and reviews the progress that has been made to date with their clinical assessment. PMID:21562972

  13. Characterization and biodistribution of recombinant and recombinant/chimeric constructs of monoclonal antibody B72. 3

    SciTech Connect

    Colcher, D.; Milenic, D.; Roselli, M.; Raubitschek, A.; Yarranton, G.; King, D.; Adair, J.; Whittle, N.; Bodmer, M.; Schlom, J.

    1989-04-01

    Radiolabeled B72.3 has been administered both i.v. and i.p. in patients with colorectal and ovarian cancer as well as other carcinomas and has been shown to selectively bind to approximately 70-80% of metastatic lesions. Greater than 50% of the patients that have been treated with B72.3 have developed an immunological response to murine IgG after a single injection. In an attempt to minimize the immune response of these patients to the administered murine monoclonal antibody, we developed a recombinant form of the murine B72.3 as well as a recombinant/chimeric antibody, using the variable regions of the murine B72.3 and human heavy chain (gamma 4) and light chain (kappa) constant regions. We report here that both the recombinant B72.3 (rB72.3) and the recombinant/chimeric B72.3 (cB72.3(gamma 4)) IgGs maintain the tissue binding and idiotypic specificity of the native murine IgG. The native B72.3, rB72.3, and cB72.3(gamma 4) IgGs were radiolabeled and the biodistribution of these IgGs was studied in athymic mice bearing human colon carcinoma xenografts (LS-174T). Differences were observed between the cB72.3(gamma 4) and the native B72.3 in the percentage of injected dose/g that localized in the tumor. The somewhat lower absolute amounts of the cB72.3(gamma 4) in the tumor are mostly likely due to the observed more rapid clearance from the blood and body of the mouse as compared to the native B72.3 and rB72.3. All three forms (native B72.3, rB72.3, and cB72.3(gamma 4)) of the IgG, however, were able to localize the colon tumor with similar radiolocalization indices (percentage of injected dose/g in tumor divided by the percentage of injected dose/g in normal tissue).

  14. Rate coefficients for N2(+)(v) dissociative recombination

    NASA Astrophysics Data System (ADS)

    Bates, D. R.; Mitchell, J. B. A.

    1991-09-01

    The data of Zipf (1980) on N2(+)(v) dissociative recombination are analyzed taking into account the fact that there is coupling due to reversible symmetrical resonance charge transfer, N2(+)(v) + N2(0) yields N2(+)(0) + N2(v). The vibrational deactivation in N2(+)(v)-Ne collisions is also considered. A reported experimental value of the vibrational deactivation coefficient is found to be much higher than can be reconciled with the results of Zipf and it is therefore rejected. The analysis shows that the recombination coefficient for N2(+)(0) is about 2.6 x 10 exp-7 cu cm/s at 300 K and that recombination coefficients for N2(+)(1) and N2(+)(2) are substantially smaller. It is concluded that these coefficients conflict with the dissociative recombination cross section vs energy curve obtained by the merged beam method.

  15. [Processing and Modification of Recombinant Spider Silk Proteins].

    PubMed

    Liu, Bin; Wang, Tao; Liu, Xiaobing; Luo, Yongen

    2015-08-01

    Due to its special sequence structure, spider silk protein has unique physical and chemical properties, mechanical properties and excellent biological properties. With the expansion of the application value of spider silk in many fields as a functional material, progress has been made in the studies on the expression of recombinant spider silk proteins through many host systems by gene recombinant techniques. Recombinant spider silk proteins can be processed into high performance fibers, and a wide range of nonfibrous morphologies. Moreover, for their excellent biocompatibility and low immune response they are ideal for biomedical applications. Here we review the process and mechanism of preparation in vitro, chemistry and genetic engineering modification on recombinant spider silk protein. PMID:26710473

  16. Experimental study of columnar recombination in fission chambers

    NASA Astrophysics Data System (ADS)

    Filliatre, P.; Lamirand, V.; Geslot, B.; Jammes, C.

    2016-05-01

    In this paper, we present experimental saturation curves of a small gap miniature fission chamber obtained in the MINERVE reactor. The chamber is filled with argon at various pressures, and the fissile material can be coated on the anode, cathode, or both. For analyzing the recombination regime, we consider a model of columnar recombination and discuss its applicability to our chamber. By applying this model to the data, it is possible to estimate the ratio between the recombination coefficient k and an effective column radius b, appearing in the model, to be k / b =(2.5 ± 0.9) ×10-6m2 / s for argon. From these results, a routine measurement of the recombination regime is proposed in order to detect gas leakage. This online diagnosis would be beneficial in terms of lifetime and reliability of the neutron instrumentation of nuclear reactors.

  17. Interchromosomal recombination is suppressed in mammalian somatic cells.

    PubMed Central

    Shulman, M J; Collins, C; Connor, A; Read, L R; Baker, M D

    1995-01-01

    Homologous recombination occurs intrachromosomally as well as interchromosomally, both in mitotic (somatic) cells as well as meiotically in the germline. These different processes can serve very different purposes in maintaining the integrity of the organism and in enhancing diversity in the species. As shown here, comparison of the frequencies of intra- and interchromosomal recombination in meiotic and mitotic cells of both mouse and yeast argues that interchromosomal recombination is particularly low in mitotic cells of metazoan organisms. This result in turn suggests that the recombination machinery of metazoa might be organized to avoid the deleterious effects of homozygotization in somatic cells while still deriving the benefits of species diversification and of DNA repair. Images PMID:7664750

  18. DNA damage tolerance by recombination: Molecular pathways and DNA structures.

    PubMed

    Branzei, Dana; Szakal, Barnabas

    2016-08-01

    Replication perturbations activate DNA damage tolerance (DDT) pathways, which are crucial to promote replication completion and to prevent fork breakage, a leading cause of genome instability. One mode of DDT uses translesion synthesis polymerases, which however can also introduce mutations. The other DDT mode involves recombination-mediated mechanisms, which are generally accurate. DDT occurs prevalently postreplicatively, but in certain situations homologous recombination is needed to restart forks. Fork reversal can function to stabilize stalled forks, but may also promote error-prone outcome when used for fork restart. Recent years have witnessed important advances in our understanding of the mechanisms and DNA structures that mediate recombination-mediated damage-bypass and highlighted principles that regulate DDT pathway choice locally and temporally. In this review we summarize the current knowledge and paradoxes on recombination-mediated DDT pathways and their workings, discuss how the intermediate DNA structures may influence genome integrity, and outline key open questions for future research. PMID:27236213

  19. Auger recombination in sodium-iodide scintillators from first principles

    SciTech Connect

    McAllister, Andrew; Åberg, Daniel; Schleife, André; Kioupakis, Emmanouil

    2015-04-06

    Scintillator radiation detectors suffer from low energy resolution that has been attributed to non-linear light yield response to the energy of the incident gamma rays. Auger recombination is a key non-radiative recombination channel that scales with the third power of the excitation density and may play a role in the non-proportionality problem of scintillators. In this work, we study direct and phonon-assisted Auger recombination in NaI using first-principles calculations. Our results show that phonon-assisted Auger recombination, mediated primarily by short-range phonon scattering, dominates at room temperature. We discuss our findings in light of the much larger values obtained by numerical fits to z-scan experiments.

  20. Nonradiative recombination of excitons in semimagnetic quantum dots

    SciTech Connect

    Chernenko, A. V.

    2015-12-15

    The mechanisms of the nonradiative recombination of excitons in neutral and charged quantum dots based on II–VI semimagnetic semiconductors are investigated. It is shown that, along with the dipole–dipole and direct-exchange mechanisms, there is one more mechanism referred to as the indirect-exchange mechanism and related to sp–d mixing. The selection rules for nonradiative recombination by exchange mechanisms are subsequently derived. The dependence of the efficiency of all recombination mechanisms on the quantum-dot size is studied. The experimentally observed growth in the intracenter photoluminescence intensity with decreasing size of dots and nanocrystals is accounted for. Methods for experimental determination of the contributions of different mechanisms to nonradiative recombination are discussed.

  1. New strategies for genetic engineering Pseudomonas syringae using recombination

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Here we report that DNA oligonucleotides (oligos) introduced directly into bacteria by electroporation can recombine with the bacterial chromosome. This phenomenon was identified in Pseudomonas syringae and we subsequently found that Escherichia coli, Salmonella typhimurium and Shigella flexneri are...

  2. 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

  3. Paracentric inversions do not normally generate monocentric recombinant chromosomes

    SciTech Connect

    Sutherland, G.R.; Callen, D.F.; Gardner, R.J.M.

    1995-11-20

    Dr. Pettenati et al. recently reported a review of paracentric inversions in humans in which they concluded that carriers of these have a 3.8% risk of viable offspring with recombinant chromosomes. We are of the view that there are serious problems with this estimate which should be much closer to zero. The only recombinant chromosomes which can be generated by a paracentric inversion undergoing a normal meiotic division are dicentrics and acentric fragments. Only two such cases were found by Pettenati et al. Several of the alleged monocentric recombinants were originally reported as arising from parental insertions (3-break rearrangements) and it is not legitimate to include them in any analysis of paracentric inversions. Any monocentric recombinant chromosome can only arise from a paracentric inversion by some abnormal process which must involve chromatid breakage and reunion. 4 refs.

  4. Meiotic recombination in normal and cloned bulls and their offspring

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Homologous chromosome pairing and recombination are essential components of meiosis and sexual reproduction. The reshuffling of genetic material through breakage and reunion of chromatids ensure proper segregation of homologous chromosomes in reduction division and genetic diversity in the progeny....

  5. Effect Of Auger Recombination In An Ion Track

    NASA Technical Reports Server (NTRS)

    Edmonds, Larry D.

    1993-01-01

    Report presents theoretical calculations of contribution of Auger recombination to depletion of charge carriers from ionization track left by passage of energetic heavy ion through silicon-based electronic device.

  6. Iron ionization and recombination rates and ionization equilibrium

    NASA Technical Reports Server (NTRS)

    Arnaud, M.; Raymond, J.

    1992-01-01

    In the past few years important progress has been made on the knowledge of ionization and recombination rates of iron, an astrophysically abundant heavy element and a major impurity in laboratory fusion devices. We make a critical review of the existing data on ionization and dielectronic recombination and present new computations of radiative recombination rate coefficients of Fe(+14) through Fe(+25) using the photoionization cross sections of Clark et al. (1986). We provide analytical fits to the recommended data (direct ionization and excitation-autoionization cross sections; radiative and dielectronic recombination rate coefficients). Finally we determine the iron ionic fractions at ionization equilibrium and compare them with previous computations as well as with observational data.

  7. Evidence for recombination in scorpion mitochondrial DNA (Scorpiones: Buthidae)

    PubMed Central

    Gantenbein, Benjamin; Fet, Victor; Gantenbein-Ritter, Iris A; Balloux, François

    2005-01-01

    There has been very little undisputed evidence for recombination in animal mitochondrial DNA (mtDNA) provided so far. Previous unpublished results suggestive of mtDNA recombination in the scorpion family Buthidae, together with cytological evidence for a unique mechanism of mitochondrial fusion in that family, prompted us to investigate this group in more details. First, we sequenced the complete mtDNA genome of Mesobuthus gibbosus, and chose two genes opposing each other (16S and coxI). We then sequenced 150 individuals from the natural populations of four species of Buthidae (Old World genera Buthus and Mesobuthus). We observed strong evidence for widespread recombination through highly significant negative correlations between linkage disequilibrium and physical distance in three out of four species. The evidence is further confirmed when using five other tests for recombination and by the presence of a high amount of homoplasy in phylogenetic trees. PMID:15870032

  8. Products of Dissociative Recombination in the Ionosphere

    NASA Technical Reports Server (NTRS)

    Cosby, Philip

    1996-01-01

    SRI International undertook a novel experimental measurement of the product states formed by dissociative recombination (DR) of O2(+), NO(+), and N2(+) as a function of both electron energy and reactant ion vibrational level. For these measurements we used a recently developed experimental technique for measuring dissociation product distributions that allows both the branching ratios to be accurately determined and the electronic and rovibrational state composition of the reactant ions to be specified. DR is the dominant electron loss mechanism in all regions of the ionosphere. In this process, electron attachment to the molecular ion produces an unstable neutral molecule that rapidly dissociates. For a molecular ion such as O2(+), the dissociation recombination reaction is (1) O2(+) + e yields O + O + W. The atomic products of this reaction, in this case two oxygen atoms, can be produced in a variety of excited states and with a variety of kinetic energies, as represented by W in Eq. (1). These atoms are not only active in the neutral chemistry of the ionosphere, but are also especially important because their optical emissions are often used to infer in situ concentrations of the parent molecular ion and ambient electron densities. Many laboratory measurements have been made of DR reaction rates under a wide range of electron temperatures, but very little is known about the actual distributions among the final states of the atomic products. This lack of knowledge seriously limits the validity and effectiveness of efforts to model both natural and man-made ionospheric disturbances. Bates recently identified major deficiencies in the currently accepted branching ratios for O2(+) as they relate to blue and green line emission measurements in the nocturnal F-region. During our two-year effort, we partially satisfied our ambitious goals. We constructed and operated a variable pressure, electron-impact ion source and a high pressure, hollow-cathode discharge ion

  9. Therapeutic use of recombinant methionyl human leptin.

    PubMed

    Vatier, Camille; Gautier, Jean-François; Vigouroux, Corinne

    2012-10-01

    Recombinant methionyl human leptin (r-metHuLeptin) was first used as a replacement therapy in patients bearing inactivating mutations in the leptin gene. In this indication, it was shown since 1999 to be very efficient in inducing a dramatic weight loss in rare children and adults with severe obesity due to the lack of leptin. These first clinical trials clearly showed that r-metHuLeptin acted centrally to reduce food intake, inducing loss of fat mass, and to correct metabolic alterations, immune and neuroendocrine defects. A few years later, r-metHuLeptin was also shown to reverse the metabolic complications associated with lipodystrophic syndromes, due to primary defects in fat storage, which induce leptin deficiency. The beneficial effects, which could be mediated by central and/or peripheral mechanisms, are thought to mainly involve the lowering effects of leptin on ectopic lipid storage, in particular in liver and muscles, reducing insulin resistance. Interestingly, r-metHuLeptin therapy also reversed the hypothalamic-pituitary-gonadal axis dysfunctions associated with hypothalamic amenorrhea. However, if r-metHuLeptin treatment has been shown to be dramatically efficient in leptin-deficient states, its very limited effect in inducing weight loss in common obese patients revealed that, in patients with adequate leptin secretion, mechanisms of leptin resistance and leptin tolerance prevent r-metHuLeptin from inducing any additional effects. This review will present the current data about the effects of r-metHuLeptin therapy in humans, and discuss the recent perspectives of this therapy in new indications. PMID:22464954

  10. Integrated continuous production of recombinant therapeutic proteins.

    PubMed

    Warikoo, Veena; Godawat, Rahul; Brower, Kevin; Jain, Sujit; Cummings, Daniel; Simons, Elizabeth; Johnson, Timothy; Walther, Jason; Yu, Marcella; Wright, Benjamin; McLarty, Jean; Karey, Kenneth P; Hwang, Chris; Zhou, Weichang; Riske, Frank; Konstantinov, Konstantin

    2012-12-01

    In the current environment of diverse product pipelines, rapidly fluctuating market demands and growing competition from biosimilars, biotechnology companies are increasingly driven to develop innovative solutions for highly flexible and cost-effective manufacturing. To address these challenging demands, integrated continuous processing, comprised of high-density perfusion cell culture and a directly coupled continuous capture step, can be used as a universal biomanufacturing platform. This study reports the first successful demonstration of the integration of a perfusion bioreactor and a four-column periodic counter-current chromatography (PCC) system for the continuous capture of candidate protein therapeutics. Two examples are presented: (1) a monoclonal antibody (model of a stable protein) and (2) a recombinant human enzyme (model of a highly complex, less stable protein). In both cases, high-density perfusion CHO cell cultures were operated at a quasi-steady state of 50-60 × 10(6) cells/mL for more than 60 days, achieving volumetric productivities much higher than current perfusion or fed-batch processes. The directly integrated and automated PCC system ran uninterrupted for 30 days without indications of time-based performance decline. The product quality observed for the continuous capture process was comparable to that for a batch-column operation. Furthermore, the integration of perfusion cell culture and PCC led to a dramatic decrease in the equipment footprint and elimination of several non-value-added unit operations, such as clarification and intermediate hold steps. These findings demonstrate the potential of integrated continuous bioprocessing as a universal platform for the manufacture of various kinds of therapeutic proteins. PMID:22729761

  11. Dissociative Recombination of Molecular Ions for Astrochemistry

    NASA Astrophysics Data System (ADS)

    Novotny, Oldrich; Becker, A.; Buhr, H.; Fleischmann, Andreas; Gamer, Lisa; Geppert, W.; Krantz, C.; Kreckel, H.; Schwalm, D.; Spruck, K.; Wolf, A.; Savin, Daniel Wolf

    2014-06-01

    Dissociative recombination (DR) of molecular ions is a key chemical process in the cold interstellar medium (ISM). DR affects the composition, charge state, and energy balance of such environments. Astrochemical models of the ISM require reliable total DR cross sections as well as knowledge of the chemical composition of the neutral DR products. We have systematically measured DR for many astrophysically relevant molecular ions utilizing the TSR storage ring at the Max-Planck-Institute for Nuclear Physics (MPIK) in Heidelberg, Germany. We used the merged ion-electron beam technique combined with an energy- and position-sensitive imaging detector and are able to study DR down to plasma temperatures as low as 10 K. The DR count rate is used to obtain an absolute merged beams DR rate coefficient from which we can derive a thermal rate coefficient needed for plasma models. Additionally we determine the masses of the DR products by measuring their kinetic energy in the laboratory reference frame. This allows us to assign particular DR fragmentation channels and to obtain their branching ratios. All this information is particularly important for understanding DR of heteronuclear polyatomic ions. We will present DR results for several ions recently investigated at TSR. A new Cryogenic Storage Ring (CSR) is currently being commissioned at MPIK. With the chamber cooled down to ~10 K and a base pressure better than 10-13 mbar, this setup will allow internal cooling of the stored ions down to their rotational ground states, thus opening a new era in DR experiments. New technological challenges arise due to the ultracold, ultra-high vacuum environment of the CSR and thus the detection techniques used at TSR cannot be easily transferred to CSR. We will present new approaches for DR fragment detection in cryogenic environment. This work is supported in part by NASA and the NSF.

  12. The Time Scale of Recombination Rate Evolution in Great Apes.

    PubMed

    Stevison, Laurie S; Woerner, August E; Kidd, Jeffrey M; Kelley, Joanna L; Veeramah, Krishna R; McManus, Kimberly F; Bustamante, Carlos D; Hammer, Michael F; Wall, Jeffrey D

    2016-04-01

    We present three linkage-disequilibrium (LD)-based recombination maps generated using whole-genome sequence data from 10 Nigerian chimpanzees, 13 bonobos, and 15 western gorillas, collected as part of the Great Ape Genome Project (Prado-Martinez J, et al. 2013. Great ape genetic diversity and population history. Nature 499:471-475). We also identified species-specific recombination hotspots in each group using a modified LDhot framework, which greatly improves statistical power to detect hotspots at varying strengths. We show that fewer hotspots are shared among chimpanzee subspecies than within human populations, further narrowing the time scale of complete hotspot turnover. Further, using species-specific PRDM9 sequences to predict potential binding sites (PBS), we show higher predicted PRDM9 binding in recombination hotspots as compared to matched cold spot regions in multiple great ape species, including at least one chimpanzee subspecies. We found that correlations between broad-scale recombination rates decline more rapidly than nucleotide divergence between species. We also compared the skew of recombination rates at centromeres and telomeres between species and show a skew from chromosome means extending as far as 10-15 Mb from chromosome ends. Further, we examined broad-scale recombination rate changes near a translocation in gorillas and found minimal differences as compared to other great ape species perhaps because the coordinates relative to the chromosome ends were unaffected. Finally, on the basis of multiple linear regression analysis, we found that various correlates of recombination rate persist throughout the African great apes including repeats, diversity, and divergence. Our study is the first to analyze within- and between-species genome-wide recombination rate variation in several close relatives. PMID:26671457

  13. Fixation of Emerging Interviral Recombinants in Cucumber Mosaic Virus Populations

    PubMed Central

    Pita, Justin S.

    2013-01-01

    Interstrain recombinants were observed in the progenies of the Cucumber mosaic virus (CMV) reassortant L1L2F3 containing RNAs 1 and 2 from LS-CMV and RNA 3 from Fny-CMV. We characterized these recombinants, and we found that their fixation was controlled by the nature of the replicating RNAs 1 and 2. We demonstrate that the 2b gene partially affects this fixation process, but only in the context of homologous RNAs 1 and 2. PMID:23115282

  14. Strongly Enhanced Recombination via Two-Center Electronic Correlations

    SciTech Connect

    Mueller, C.; Voitkiv, A. B.; Lopez-Urrutia, J. R. Crespo; Harman, Z.

    2010-06-11

    In the presence of a neighboring atom, electron-ion recombination can proceed resonantly via excitation of an electron in the atom, with subsequent relaxation through radiative decay. It is shown that this two-center dielectronic process can largely dominate over single-center radiative recombination at internuclear distances as large as several nanometers. The relevance of the predicted process is demonstrated by using examples of water-dissolved alkali cations and warm dense matter.

  15. Negative-continuum dielectronic recombination for heavy ions

    SciTech Connect

    Artemyev, A.N.; Yerokhin, V.A.; Beier, T.; Kozhuharov, C.; Eichler, J.; Klasnikov, A.E.; Shabaev, V.M.; Stoehlker, T.

    2003-05-01

    The process of recombination of an electron with a bare heavy nucleus via the creation of a free-positron-bound-electron pair is considered. This process is denoted as 'negative-continuum dielectronic recombination' because it results in the capture of an incident electron into a bound state accompanied by a transition of a negative-continuum electron into a bound state. The calculations are performed for a wide range of incident electron energies for Z=82 and 92.

  16. Recombinant Protein Production and Insect Cell Culture and Process

    NASA Technical Reports Server (NTRS)

    Spaulding, Glenn F. (Inventor); Goodwin, Thomas J. (Inventor); OConnor, Kim C. (Inventor); Francis, Karen M. (Inventor); Andrews, Angela D. (Inventor); Prewett, Tracey L. (Inventor)

    1997-01-01

    A process has been developed for recombinant production of selected polypeptides using transformed insect cells cultured in a horizontally rotating culture vessel modulated to create low shear conditions. A metabolically transformed insect cell line is produced using the culture procedure regardless of genetic transformation. The recombinant polypeptide can be produced by an alternative process using virtually infected or stably transformed insect cells containing a gene encoding the described polypeptide. The insect cells can also be a host for viral production.

  17. Recombinant protein production and insect cell culture and process

    NASA Technical Reports Server (NTRS)

    Spaulding, Glenn (Inventor); Prewett, Tacey (Inventor); Goodwin, Thomas (Inventor); Francis, Karen (Inventor); Andrews, Angela (Inventor); Oconnor, Kim (Inventor)

    1993-01-01

    A process has been developed for recombinant production of selected polypeptides using transformed insect cells cultured in a horizontally rotating culture vessel modulated to create low shear conditions. A metabolically transformed insect cell line is produced using the culture procedure regardless of genetic transformation. The recombinant polypeptide can be produced by an alternative process using the cultured insect cells as host for a virus encoding the described polypeptide such as baculovirus. The insect cells can also be a host for viral production.

  18. Recombinant HT{sub m4} gene, protein and assays

    DOEpatents

    Lim, B.; Adra, C.N.; Lelias, J.M.

    1996-09-03

    The invention relates to a recombinant DNA molecule which encodes a HT{sub m4} protein, a transformed host cell which has been stably transfected with a DNA molecule which encodes a HT{sub m4} protein and a recombinant HT{sub m4} protein. The invention also relates to a method for detecting the presence of a hereditary atopy. 2 figs.

  19. Green factory: plants as bioproduction platforms for recombinant proteins.

    PubMed

    Xu, Jianfeng; Dolan, Maureen C; Medrano, Giuliana; Cramer, Carole L; Weathers, Pamela J

    2012-01-01

    Molecular farming, long considered a promising strategy to produce valuable recombinant proteins not only for human and veterinary medicine, but also for agriculture and industry, now has some commercially available products. Various plant-based production platforms including whole-plants, aquatic plants, plant cell suspensions, and plant tissues (hairy roots) have been compared in terms of their advantages and limits. Effective recombinant strategies are summarized along with descriptions of scalable culture systems and examples of commercial progress and success. PMID:21924345

  20. Unified Theoretical Approach to Electron-Ion Recombination

    NASA Astrophysics Data System (ADS)

    Pradhan, Anil

    1998-05-01

    Electron-ion recombination occurs via the electron continuum and embedded resonances that are coupled to each other and thus unified in nature. However, theoretical methods generally separate the two as ``radiative recombination" (RR), referring to recombination through the non-resonant continuum, and ``di-electronic recombination" (DR) through the autoionizing resonances. A computationally unified scheme(S.N. Nahar and A.K. Pradhan, Phys.Rev.Lett.) 68,1488(1992); H.L. Zhang and A.K. Pradhan, Phys.Rev.Lett. 78,195(1997) is described that employs the coupled channel R-matrix method, and detailed balance (the Milne relation), to obtain photo-recombination cross sections including both the continuum and resonant recombination in an ab intio manner. In the energy region corresponding to high-n Rydberg resonances, where the background recombination (RR) is negligibly small, a precise theory of DR (R.H. Bell and M.J. Seaton, J.Phys.B) 18,1589(1985) yields DR collision strengths consistent with their threshold behaviour leading to the cross section for electron impact excitation. For highly charged ions the relativistic fine structure and the effect of radiation damping of resonances are considered. Theoretical cross sections agree well with recent experiments on ion stograge rings and the electron-beam-ion-trap (EBIT). Total electron-ion recombination rates can be obtained for practical applications. For many complex atomic systems, such as the important Iron-peak elements, the new results differ considerably from those hitherto available.

  1. Fine-scale recombination and adaptive radiation could be linked.

    PubMed

    Bodilis, Josselin

    2013-09-15

    The difficult reconstruction of the evolutionary history of the major surface protein gene oprF highlighted an adaptive radiation in the Pseudomonas fluorescens group. The recent work of Hao (2013) showed that partial recombination events in oprF gene occurred specifically in a P. fluorescens lineage under ecological niche segregation. So, I suggest that identification of lineage-specific fine-scale recombination may be a way to detect putative adaptive radiation in bacteria. PMID:23774687

  2. Generation of monoclonal antibodies to recombinant vascular endothelial growth factor.

    PubMed

    Shein, S A; Gurina, O I; Leopol'd, A V; Baklaushev, V P; Korchagina, A A; Grinenko, N F; Ivanova, N V; Volgina, N E; Ryabukhin, I A; Chekhonin, V P

    2012-05-01

    Female BALB/c mice were subcutaneously immunized with recombinant VEGF-164. After 3 immunization cycles, splenic B cells from immunized mouse were fused with immortalized myeloma culture SP2/0-Ag14 cells. Screening of hybrid cells producing anti-VEGF antibodies was performed by ELISA and immunocytochemical analysis on cultured C6 glioma cells. Subsequent cloning yielded hybridoma stably expressing monoclonal anti-VEGF antibodies recognizing recombinant and native VEGF. PMID:22808513

  3. Recombinant TCR ligand reverses clinical signs and CNS damage of EAE induced by recombinant human MOG.

    PubMed

    Sinha, Sushmita; Subramanian, Sandhya; Emerson-Webber, Ashley; Lindner, Maren; Burrows, Gregory G; Grafe, Marjorie; Linington, Christopher; Vandenbark, Arthur A; Bernard, Claude C A; Offner, Halina

    2010-06-01

    Increasing evidence suggests that in addition to T cell-dependent effector mechanisms, autoantibodies are also involved in the pathogenesis of MS, including demyelinating antibodies specific for myelin oligodendrocyte glycoprotein (MOG). Our previous studies have demonstrated that recombinant T cell receptor ligands (RTLs) are very effective for treating T cell-mediated experimental autoimmune encephalomyelitis (EAE). In order to expand the scope of RTL therapy in MS patients, it was of interest to study RTL treatment of EAE involving a demyelinating antibody component. Therefore, we evaluated the therapeutic effects of RTL551, specific for T cells reactive to mouse (m)MOG-35-55 peptide, on EAE induced with recombinant human (rh)MOG in C57BL/6 mice. We report that RTL551 therapy can reverse disease progression and reduce demyelination and axonal damage induced by rhMOG without suppressing the anti-MOG antibody response. This result suggests that T cell-mediated inflammation and associated blood-brain barrier dysfunction are the central contributors to EAE pathogenesis and that successful regulation of these key players restricts potential damage by demyelinating antibodies. The results of our study lend support for the use of RTL therapy for treatment of MS subjects whose disease includes inflammatory T cells as well as those with an additional antibody component. PMID:19789980

  4. Generation of Food-Grade Recombinant Lactic Acid Bacterium Strains by Site-Specific Recombination

    PubMed Central

    Martín, M. Cruz; Alonso, Juan C.; Suárez, Juan E.; Alvarez, Miguel A.

    2000-01-01

    The construction of a delivery and clearing system for the generation of food-grade recombinant lactic acid bacterium strains, based on the use of an integrase (Int) and a resolvo-invertase (β-recombinase) and their respective target sites (attP-attB and six, respectively) is reported. The delivery system contains a heterologous replication origin and antibiotic resistance markers surrounded by two directly oriented six sites, a multiple cloning site where passenger DNA could be inserted (e.g., the cI gene of bacteriophage A2), the int gene, and the attP site of phage A2. The clearing system provides a plasmid-borne gene encoding β-recombinase. The nonreplicative vector-borne delivery system was transformed into Lactobacillus casei ATCC 393 and, by site-specific recombination, integrated as a single copy in an orientation- and Int-dependent manner into the attB site present in the genome of the host strain. The transfer of the clearing system into this strain, with the subsequent expression of the β-recombinase, led to site-specific DNA resolution of the non-food-grade DNA. These methods were validated by the construction of a stable food-grade L. casei ATCC 393-derived strain completely immune to phage A2 infection during milk fermentation. PMID:10831443

  5. Posttranslational modifications and activity of natural and recombinant tissue factor

    PubMed Central

    Butenas, Saulius; Krudysz-Amblo, Jolanta; Mann, Kenneth G

    2010-01-01

    Tissue factor is a membrane protein, which in a complex with factor VIIa initiates in vivo blood coagulation. Due to the scarcity of natural tissue factor protein, most studies have relied upon recombinant tissue factor forms. However, there have been only cursory experimental comparisons of natural and recombinant tissue factor proteins. Our preliminary data suggested that placental tissue factor in a complex with factor VIIa was more efficient activator of factor X than the recombinant protein. After deglycosylation, both forms of tissue factor showed almost an identical activity in the extrinsic factor Xase. Analyses using tryptic digestion and mass-spectrometry revealed that the levels of glycosylation and the composition of carbohydrates present in natural placental tissue factor were different than those in its recombinant counterpart. These data indicate that natural and recombinant tissue factor proteins differ in their posttranslational modifications and that these differences translate into different cofactor activity. Thus the use of recombinant tissue factor proteins for the quantitation of natural tissue factor is misleading. PMID:20138335

  6. Recombination Suppression by Heterozygous Robertsonian Chromosomes in the Mouse

    PubMed Central

    Davisson, M. T.; Akeson, E. C.

    1993-01-01

    Robertsonian chromosomes are metacentric chromosomes formed by the joining of two telocentric chromosomes at their centromere ends. Many Robertsonian chromosomes of the mouse suppress genetic recombination near the centromere when heterozygous. We have analyzed genetic recombination and meiotic pairing in mice heterozygous for Robertsonian chromosomes and genetic markers to determine (1) the reason for this recombination suppression and (2) whether there are any consistent rules to predict which Robertsonian chromosomes will suppress recombination. Meiotic pairing was analyzed using synaptonemal complex preparations. Our data provide evidence that the underlying mechanism of recombination suppression is mechanical interference in meiotic pairing between Robertsonian chromosomes and their telocentric partners. The fact that recombination suppression is specific to individual Robertsonian chromosomes suggests that the pairing delay is caused by minor structural differences between the Robertsonian chromosomes and their telocentric homologs and that these differences arise during Robertsonian formation. Further understanding of this pairing delay is important for mouse mapping studies. In 10 mouse chromosomes (3, 4, 5, 6, 8, 9, 10, 11, 15 and 19) the distances from the centromeres to first markers may still be underestimated because they have been determined using only Robertsonian chromosomes. Our control linkage studies using C-band (heterochromatin) markers for the centromeric region provide improved estimates for the centromere-to-first-locus distance in mouse chromosomes 1, 2 and 16. PMID:8454207

  7. RPA homologs and ssDNA processing during meiotic recombination.

    PubMed

    Ribeiro, Jonathan; Abby, Emilie; Livera, Gabriel; Martini, Emmanuelle

    2016-06-01

    Meiotic homologous recombination is a specialized process that involves homologous chromosome pairing and strand exchange to guarantee proper chromosome segregation and genetic diversity. The formation and repair of DNA double-strand breaks (DSBs) during meiotic recombination differs from those during mitotic recombination in that the homologous chromosome rather than the sister chromatid is the preferred repair template. The processing of single-stranded DNA (ssDNA) formed on intermediate recombination structures is central to driving the specific outcomes of DSB repair during meiosis. Replication protein A (RPA) is the main ssDNA-binding protein complex involved in DNA metabolism. However, the existence of RPA orthologs in plants and the recent discovery of meiosis specific with OB domains (MEIOB), a widely conserved meiosis-specific RPA1 paralog, strongly suggest that multiple RPA complexes evolved and specialized to subdivide their roles during DNA metabolism. Here we review ssDNA formation and maturation during mitotic and meiotic recombination underlying the meiotic specific features. We describe and discuss the existence and properties of MEIOB and multiple RPA subunits in plants and highlight how they can provide meiosis-specific fates to ssDNA processing during homologous recombination. Understanding the functions of these RPA homologs and how they interact with the canonical RPA subunits is of major interest in the fields of meiosis and DNA repair. PMID:26520106

  8. Recombinant medaka (Oryzias melastigmus) pro-hepcidin: Multifunctional characterization.

    PubMed

    Cai, Ling; Cai, Jing-Jing; Liu, Hai-Peng; Fan, Dan-Qing; Peng, Hui; Wang, Ke-Jian

    2012-02-01

    Recently, two hepcidin variant genes (Om-hep1 and Om-hep2) were identified in a model fish marine medaka and both were highly induced in vivo with bacterial challenge, suggesting that the medaka hepcidin may have a similar function to other reported teleostean hepcidins. In the present study, the antibacterial, antiviral and antitumor activities of Om-hep1 were determined using its synthetic and recombinant pro-peptides. The recombinant pro-hepcidin1 was expressed in Escherichia coli and an effective method to produce recombinant Pro-Omhep1 was developed in order to obtain a right folded structure. The results showed that both the synthetic mature peptide and recombinant pro-peptide had similar antibacterial activity against Gram-positive and negative bacteria. In particular, both the synthetic mature Om-hep1 and recombinant Pro-Omhep1 inhibited the viral replication of white spot syndrome virus in the hematopoietic tissue cells of the crayfish Cherax quadricarinatus. Om-hep1 also presented antitumor activity on the cultured human hepatocellular carcinoma cells. In addition, the antimicrobial mechanism of Om-hep1 was measured and it was found that Om-hep1 was likely to be non-membranolytic. The recombinant Pro-Omhep1 performed better biological activity compared to the synthetic mature Om-hep1. This study suggested that Om-hep1 was likely to be an important multifunction protein involved in various resistance actions in the marine medaka immune system. PMID:22051539

  9. Charge relaxation and recombination in photo-excited Mott insulators

    NASA Astrophysics Data System (ADS)

    Prelovšek, P.; Lenarčič, Z.

    2016-04-01

    Recent femtosecond pump-probe experiments on Mott insulators reveal charge recombination, which is in picosecond range, i.e., much faster than in clean bandgap semiconductors although excitation gaps in Mott insulators are even larger. The charge response in photo-excited insulators can be generally divided in femtosecond transient relaxation of charge excitations, which are holons and doublons, and a second slower, but still very fast, holon-doublon (HD) recombination. We present a theory of the recombination rate of the excited HD pairs, based on the two-dimensional (2D) model relevant for cuprates, which shows that such fast processes can be explained even quantitatively with the multi-magnon emission. We show that the condition for the exponential decay as observed in the experiment is the existence of the exciton, i.e., the bound HD pair. Its recombination rate is exponentially dependent on the charge gap and on the magnon energy, while the ultrafast process can be traced back to strong charge-spin coupling. We comment also fast recombination times in the one-dimensional (1D) Mott insulators, as e.g., organic salts. The recombination rate in the latter cases can be explained with the stronger coupling with phonon excitations.

  10. Genetic diversity and recombination analysis of sweepoviruses from Brazil

    PubMed Central

    2012-01-01

    Background Monopartite begomoviruses (genus Begomovirus, family Geminiviridae) that infect sweet potato (Ipomoea batatas) around the world are known as sweepoviruses. Because sweet potato plants are vegetatively propagated, the accumulation of viruses can become a major constraint for root production. Mixed infections of sweepovirus species and strains can lead to recombination, which may contribute to the generation of new recombinant sweepoviruses. Results This study reports the full genome sequence of 34 sweepoviruses sampled from a sweet potato germplasm bank and commercial fields in Brazil. These sequences were compared with others from public nucleotide sequence databases to provide a comprehensive overview of the genetic diversity and patterns of genetic exchange in sweepoviruses isolated from Brazil, as well as to review the classification and nomenclature of sweepoviruses in accordance with the current guidelines proposed by the Geminiviridae Study Group of the International Committee on Taxonomy of Viruses (ICTV). Co-infections and extensive recombination events were identified in Brazilian sweepoviruses. Analysis of the recombination breakpoints detected within the sweepovirus dataset revealed that most recombination events occurred in the intergenic region (IR) and in the middle of the C1 open reading frame (ORF). Conclusions The genetic diversity of sweepoviruses was considerably greater than previously described in Brazil. Moreover, recombination analysis revealed that a genomic exchange is responsible for the emergence of sweepovirus species and strains and provided valuable new information for understanding the diversity and evolution of sweepoviruses. PMID:23082767

  11. Recombination, chromosome number and eusociality in the Hymenoptera

    PubMed Central

    Ross, L; Blackmon, H; Lorite, P; Gokhman, V E; Hardy, N B

    2015-01-01

    Extraordinarily high rates of recombination have been observed in some eusocial species. The most popular explanation is that increased recombination increases genetic variation among workers, which in turn increases colony performance, for example by increasing parasite resistance. However, support for the generality of higher recombination rates among eusocial organisms remains weak, due to low sample size and a lack of phylogenetic independence of observations. Recombination rate, although difficult to measure directly, is correlated with chromosome number. As predicted, several authors have noted that chromosome numbers are higher among the eusocial species of Hymenoptera (ants, bees and wasps). Here, we present a formal comparative analysis of karyotype data from 1567 species of Hymenoptera. Contrary to earlier studies, we find no evidence for an absolute difference between chromosome number in eusocial and solitary species of Hymenoptera. However, we find support for an increased rate of chromosome number change in eusocial taxa. We show that among eusocial taxa colony size is able to explain some of the variation in chromosome number: intermediate-sized colonies have more chromosomes than those that are either very small or very large. However, we were unable to detect effects of a number of other colony characteristics predicted to affect recombination rate – including colony relatedness and caste number. Taken together, our results support the view that a eusocial lifestyle has led to variable selection pressure for increased recombination rates, but that identifying the factors contributing to this variable selection will require further theoretical and empirical effort. PMID:25382409

  12. Meiotic recombination analysis in female ducks (Anas platyrhynchos).

    PubMed

    Pigozzi, M I; Del Priore, L

    2016-06-01

    Meiotic recombination in female ducks was directly studied by immunolocalization of MLH1 protein, a mismatch repair protein of mature recombination nodules. In total, 6820 crossovers were scored along the autosomal synaptonemal complexes in 122 meiotic nuclei. From this analysis we predict that the female map length of the duck is 2845 cM, with a genome wide recombination rate of 2 cM/Mb. MLH1-focus mapping along the six largest bivalents shows regional variations of recombination frequencies that can be linked to differences in chromosome morphology. From this MLH1 mapping it can be inferred that distally located markers will appear more separated in genetic maps than physically equidistant markers located near the centromeres on bivalents 1 and 2. Instead, markers at interstitial positions on the acrocentric bivalents 3-6 will appear more tightly linked than expected on the basis of their physical distance because recombination is comparatively lower at the mid region of these chromosomes. The present results provide useful information to complement linkage mapping in ducks and extend previous knowledge about the variation of recombination rates among domestic Galloanserae. PMID:27115519

  13. Immunoglobulin/Myc recombinations in murine Peyer's patch follicles.

    PubMed

    Müller, J R; Mushinski, E B; Williams, J A; Hausner, P F

    1997-09-01

    Immunoglobulin heavy chain (Igh)/Myc recombinations are a hallmark of pristane-induced mouse plasmacytomas but are also frequently found in non-tumorous tissues. Here we describe for the first time a PCR-based technique for detecting fusions between Igh mu or Igh alpha and Myc in situ. Igh/Myc recombinations were found in transplanted and primary plasmacytomas. In addition, the gut-associated lymphoid tissues of plasmacytoma-free BALB/c mice were investigated for the presence of Igh/Myc fusions. Igh/Myc rearrangements were detected in Peyer's patch follicles and in the intestinal lamina propria both in normal mice and in mice shortly after pristane treatment. The sequence analysis showed that i) three to five different Igh/Myc hybrid sequences were present in individual follicles, ii) Igh/Myc recombinations can be subjected to additional switch recombinations as shown by related sequences in neighboring cells, and iii) cells harboring these rearrangements migrate into the adjacent lamina propria. The results indicate that Peyer's patches are a hyper-recombinogenic tissue. Myc recombination-positive cells are present in at least 100-fold more frequently than expected if recombinations were random, which suggests that this kind of trans-chromosomal rearrangement may be targeted. PMID:9290947

  14. Recombination Coefficient Measurements of O and N radicals

    NASA Astrophysics Data System (ADS)

    Singh, Harmeet; Coburn, John; Graves, David

    1999-10-01

    Surface recombination of radicals in low-pressure high-density plasmas has direct influence on the neutral and ionic composition of the plasma. While, electron impact dissociation of molecules is the dominant mechanism for creation of radicals, the surface recombination of radicals is often expected to be the dominant loss mechanism. We have a combination of measurements and a model to determine the recombination coefficients of O and N, to O2 and N2, respectively on the stainless steel walls of our inductively coupled plasma chamber. The radial variation of the electron energy distribution function (EEDF) is measured using a tuned, cylindrical Langmuir probe. The number density of the molecular species is measured using line-of-sight modulated beam mass spectrometry. The mass spectrometer is differentially pumped in three stages to ensure a good beam to background signal ratio. The radical absolute number density is measured using appearance potential mass spectrometry with the aforementioned mass spectrometer. The recombination coefficient is calculated using a balance of the volume-generation and surface-loss rates of the radicals in the plasma. The generation rate of the radicals is calculated using the number density measurements of the parent molecule and the spatially resolved EEDFs. At approximately 330 K on stainless steel, the recombination coefficient for O is 0.16, and recombination coefficient for N is 0.07.

  15. Seroreactive recombinant herpes simplex virus type 2-specific glycoprotein G.

    PubMed Central

    Parkes, D L; Smith, C M; Rose, J M; Brandis, J; Coates, S R

    1991-01-01

    The herpes simplex virus type 2 (HSV-2) genome codes for an envelope protein, glycoprotein G (gG), which contains predominantly type 2-specific epitopes. A portion of this gG gene has been expressed as a fusion protein in Escherichia coli. Expression was regulated by a lambda phage pL promoter. The 60,000-molecular-weight recombinant protein was purified by ion-exchange chromatography. Amino acid sequence analysis confirmed the N terminus of the purified protein. Mice immunized with recombinant gG developed antibodies reactive with native HSV-2 protein, but not with HSV-1 protein, in an indirect immunofluorescence assay. The serological activity of this purified recombinant gG protein was evaluated by immunoblot assay. This protein was reactive with an HSV-2 gG monoclonal antibody. It was also reactive with HSV-2 rabbit antiserum but not with HSV-1 rabbit antiserum. Of 15 patient serum samples known to have antibody to HSV-2, 14 were reactive with this recombinant type 2-specific gG protein, and none of 15 HSV antibody-negative patient serum samples showed reactivity. In agreement with the expected prevalence of HSV-2 infection, 27.6% of 134 serum samples from random normal individuals had antibodies reactive with recombinant gG. This recombinant gG protein may be of value in detecting HSV-2-specific antibody responses in patients infected with HSV-2. Images PMID:1653787

  16. Electron-ion recombination of FetII

    NASA Astrophysics Data System (ADS)

    Nahar, Sultana N.

    1997-03-01

    A complete treatment of electron-ion recombination of e+Fe III-->Fe II employing a unified method is presented. The treatment incorporates both the radiative and dielectronic recombinations in a self-consistent manner. Total recombination rate coefficients are obtained from photoionization cross sections, and from collision strengths for dielectronic recombination calculated using the precise theory of Bell and Seaton [J. Phys. B 18, 1589 (1985)]. Large-scale computations for both of these quantities are carried out in the close coupling approximation using the R-matrix method with an eigenfunction expansion that includes 83 states of Fe III dominated by the ground 3d6, and the excited 3d54s and 3d54p configurations. Both the total and state-specific recombination rate coefficients are obtained. Comparison of the present results with the previous ones shows considerable difference for most of the temperature regions. The present results provide accurate and self-consistent recombination rates, in the temperature range of practical applications (T<105 K), for ionization balance in photoionization models employing the detailed photoionization cross sections from the Opacity Project.

  17. Electron-ion recombination of FeII

    SciTech Connect

    Nahar, S.N.

    1997-03-01

    A complete treatment of electron-ion recombination of e+FeIII{r_arrow}FeII employing a unified method is presented. The treatment incorporates both the radiative and dielectronic recombinations in a self-consistent manner. Total recombination rate coefficients are obtained from photoionization cross sections, and from collision strengths for dielectronic recombination calculated using the precise theory of Bell and Seaton [J. Phys. B {bold 18}, 1589 (1985)]. Large-scale computations for both of these quantities are carried out in the close coupling approximation using the R-matrix method with an eigenfunction expansion that includes 83 states of FeIII dominated by the ground 3d{sup 6}, and the excited 3d{sup 5}4s and 3d{sup 5}4p configurations. Both the total and state-specific recombination rate coefficients are obtained. Comparison of the present results with the previous ones shows considerable difference for most of the temperature regions. The present results provide accurate and self-consistent recombination rates, in the temperature range of practical applications (T{lt}10{sup 5} K), for ionization balance in photoionization models employing the detailed photoionization cross sections from the Opacity Project. {copyright} {ital 1997} {ital The American Physical Society}

  18. Distribution of Recombination Crossovers and the Origin of Haplotype Blocks: The Interplay of Population History, Recombination, and Mutation

    PubMed Central

    Wang, Ning; Akey, Joshua M.; Zhang, Kun; Chakraborty, Ranajit; Jin, Li

    2002-01-01

    Recent studies suggest that haplotypes are arranged into discrete blocklike structures throughout the human genome. Here, we present an alternative haplotype block definition that assumes no recombination within each block but allows for recombination between blocks, and we use it to study the combined effects of demographic history and various population genetic parameters on haplotype block characteristics. Through extensive coalescent simulations and analysis of published haplotype data on chromosome 21, we find that (1) the combined effects of population demographic history, recombination, and mutation dictate haplotype block characteristics and (2) haplotype blocks can arise in the absence of recombination hot spots. Finally, we provide practical guidelines for designing and interpreting studies investigating haplotype block structure. PMID:12384857

  19. “Genome-wide recombination and chromosome segregation in human oocytes and embryos reveal selection for maternal recombination rates”

    PubMed Central

    Natesan, Senthilkumar A.; Joshi, Hrishikesh A.; Cimadomo, Danilo; Griffin, Darren K.; Sage, Karen; Summers, Michael C.; Thornhill, Alan R.; Housworth, Elizabeth; Herbert, Alex D.; Rienzi, Laura; Ubaldi, Filippo M.; Handyside, Alan H.; Hoffmann, Eva R.

    2015-01-01

    Crossover recombination reshuffles genes and prevents errors in segregation that lead to extra or missing chromosomes (aneuploidy) in human eggs, a major cause of pregnancy failure and congenital disorders. Here, we generate genome-wide maps of crossovers and chromosome segregation patterns by recovering all three products of single female meioses. Genotyping > 4 million informative single-nucleotide polymorphisms (SNPs) from 23 complete meioses allowed us to map 2,032 maternal and 1,342 paternal crossovers and to infer the segregation patterns of 529 chromosome pairs. We uncover a novel reverse chromosome segregation pattern in which both homologs separate their sister chromatids at meiosis I; detect selection for higher recombination rates in the female germline by the elimination of aneuploid embryos; and report chromosomal drive against non-recombinant chromatids at meiosis II. Collectively, our findings reveal that recombination not only affects homolog segregation at meiosis I but also the fate of sister chromatids at meiosis II. PMID:25985139

  20. Genome-wide maps of recombination and chromosome segregation in human oocytes and embryos show selection for maternal recombination rates.

    PubMed

    Ottolini, Christian S; Newnham, Louise J; Capalbo, Antonio; Natesan, Senthilkumar A; Joshi, Hrishikesh A; Cimadomo, Danilo; Griffin, Darren K; Sage, Karen; Summers, Michael C; Thornhill, Alan R; Housworth, Elizabeth; Herbert, Alex D; Rienzi, Laura; Ubaldi, Filippo M; Handyside, Alan H; Hoffmann, Eva R

    2015-07-01

    Crossover recombination reshuffles genes and prevents errors in segregation that lead to extra or missing chromosomes (aneuploidy) in human eggs, a major cause of pregnancy failure and congenital disorders. Here we generate genome-wide maps of crossovers and chromosome segregation patterns by recovering all three products of single female meioses. Genotyping >4 million informative SNPs from 23 complete meioses allowed us to map 2,032 maternal and 1,342 paternal crossovers and to infer the segregation patterns of 529 chromosome pairs. We uncover a new reverse chromosome segregation pattern in which both homologs separate their sister chromatids at meiosis I; detect selection for higher recombination rates in the female germ line by the elimination of aneuploid embryos; and report chromosomal drive against non-recombinant chromatids at meiosis II. Collectively, our findings show that recombination not only affects homolog segregation at meiosis I but also the fate of sister chromatids at meiosis II. PMID:25985139

  1. Urinary hMG (Meropur) versus recombinant FSH plus recombinant LH (Pergoveris) in IVF: a multicenter, prospective, randomized controlled trial.

    PubMed

    Pacchiarotti, Alessandro; Sbracia, Marco; Frega, Antonio; Selman, Helmy; Rinaldi, Leonardo; Pacchiarotti, Arianna

    2010-11-01

    To compare IVF outcome in ovarian stimulation protocols with recombinant FSH plus recombinant LH versus hMG, 122 patients were randomized into two study groups: group A, patients treated with urinary hMG, and group B, patients treated with rFSH plus rLH. The two groups proved to be comparable to the main IVF outcome (pregnancy rate, implantation rate, oocytes, and embryos quality), with an increasing risk of ovarian hyperstimulation in the Pergoveris group. PMID:20537626

  2. Differential Requirements of Singleplex and Multiplex Recombineering of Large DNA Constructs

    PubMed Central

    Reddy, Thimma R.; Kelsall, Emma J.; Fevat, Léna M. S.; Munson, Sarah E.; Cowley, Shaun M.

    2015-01-01

    Recombineering is an in vivo genetic engineering technique involving homologous recombination mediated by phage recombination proteins. The use of recombineering methodology is not limited by size and sequence constraints and therefore has enabled the streamlined construction of bacterial strains and multi-component plasmids. Recombineering applications commonly utilize singleplex strategies and the parameters are extensively tested. However, singleplex recombineering is not suitable for the modification of several loci in genome recoding and strain engineering exercises, which requires a multiplex recombineering design. Defining the main parameters affecting multiplex efficiency especially the insertion of multiple large genes is necessary to enable efficient large-scale modification of the genome. Here, we have tested different recombineering operational parameters of the lambda phage Red recombination system and compared singleplex and multiplex recombineering of large gene sized DNA cassettes. We have found that optimal multiplex recombination required long homology lengths in excess of 120 bp. However, efficient multiplexing was possible with only 60 bp of homology. Multiplex recombination was more limited by lower amounts of DNA than singleplex recombineering and was greatly enhanced by use of phosphorothioate protection of DNA. Exploring the mechanism of multiplexing revealed that efficient recombination required co-selection of an antibiotic marker and the presence of all three Red proteins. Building on these results, we substantially increased multiplex efficiency using an ExoVII deletion strain. Our findings elucidate key differences between singleplex and multiplex recombineering and provide important clues for further improving multiplex recombination efficiency. PMID:25954970

  3. Recombinant thrombomodulin for secondary thrombotic thrombocytopenic purpura.

    PubMed

    Nakamura, Kensuke; Inokuchi, Ryota; Hiruma, Takahiro; Ohshima, Kazuma; Sonoo, Tomohiro; Tokunaga, Kurato; Doi, Kent; Nakajima, Susumu

    2016-06-01

    In the pathogenesis of thrombotic thrombocytopenic purpura (TTP), reductions in the enzyme activity of ADAMTS13, which cuts ultralarge von Willebrand multimers, generates shear stress on the microvascular endothelium, leading to platelet aggregation and the formation of a thrombus. ADAMTS13 activity is markedly decreased in typical TTP, but is only mildly reduced in secondary TTP, which concomitantly develops with primary disease. The latter develops with septic disseminated intravascular coagulation (DIC) and often causes organ failure. Recombinant thrombomodulin (rTM) is a drug that is used to treat DIC and may also remit TTP because it improves vascular endothelial dysfunction. Therefore, we herein investigated the efficacy of rTM in patients treated for the pathology of secondary TTP. Patients who were admitted to the Emergency and Critical Care Center of our hospital and met the following conditions were extracted and retrospectively analyzed: hemolytic anemia accompanied by fragmented red blood cells (Hb < 12 g/dL or lower); thrombocytopenia (<100 × 10/μL); and ADAMTS13 activity <50%. Sixteen patients were included and accompanied by Kidney Disease: Improving Global Outcomes (KDIGO) stage 2 or more severe nephropathy and DIC. Eleven and 5 patients treated with and without rTM (the rTM and non-rTM treatment groups, respectively) were compared, and no significant difference was noted in their basic characteristics, such as background disease and severity. No significant difference was observed in survival rates; however, the platelet count, which is an important outcome of treatments for TTP, significantly increased in the rTM treatment group: 3.3 ± 2.6→11.3 ± 14.6 versus 3.5 ± 3.7→5.7 ± 3.9 (×1000/μL) (P = 0.034). Thrombotic thrombocytopenic purpura originally requires invasive treatments and its prognosis is not favorable. Blood thrombomodulin levels also markedly increase due to vascular endothelial dysfunction

  4. Transforming the treatment for hemophilia B patients: update on the clinical development of recombinant fusion protein linking recombinant coagulation factor IX with recombinant albumin (rIX-FP).

    PubMed

    Santagostino, Elena

    2016-05-01

    Recombinant fusion protein linking recombinant coagulation factor IX with recombinant albumin (rIX-FP; Idelvion®(†)) is an innovative new treatment designed to extend the half-life of factor IX (FIX) and ease the burden of care for hemophilia B patients. The rIX-FP clinical development program - PROLONG-9FP - is in its advanced phases, with pivotal studies in previously treated adults, adolescents, and pediatrics now completed. Across all age groups studied, rIX-FP has demonstrated a markedly improved pharmacokinetic profile compared with plasma-derived and recombinant FIX treatments, with a 30-40% higher incremental recovery, an approximately 5-fold longer half-life, a lower clearance, and a greater area under the curve. rIX-FP has been very well tolerated with an excellent safety profile. In the pivotal studies, there have been no reports of FIX inhibitors or antidrug antibodies, and few treatment-related adverse events have been observed. Prophylactic regimens of rIX-FP administered once weekly to once every 14 days have been highly effective. When used for surgical prophylaxis, a single infusion of rIX-FP has been sufficient to maintain hemostasis, even during major orthopedic surgery. An ongoing study is now enrolling previously untreated patients and evaluating the possibility of extending the dosing interval to every 21 days. There is little doubt that rIX-FP will transform the treatment of hemophilia B. PMID:27288064

  5. Enhanced Proteolytic Processing of Recombinant Human Coagulation Factor VIII B-Domain Variants by Recombinant Furins.

    PubMed

    Demasi, Marcos A; de S Molina, Erika; Bowman-Colin, Christian; Lojudice, Fernando H; Muras, Angelita; Sogayar, Mari C

    2016-06-01

    Recombinant human factor VIII (rFVIII) is used in replacement therapy for hemophilia A. Current research efforts are focused on bioengineering rFVIII molecules to improve its secretion efficiency and stability, limiting factors for its efficient production. However, high expression yield in mammalian cells of these rFVIII variants is generally associated with limited proteolytic processing. Non-processed single-chain polypeptides constitute non-natural FVIII molecule configurations with unpredictable toxicity and/or antigenicity. Our main objective was to demonstrate the feasibility of promoting full-proteolytic processing of an rFVIII variant retaining a portion of the B-domain, converting it into the smallest natural activatable form of rFVIII, while keeping its main advantage, i.e., improved secretion efficiency. We generated and employed a CHO-DG44 cell clone producing an rFVIII variant retaining a portion of the B-domain and the FVIII native cleavage site between Arg(1648) and Glu(1649). By bioengineering CHO-DG44 cells to express stably the recombinant human endoproteases PACE, PACE-SOL, PCSK5, PCSK6, or PCKS7, we were able to achieve complete intra- or extracellular proteolytic processing of this rFVIII variant. Additionally, our quantitative data indicated that removal of the B-domain segment by intracellular proteolytic processing does not interfere with this rFVIII variant secretion efficiency. This work also provides the first direct evidence of (1) intracellular cleavage at the Arg(1648) FVIII processing site promoted by wild-type PACE and PCSK7 and (2) proteolytic processing at the Arg(1648) FVIII processing site by PCSK6. PMID:27126696

  6. Genome-wide recombination dynamics are associated with phenotypic variation in maize.

    PubMed

    Pan, Qingchun; Li, Lin; Yang, Xiaohong; Tong, Hao; Xu, Shutu; Li, Zhigang; Li, Weiya; Muehlbauer, Gary J; Li, Jiansheng; Yan, Jianbing

    2016-05-01

    Meiotic recombination is a major driver of genetic diversity, species evolution, and agricultural improvement. Thus, an understanding of the genetic recombination landscape across the maize (Zea mays) genome will provide insight and tools for further study of maize evolution and improvement. Here, we used c. 50 000 single nucleotide polymorphisms to precisely map recombination events in 12 artificial maize segregating populations. We observed substantial variation in the recombination frequency and distribution along the ten maize chromosomes among the 12 populations and identified 143 recombination hot regions. Recombination breakpoints were partitioned into intragenic and intergenic events. Interestingly, an increase in the number of genes containing recombination events was accompanied by a decrease in the number of recombination events per gene. This kept the overall number of intragenic recombination events nearly invariable in a given population, suggesting that the recombination variation observed among populations was largely attributed to intergenic recombination. However, significant associations between intragenic recombination events and variation in gene expression and agronomic traits were observed, suggesting potential roles for intragenic recombination in plant phenotypic diversity. Our results provide a comprehensive view of the maize recombination landscape, and show an association between recombination, gene expression and phenotypic variation, which may enhance crop genetic improvement. PMID:26720856

  7. CRISPR-Cas9-assisted recombineering in Lactobacillus reuteri.

    PubMed

    Oh, Jee-Hwan; van Pijkeren, Jan-Peter

    2014-01-01

    Clustered regularly interspaced palindromic repeats (CRISPRs) and the CRISPR-associated (Cas) nuclease protect bacteria and archeae from foreign DNA by site-specific cleavage of incoming DNA. Type-II CRISPR-Cas systems, such as the Streptococcus pyogenes CRISPR-Cas9 system, can be adapted such that Cas9 can be guided to a user-defined site in the chromosome to introduce double-stranded breaks. Here we have developed and optimized CRISPR-Cas9 function in the lactic acid bacterium Lactobacillus reuteri ATCC PTA 6475. We established proof-of-concept showing that CRISPR-Cas9 selection combined with single-stranded DNA (ssDNA) recombineering is a realistic approach to identify at high efficiencies edited cells in a lactic acid bacterium. We show for three independent targets that subtle changes in the bacterial genome can be recovered at efficiencies ranging from 90 to 100%. By combining CRISPR-Cas9 and recombineering, we successfully applied codon saturation mutagenesis in the L. reuteri chromosome. Also, CRISPR-Cas9 selection is critical to identify low-efficiency events such as oligonucleotide-mediated chromosome deletions. This also means that CRISPR-Cas9 selection will allow identification of recombinant cells in bacteria with low recombineering efficiencies, eliminating the need for ssDNA recombineering optimization procedures. We envision that CRISPR-Cas genome editing has the potential to change the landscape of genome editing in lactic acid bacteria, and other Gram-positive bacteria. PMID:25074379

  8. Synonymous and nonsynonymous distances help untangle convergent evolution and recombination.

    PubMed

    Chi, Peter B; Chattopadhyay, Sujay; Lemey, Philippe; Sokurenko, Evgeni V; Minin, Vladimir N

    2015-08-01

    When estimating a phylogeny from a multiple sequence alignment, researchers often assume the absence of recombination. However, if recombination is present, then tree estimation and all downstream analyses will be impacted, because different segments of the sequence alignment support different phylogenies. Similarly, convergent selective pressures at the molecular level can also lead to phylogenetic tree incongruence across the sequence alignment. Current methods for detection of phylogenetic incongruence are not equipped to distinguish between these two different mechanisms and assume that the incongruence is a result of recombination or other horizontal transfer of genetic information. We propose a new recombination detection method that can make this distinction, based on synonymous codon substitution distances. Although some power is lost by discarding the information contained in the nonsynonymous substitutions, our new method has lower false positive probabilities than the comparable recombination detection method when the phylogenetic incongruence signal is due to convergent evolution. We apply our method to three empirical examples, where we analyze: (1) sequences from a transmission network of the human immunodeficiency virus, (2) tlpB gene sequences from a geographically diverse set of 38 Helicobacter pylori strains, and (3) hepatitis C virus sequences sampled longitudinally from one patient. PMID:26061623

  9. Electron-ion Recombination and Photoionization of P II

    NASA Astrophysics Data System (ADS)

    Nahar, Sultana

    2016-05-01

    Study of the inverse processes of photoionization and electron-ion recombination of P II will be reported. It is a highly reactive ion and has been difficult to detect without detailed information of its interactions. Although a low charged ion, present study shows features in photoionization resulting from relativistic fine structure couplings at low energy region near the ionization threshold of many levels. Unified method under the framework of close coupling approximation and R-matrix method and an extension of Bell and Seaton theory has been used to study the inverse processes. The method gives the level-specific as well as the total recombination rate coefficients which include both the radiative recombination (RR) and dielectronic recombination (DR) in a precise manner. The present results include level specific rates and photoionization cross sections of 475 fine structure levels with n <= 10. Preliminary results on the total recombination rates show considerable interference of RR and DR around 4000 K and a DR peak around 105 K. NSF,DOE,OSC.

  10. Suppression of auger recombination in ""giant"" core/shell nanocrystals

    SciTech Connect

    Garcia Santamaria, Florencio; Vela, Javier; Schaller, Richard D; Hollingsworth, Jennifer A; Klimov, Victor I; Chen, Yongfen

    2009-01-01

    Many potential applications of semiconductor nanocrystals are hindered by nonradiative Auger recombination wherein the electron-hole (exciton) recombination energy is transferred to a third charge carrier. This process severely limits the lifetime and bandwidth of optical gain, leads to large nonradiative losses in light emitting diodes and photovoltaic cells, and is believed to be responsible for intermittency ('blinking') of emission from single nanocrystals. The development of nanostructures in which Auger recombination is suppressed has been a longstanding goal in colloidal nanocrystal research. Here, we demonstrate that such suppression is possible using so-called 'giant' nanocrystals that consist of a small CdSe core and a thick CdS shell. These nanostructures exhibit a very long biexciton lifetime ({approx}10 ns) that is likely dominated by radiative decay instead of non-radiative Auger recombination. As a result of suppressed Auger recombination, even high-order multiexcitons exhibit high emission efficiencies, which allows us to demonstrate optical amplification with an extraordinarily large bandwidth (>500 me V) and record low excitation thresholds.

  11. Epistasis and the selective advantage of sex and recombination

    NASA Astrophysics Data System (ADS)

    de Oliveira, Viviane M.; da Silva, Juliana K.; Campos, Paulo R. A.

    2008-09-01

    The understanding of the central mechanisms favoring sex and recombination in real populations is one of the fundamental issues in evolutionary biology. Based on a previous stochastic formulation for the study of sex, here we aim to investigate the conditions under which epistasis favors the fixation of the sexual mode of reproduction in a given population. In addition, we try to identify the evolutionary forces which contribute to this process. One considers a finite population model which assumes the existence of a recombination modifier allele that can activate the recombination mechanism. We have found that sex is very little favored in a scenario of antagonistic epistasis, and this advantage only occurs in a narrow range of values of the selection coefficient sd . On the other hand, synergistic epistasis favors recombination in a very broad domain. However, the major mechanism contributing to the spreading of the modifier allele depends on the range of values of sd . At large sd , background selection favors recombination since it increases the efficacy of selection, while at low sd Muller’s ratchet is the leading mechanism.

  12. Diversity, Mutation and Recombination Analysis of Cotton Leaf Curl Geminiviruses

    PubMed Central

    Saleem, Huma; Nahid, Nazia; Shakir, Sara; Ijaz, Sehrish; Murtaza, Ghulam; Khan, Asif Ali; Mubin, Muhammad; Nawaz-ul-Rehman, Muhammad Shah

    2016-01-01

    The spread of cotton leaf curl disease in China, India and Pakistan is a recent phenomenon. Analysis of available sequence data determined that there is a substantial diversity of cotton-infecting geminiviruses in Pakistan. Phylogenetic analyses indicated that recombination between two major groups of viruses, cotton leaf curl Multan virus (CLCuMuV) and cotton leaf curl Kokhran virus (CLCuKoV), led to the emergence of several new viruses. Recombination detection programs and phylogenetic analyses showed that CLCuMuV and CLCuKoV are highly recombinant viruses. Indeed, CLCuKoV appeared to be a major donor virus for the coat protein (CP) gene, while CLCuMuV donated the Rep gene in the majority of recombination events. Using recombination free nucleotide datasets the substitution rates for CP and Rep genes were determined. We inferred similar nucleotide substitution rates for the CLCuMuV-Rep gene (4.96X10-4) and CLCuKoV-CP gene (2.706X10-4), whereas relatively higher substitution rates were observed for CLCuMuV-CP and CLCuKoV-Rep genes. The combination of sequences with equal and relatively low substitution rates, seemed to result in the emergence of viral isolates that caused epidemics in Pakistan and India. Our findings also suggest that CLCuMuV is spreading at an alarming rate, which can potentially be a threat to cotton production in the Indian subcontinent. PMID:26963635

  13. Recombination and the maintenance of plant organelle genome stability.

    PubMed

    Maréchal, Alexandre; Brisson, Normand

    2010-04-01

    Like their nuclear counterpart, the plastid and mitochondrial genomes of plants have to be faithfully replicated and repaired to ensure the normal functioning of the plant. Inability to maintain organelle genome stability results in plastid and/or mitochondrial defects, which can lead to potentially detrimental phenotypes. Fortunately, plant organelles have developed multiple strategies to maintain the integrity of their genetic material. Of particular importance among these processes is the extensive use of DNA recombination. In fact, recombination has been implicated in both the replication and the repair of organelle genomes. Revealingly, deregulation of recombination in organelles results in genomic instability, often accompanied by adverse consequences for plant fitness. The recent identification of four families of proteins that prevent aberrant recombination of organelle DNA sheds much needed mechanistic light on this important process. What comes out of these investigations is a partial portrait of the recombination surveillance machinery in which plants have co-opted some proteins of prokaryotic origin but have also evolved whole new factors to keep their organelle genomes intact. These new features presumably optimized the protection of plastid and mitochondrial genomes against the particular genotoxic stresses they face. PMID:20180912

  14. Application of internal standard method in recombinant luminescent bacteria test.

    PubMed

    Wang, Yong-Zhi; Li, Dan; He, Miao

    2015-09-01

    Mercury and its organic compounds have been of severe concern worldwide due to their damage to the ecosystem and human health. The development of effective and affordable technology to monitor and signal the presence of bioavailable mercury is an urgent need. The Mer gene is a mercury-responsive resistant gene, and a mercury-sensing recombinant luminescent bacterium using the Mer gene was constructed in this study. The mer operon from marine Pseudomonas putida strain SP1 was amplified and fused with prompterless luxCDABE in the pUCD615 plasmid within Escherichia coli cells, resulting in pTHE30-E. coli. The recombinant strain showed high sensitivity and specificity. The detection limit of Hg(2+) was 5nmol/L, and distinct luminescence could be detected in 30min. Cd(2+), Cu(2+), Zn(2+), Ca(2+), Pb(2+), Mg(2+), Mn(2+), and Al(3+) did not interfere with the detection over a range of 10(-5)-1mM. Application of recombinant luminescent bacteria testing in environmental samples has been a controversial issue: especially for metal-sensing recombinant strains, false negatives caused by high cytotoxicity are one of the most important issues when applying recombinant luminescent bacteria in biomonitoring of heavy metals. In this study, by establishing an internal standard approach, the false negative problem was overcome; furthermore, the method can also help to estimate the suspected mercury concentration, which ensures high detection sensitivity of bioavailable Hg(2+). PMID:26354701

  15. Recombination spot identification Based on gapped k-mers

    PubMed Central

    Wang, Rong; Xu, Yong; Liu, Bin

    2016-01-01

    Recombination is crucial for biological evolution, which provides many new combinations of genetic diversity. Accurate identification of recombination spots is useful for DNA function study. To improve the prediction accuracy, researchers have proposed several computational methods for recombination spot identification. The k-mer feature is one of the most useful features for modeling the properties and function of DNA sequences. However, it suffers from the inherent limitation. If the value of word length k is large, the occurrences of k-mers are closed to a binary variable, with a few k-mers present once and most k-mers are absent. This usually causes the sparse problem and reduces the classification accuracy. To solve this problem, we add gaps into k-mer and introduce a new feature called gapped k-mer (GKM) for identification of recombination spots. By using this feature, we present a new predictor called SVM-GKM, which combines the gapped k-mers and Support Vector Machine (SVM) for recombination spot identification. Experimental results on a widely used benchmark dataset show that SVM-GKM outperforms other highly related predictors. Therefore, SVM-GKM would be a powerful predictor for computational genomics. PMID:27030570

  16. Novel Heterotypic Rox Sites for Combinatorial Dre Recombination Strategies

    PubMed Central

    Chuang, Katherine; Nguyen, Eileen; Sergeev, Yuri; Badea, Tudor C.

    2015-01-01

    Site-specific recombinases (SSRs) such as Cre are widely used in gene targeting and genetic approaches for cell labeling and manipulation. They mediate DNA strand exchange between two DNA molecules at dedicated recognition sites. Precise understanding of the Cre recombination mechanism, including the role of individual base pairs in its loxP target site, guided the generation of mutant lox sites that specifically recombine with themselves but not with the wild type loxP. This has led to the development of a variety of combinatorial Cre-dependent genetic strategies, such as multicolor reporters, irreversible inversions, or recombination-mediated cassette exchange. Dre, a Cre-related phage integrase that recognizes roxP sites, does not cross-react with the Cre-loxP system, but has similar recombination efficiency. We have previously described intersectional genetic strategies combining Dre and Cre. We now report a mutagenesis screen aimed at identifying roxP base pairs critical for self-recognition. We describe several rox variant sites that are incompatible with roxP, but are able to efficiently recombine with themselves in either purified systems or bacterial and eukaryotic tissue culture systems. These newly identified rox sites are not recognized by Cre, thus enabling potential combinatorial strategies involving Cre, Dre, and target loci including multiple loxP and roxP variants. PMID:26715092

  17. Recombination spot identification Based on gapped k-mers.

    PubMed

    Wang, Rong; Xu, Yong; Liu, Bin

    2016-01-01

    Recombination is crucial for biological evolution, which provides many new combinations of genetic diversity. Accurate identification of recombination spots is useful for DNA function study. To improve the prediction accuracy, researchers have proposed several computational methods for recombination spot identification. The k-mer feature is one of the most useful features for modeling the properties and function of DNA sequences. However, it suffers from the inherent limitation. If the value of word length k is large, the occurrences of k-mers are closed to a binary variable, with a few k-mers present once and most k-mers are absent. This usually causes the sparse problem and reduces the classification accuracy. To solve this problem, we add gaps into k-mer and introduce a new feature called gapped k-mer (GKM) for identification of recombination spots. By using this feature, we present a new predictor called SVM-GKM, which combines the gapped k-mers and Support Vector Machine (SVM) for recombination spot identification. Experimental results on a widely used benchmark dataset show that SVM-GKM outperforms other highly related predictors. Therefore, SVM-GKM would be a powerful predictor for computational genomics. PMID:27030570

  18. Recombination coefficients of O and N radicals on stainless steel

    NASA Astrophysics Data System (ADS)

    Singh, Harmeet; Coburn, J. W.; Graves, David B.

    2000-09-01

    Surface recombination coefficients of O and N radicals in pure O2 and N2 plasmas, respectively, have been estimated on the stainless steel walls of a low-pressure inductively coupled plasma reactor. The recombination coefficients are estimated using a steady state plasma model describing the balance between the volume generation of the radicals from electron-impact dissociation of the parent molecules, and the loss of the radicals due to surface recombination. The model uses radical and parent molecule number densities and the electron energy distribution function (EEDF) as input parameters. We have measured the radical number density using appearance potential mass spectrometry. The parent neutral number density is measured using mass spectrometry. The EEDF is measured using a Langmuir probe. The recombination coefficient of O radicals on stainless steel walls at approximately 330 K is estimated to be 0.17±0.02, and agrees well with previous measurements. The recombination coefficient of N radicals is estimated to be 0.07±0.02 on stainless steel at 330 K.

  19. Recombinant host cells and media for ethanol production

    DOEpatents

    Wood, Brent E; Ingram, Lonnie O; Yomano, Lorraine P; York, Sean W

    2014-02-18

    Disclosed are recombinant host cells suitable for degrading an oligosaccharide that have been optimized for growth and production of high yields of ethanol, and methods of making and using these cells. The invention further provides minimal media comprising urea-like compounds for economical production of ethanol by recombinant microorganisms. Recombinant host cells in accordance with the invention are modified by gene mutation to eliminate genes responsible for the production of unwanted products other than ethanol, thereby increasing the yield of ethanol produced from the oligosaccharides, relative to unmutated parent strains. The new and improved strains of recombinant bacteria are capable of superior ethanol productivity and yield when grown under conditions suitable for fermentation in minimal growth media containing inexpensive reagents. Systems optimized for ethanol production combine a selected optimized minimal medium with a recombinant host cell optimized for use in the selected medium. Preferred systems are suitable for efficient ethanol production by simultaneous saccharification and fermentation (SSF) using lignocellulose as an oligosaccharide source. The invention also provides novel isolated polynucleotide sequences, polypeptide sequences, vectors and antibodies.

  20. Enhancing radiotherapy through a greater understanding of homologous recombination

    PubMed Central

    Barker, Christopher A.; Powell, Simon N.

    2016-01-01

    Radiotherapy for the treatment of cancer can cause a wide range of cellular effects, the most biologically potent of which is the double strand break in DNA. The process of repairing DNA double strand breaks involves one of two major mechanisms: non-homologous end-joining or homologous recombination. In this review, we review the molecular mechanisms of homologous recombination, in particular as it relates to the repair of DNA damage from ionizing radiation. We also present specific situations where homologous recombination may be dysfunctional in human cancers, and how this functional abnormality can be recognized. We also discuss the therapeutic opportunities that can be exploited based on deficiencies in homologous recombination at various steps in the DNA repair pathway. Side-by-side with these potential therapeutic opportunities, we review the contemporary clinical trials in which strategies to exploit these defects in homologous recombination can be enhanced by the use of radiotherapy in conjunction with biologically-targeted agents. We conclude that the field of radiation oncology has only scratched the surface of a potentially highly efficacious therapeutic strategy. PMID:20832019

  1. Directed Ig class switch recombination in activated murine B cells.

    PubMed Central

    Winter, E; Krawinkel, U; Radbruch, A

    1987-01-01

    Immunoglobulin class switch recombination occurs at frequencies of up to 10%/cell/generation in activated murine B-lymphocytes. We analysed cH gene rearrangements and switch recombinations from active and inactive IgH loci of B-cells activated in various ways and immortalized by cell fusion. Although about half of the IgM+ cells show rearrangement of c mu genes, the deletion of c mu is a rare event. Half of the IgG3+ and IgG1+ cells show rearrangement of c mu genes on the inactive IgH locus and the other half of the IgG+ cells have deleted c mu from both IgH loci by switch recombination. This recombination is directed to the same switch regions on both IgH loci in 60-80% of all cases. Interleukin 4 may play a critical role in programming murine B-lymphocytes for specific switch recombination. Images Fig. 1. Fig. 2. Fig. 6. PMID:3038529

  2. Auger recombination in InN from first principles

    NASA Astrophysics Data System (ADS)

    McAllister, Andrew; Kioupakis, Emmanouil

    Group-III Nitride materials are used in numerous electronic and optoelectronic devices including solid-state lighting, energy conversion, sensor technologies, and high-power electronics. Indium nitride in particular is interesting for fast electronics and optoelectronics in the infrared. Auger recombination is a non-radiative carrier recombination process that would limit the efficiency of these devices. The small band gap (0.7 eV) and the high intrinsic free-electron concentrations in InN possibly make Auger recombination particularly important in this material. We used first-principles computational methods to determine the Auger recombination rates in InN. Our results suggest that direct Auger recombination is dominant in this material and that phonon-assisted Auger processes are not as important as in wider-gap nitrides such as GaN. This research was supported by the National Science Foundation CAREER award through Grant No. DMR-1254314. Computational resources were provided by the DOE NERSC facility.

  3. Replication-Associated Recombinational Repair: Lessons from Budding Yeast

    PubMed Central

    Bonner, Jaclyn N.; Zhao, Xiaolan

    2016-01-01

    Recombinational repair processes multiple types of DNA lesions. Though best understood in the repair of DNA breaks, recombinational repair is intimately linked to other situations encountered during replication. As DNA strands are decorated with many types of blocks that impede the replication machinery, a great number of genomic regions cannot be duplicated without the help of recombinational repair. This replication-associated recombinational repair employs both the core recombination proteins used for DNA break repair and the specialized factors that couple replication with repair. Studies from multiple organisms have provided insights into the roles of these specialized factors, with the findings in budding yeast being advanced through use of powerful genetics and methods for detecting DNA replication and repair intermediates. In this review, we summarize recent progress made in this organism, ranging from our understanding of the classical template switch mechanisms to gap filling and replication fork regression pathways. As many of the protein factors and biological principles uncovered in budding yeast are conserved in higher eukaryotes, these findings are crucial for stimulating studies in more complex organisms. PMID:27548223

  4. Recombination shapes the structure of an environmental Vibrio cholerae population.

    PubMed

    Keymer, Daniel P; Boehm, Alexandria B

    2011-01-01

    Vibrio cholerae consists of pathogenic strains that cause sporadic gastrointestinal illness or epidemic cholera disease and nonpathogenic strains that grow and persist in coastal aquatic ecosystems. Previous studies of disease-causing strains have shown V. cholerae to be a primarily clonal bacterial species, but isolates analyzed have been strongly biased toward pathogenic genotypes, while representing only a small sample of the vast diversity in environmental strains. In this study, we characterized homologous recombination and structure among 152 environmental V. cholerae isolates and 13 other putative Vibrio isolates from coastal waters and sediments in central California, as well as four clinical V. cholerae isolates, using multilocus sequence analysis of seven housekeeping genes. Recombinant regions were identified by at least three detection methods in 72% of our V. cholerae isolates. Despite frequent recombination, significant linkage disequilibrium was still detected among the V. cholerae sequence types. Incongruent but nonrandom associations were observed for maximum likelihood topologies from the individual loci. Overall, our estimated recombination rate in V. cholerae of 6.5 times the mutation rate is similar to those of other sexual bacteria and appears frequently enough to restrict selection from purging much of the neutral intraspecies diversity. These data suggest that frequent recombination among V. cholerae may hinder the identification of ecotypes in this bacterioplankton population. PMID:21075874

  5. Systemic delivery of recombinant proteins by genetically modified myoblasts

    SciTech Connect

    Barr, E.; Leiden, J.M. )

    1991-12-06

    The ability to stably deliver recombinant proteins to the systemic circulation would facilitate the treatment of a variety of acquired and inherited diseases. To explore the feasibility of the use of genetically engineered myoblasts as a recombinant protein delivery system, stable transfectants of the murine C2C12 myoblast cell line were produced that synthesize and secrete high levels of human growth hormone (hGH) in vitro. Mice injected with hGH-transfected myoblasts had significant levels of hGH in both muscle and serum that were stable for at least 3 weeks after injection. Histological examination of muscles injected with {beta}-galactosidase-expressing C2C12 myoblasts demonstrated that many of the injected cells had fused to form multinucleated myotubes. Thus, genetically engineered myoblasts can be used for the stable delivery of recombinant proteins into the circulation.

  6. Interspecific plastidial recombination in the diatom genus Pseudo-nitzschia.

    PubMed

    D'Alelio, Domenico; Ruggiero, Maria Valeria

    2015-12-01

    Plastids are usually uni-parentally inherited and genetic recombination between these organelles is seldom observed. The genus Pseudo-nitzschia, a globally relevant marine diatom, features bi-parental plastid inheritance in the course of sexual reproduction. This observation inspired the recombination detection we pursued in this paper over a ~1,400-nucleotide-long region of the plastidial rbcL, a marker used in both molecular taxonomy and phylogenetic studies in diatoms. Among all the rbcL-sequences available in web-databases for Pseudo-nitzschia, 42 haplotypes were identified and grouped in five clusters by Bayesian phylogeny. Signs of hybridization were evident in four of five clusters, at both intra- and interspecific levels, suggesting that, in diatoms, (i) plastidial recombination is not absent and (ii) hybridization can play a role in speciation of Pseudo-nitzschia spp. PMID:26986997

  7. Tailoring Charge Recombination in Photoelectrodes Using Oxide Nanostructures.

    PubMed

    Iandolo, Beniamino; Wickman, Björn; Svensson, Elin; Paulsson, Daniel; Hellman, Anders

    2016-04-13

    Optimizing semiconductor devices for solar energy conversion requires an explicit control of the recombination of photogenerated electron-hole pairs. Here we show how the recombination of charge carriers can be controlled in semiconductor thin films by surface patterning with oxide nanodisks. The control mechanism relies on the formation of dipole-like electric fields at the interface that, depending on the field direction, attract or repel minority carriers from underneath the disks. The charge recombination rate can be controlled through the choice of oxide material and the surface coverage of nanodisks. We provide proof-of-principle demonstration of this approach by patterning the surface of Fe2O3, one of the most studied semiconductors for light-driven water splitting, with TiO2 and Cu2O nanodisks. We expect this method to be generally applicable to a range of semiconductor-based solar energy conversion devices. PMID:26978576

  8. Self-assembly of tunable protein suprastructures from recombinant oleosin

    PubMed Central

    Vargo, Kevin B.; Parthasarathy, Ranganath; Hammer, Daniel A.

    2012-01-01

    Using recombinant amphiphilic proteins to self-assemble suprastructures would allow precise control over surfactant chemistry and the facile incorporation of biological functionality. We used cryo-TEM to confirm self-assembled structures from recombinantly produced mutants of the naturally occurring sunflower protein, oleosin. We studied the phase behavior of protein self-assembly as a function of solution ionic strength and protein hydrophilic fraction, observing nanometric fibers, sheets, and vesicles. Vesicle membrane thickness correlated with increasing hydrophilic fraction for a fixed hydrophobic domain length. The existence of a bilayer membrane was corroborated in giant vesicles through the localized encapsulation of hydrophobic Nile red and hydrophilic calcein. Circular dichroism revealed that changes in nanostructural morphology in this family of mutants was unrelated to changes in secondary structure. Ultimately, we envision the use of recombinant techniques to introduce novel functionality into these materials for biological applications. PMID:22753512

  9. Meiotic recombination and the crossover assurance checkpoint in Caenorhabditis elegans.

    PubMed

    Yu, Zhouliang; Kim, Yumi; Dernburg, Abby F

    2016-06-01

    During meiotic prophase, chromosomes pair and synapse with their homologs and undergo programmed DNA double-strand break (DSB) formation to initiate meiotic recombination. These DSBs are processed to generate a limited number of crossover recombination products on each chromosome, which are essential to ensure faithful segregation of homologous chromosomes. The nematode Caenorhabditis elegans has served as an excellent model organism to investigate the mechanisms that drive and coordinate these chromosome dynamics during meiosis. Here we focus on our current understanding of the regulation of DSB induction in C. elegans. We also review evidence that feedback regulation of crossover formation prolongs the early stages of meiotic prophase, and discuss evidence that this can alter the recombination pattern, most likely by shifting the genome-wide distribution of DSBs. PMID:27013114

  10. RECOMBINATION RATE COEFFICIENTS OF BORON-LIKE Ne

    SciTech Connect

    Mahmood, S.; Orban, I.; Ali, S.; Schuch, R.; Glans, P.; Bleda, E. A.; Altun, Z.

    2013-07-10

    Recombination of Ne{sup 5+} was measured in a merged-beam type experiment at the heavy-ion storage ring CRYRING. In the collision energy range 0-110 eV resonances due to 2s {sup 2}2p {yields} 2s2p {sup 2} ({Delta}n = 0) and 2s {sup 2}2p {yields} 2s {sup 2}3l ({Delta}n = 1), core excitations were observed. The experimentally derived rate coefficients agree well with the calculations obtained using AUTOSTRUCTURE. At low energies, recombination is dominated by resonances belonging to the spin-forbidden 2s2p {sup 2}({sup 4} P{sub J})nl series. The energy-dependent rate coefficients were convoluted with a Maxwell-Boltzmann electron energy distribution to obtain plasma recombination rate coefficients. The data from the literature deviate from the measured results at low temperature.

  11. Cre-mediated site-specific recombination in zebrafish embryos.

    PubMed

    Thummel, Ryan; Burket, Christopher T; Brewer, Jeffrey L; Sarras, Michael P; Li, Li; Perry, Martin; McDermott, Jeffrey P; Sauer, Brian; Hyde, David R; Godwin, Alan R

    2005-08-01

    Cre-mediated site-specific recombination has become an invaluable tool for manipulation of the murine genome. The ability to conditionally activate gene expression or to generate chromosomal alterations with this same tool would greatly enhance zebrafish genetics. This study demonstrates that the HSP70 promoter can be used to inducibly control expression of an enhanced green fluorescent protein (EGFP) -Cre fusion protein. The EGFP-Cre fusion protein is capable of promoting recombination between lox sites in injected plasmids or in stably inherited transgenes as early as 2 hr post-heat shock induction. Finally, the levels of Cre expression achieved in a transgenic fish line carrying the HSP70-EGFP-cre transgene are compatible with viability and both male and female transgenic fish are fertile subsequent to induction of EGFP-Cre expression. Hence, our data suggests that Cre-mediated recombination is a viable means of manipulating gene expression in zebrafish. PMID:15977183

  12. Historical Perspectives Pertaining to the NIH Recombinant DNA Advisory Committee

    PubMed Central

    2014-01-01

    Abstract Science is host to a constantly emerging series of new paradigms, and it is this characteristic that makes science both interesting and dynamic. As a part of this continuum, it became possible to create recombinant DNA molecules. Immediately it was recognized that there was a potential for serious adverse events associated with this new technology. Following two scientific conferences at Asilomar, California, the National Institutes of Health moved quickly to create the Recombinant DNA Advisory Committee (RAC). For approximately 38 years the RAC has served as an open forum for review of various recombinant DNA experiments, and for the last 23 years it has played a pivotal role in the oversight of human gene therapy. The RAC's existence obviated the need for more restrictive governmental legislation and has supported the development of genetic interventions that are leading to actual human therapies. PMID:24444182

  13. Matter-wave recombiners for trapped Bose-Einstein condensates

    NASA Astrophysics Data System (ADS)

    Berrada, T.; van Frank, S.; Bücker, R.; Schumm, T.; Schaff, J.-F.; Schmiedmayer, J.; Julía-Díaz, B.; Polls, A.

    2016-06-01

    Interferometry with trapped atomic Bose-Einstein condensates (BECs) requires the development of techniques to recombine the two paths of the interferometer and map the accumulated phase difference to a measurable atom number difference. We have implemented and compared two recombining procedures in a double-well-based BEC interferometer. The first procedure utilizes the bosonic Josephson effect and controlled tunneling of atoms through the potential barrier, similar to laser light in an optical fiber coupler. The second one relies on the interference of the reflected and transmitted parts of the BEC wave function when impinging on the potential barrier, analogous to light impinging on a half-silvered mirror. Both schemes were implemented successfully, yielding an interferometric contrast of ˜20 % and 42% respectively. Building efficient matter-wave recombiners represents an important step towards the coherent manipulation of external quantum superposition states of BECs.

  14. Mechanisms underlying allergy vaccination with recombinant hypoallergenic allergen derivatives

    PubMed Central

    Linhart, Birgit; Valenta, Rudolf

    2015-01-01

    Hundred years ago therapeutic vaccination with allergen-containing extracts has been introduced as a clinically effective, disease-modifying, allergen-specific and long-lasting form of therapy for allergy, a hypersensitivity disease affecting more than 25% of the population. Today, the structures of most of the disease-causing allergens have been elucidated and recombinant hypoallergenic allergen derivatives with reduced allergenic activity have been engineered to reduce side effects during allergen-specific immunotherapy (SIT). These recombinant hypoallergens have been characterized in vitro, in experimental animal models and in clinical trials in allergic patients. This review provides a summary of the molecular, immunological and preclinical evaluation criteria applied for this new generation of allergy vaccines. Furthermore, we summarize the mechanisms underlying SIT with recombinant hypoallergens which are thought to be responsible for their therapeutic effect. PMID:22100888

  15. A recombineering-based gene tagging system for Arabidopsis.

    PubMed

    Alonso, Jose M; Stepanova, Anna N

    2015-01-01

    Many of the experimental approaches aimed at studying gene function heavily rely on the ability to make precise modifications in the gene's DNA sequence. Homologous recombination (HR)-based strategies provide a convenient way to create such types of modifications. HR-based DNA sequence manipulations can be enormously facilitated by expressing in E. coli a small set of bacteriophage proteins that make the exchange of DNA between a linear donor and the target DNA molecules extremely efficient. These in vivo recombineering techniques have been incorporated as essential components of the molecular toolbox in many model organisms. In this chapter, we describe the experimental procedures involved in recombineering-based tagging of an Arabidopsis gene contained in a plant transformation-ready bacterial artificial chromosome (TAC). PMID:25239749

  16. Optimising yeast as a host for recombinant protein production (review).

    PubMed

    Bonander, Nicklas; Bill, Roslyn M

    2012-01-01

    Having access to suitably stable, functional recombinant protein samples underpins diverse academic and industrial research efforts to understand the workings of the cell in health and disease. Synthesising a protein in recombinant host cells typically allows the isolation of the pure protein in quantities much higher than those found in the protein's native source. Yeast is a popular host as it is a eukaryote with similar synthetic machinery to the native human source cells of many proteins of interest, while also being quick, easy, and cheap to grow and process. Even in these cells the production of some proteins can be plagued by low functional yields. We have identified molecular mechanisms and culture parameters underpinning high yields and have consolidated our findings to engineer improved yeast cell factories. In this chapter, we provide an overview of the opportunities available to improve yeast as a host system for recombinant protein production. PMID:22454109

  17. Attenuated vaccines can recombine to form virulent field viruses.

    PubMed

    Lee, Sang-Won; Markham, Philip F; Coppo, Mauricio J C; Legione, Alistair R; Markham, John F; Noormohammadi, Amir H; Browning, Glenn F; Ficorilli, Nino; Hartley, Carol A; Devlin, Joanne M

    2012-07-13

    Recombination between herpesviruses has been seen in vitro and in vivo under experimental conditions. This has raised safety concerns about using attenuated herpesvirus vaccines in human and veterinary medicine and adds to other known concerns associated with their use, including reversion to virulence and disease arising from recurrent reactivation of lifelong chronic infection. We used high-throughput sequencing to investigate relationships between emergent field strains and vaccine strains of infectious laryngotracheitis virus (ILTV, gallid herpesvirus 1). We show that independent recombination events between distinct attenuated vaccine strains resulted in virulent recombinant viruses that became the dominant strains responsible for widespread disease in Australian commercial poultry flocks. These findings highlight the risks of using multiple different attenuated herpesvirus vaccines, or vectors, in the same populations. PMID:22798607

  18. Production of biologically active recombinant human lactoferrin in Aspergillus oryzae.

    PubMed

    Ward, P P; Lo, J Y; Duke, M; May, G S; Headon, D R; Conneely, O M

    1992-07-01

    We report the production of recombinant human lactoferrin in Aspergillus oryzae. Expression of human lactoferrin (hLF), a 78 kD glycoprotein, was achieved by placing the cDNA under the control of the A. oryzae alpha-amylase promoter and the 3' flanking region of the A. niger glucoamylase gene. Using this system, hLF is expressed and secreted into the growth medium at levels up to 25 mg/l. The recombinant lactoferrin is indistinguishable from human milk lactoferrin with respect to its size, immunoreactivity, and iron-binding capacity. The recombinant protein appears to be appropriately N-linked glycosylated and correctly processed at the N-terminus by the A. oryzae secretory apparatus. Lactoferrin is the largest heterologous protein and the first mammalian glycoprotein expressed in the Aspergillus system to date. Hence, this expression system appears suitable for the large-scale production and secretion of biologically active mammalian glycoproteins. PMID:1368268

  19. Recombinant lentivector as a genetic immunization vehicle for antitumor immunity

    PubMed Central

    He, Yukai; Munn, David; Falo, Louis D

    2011-01-01

    Summary Encouraged by remarkable successes in preventing infectious diseases and by the well established potential of immune system for controlling tumor growth, active therapeutic immunization approaches hold great promise for treating malignant tumors. In recent years, engineered recombinant viral vectors have been carefully examined as genetic immunization vehicles and have been demonstrated to induce potent T cell mediated immune responses that can control tumor growth. Very recent efforts suggest that lentivectors possess important advantages over other candidate recombinant viral vectors for genetic immunization. Here we review the development of recombinant lentivectors and the characteristics of T cell immune responses elicited by lentivector immunization, including the mechanism of T cell priming with a focus on the role of skin dendritic cells (DC) and potential applications for tumor immunotherapy. PMID:18377355

  20. An Overview of the Molecular Mechanisms of Recombinational DNA Repair.

    PubMed

    Kowalczykowski, Stephen C

    2015-11-01

    Recombinational DNA repair is a universal aspect of DNA metabolism and is essential for genomic integrity. It is a template-directed process that uses a second chromosomal copy (sister, daughter, or homolog) to ensure proper repair of broken chromosomes. The key steps of recombination are conserved from phage through human, and an overview of those steps is provided in this review. The first step is resection by helicases and nucleases to produce single-stranded DNA (ssDNA) that defines the homologous locus. The ssDNA is a scaffold for assembly of the RecA/RAD51 filament, which promotes the homology search. On finding homology, the nucleoprotein filament catalyzes exchange of DNA strands to form a joint molecule. Recombination is controlled by regulating the fate of both RecA/RAD51 filaments and DNA pairing intermediates. Finally, intermediates that mature into Holliday structures are disjoined by either nucleolytic resolution or topological dissolution. PMID:26525148

  1. Recombination and collisional X-UV lasers at ORSAY

    SciTech Connect

    Klisnick, A.; Carillon, A.; Dhez, P.; Goedtkindt, P.; Guennou, H.; Jamelot, G.; Jaegle, P.; Sureau, A.; Rus, B.; Zeitoun, P. ); Chenais-Popovics, C.; Renaudin, P.; Rancu, O.; Gauthier, J.C. ); Back, C.A. )

    1995-01-10

    In this paper we describe the progress achieved recently in our laboratory in the field of X-ray lasers. Both collisional excitation and recombination pumped systems are under investigation. We show that the 5g-4f transition in lithium-like ions could bring out a significant increase of the gain-length accessible with recombination X-ray lasers. We present preliminary results on an absorption spectroscopy experiment designed to probe the ionization state of recombination X-ray laser plasmas. Finally we report on the observation of a strong amplified signal at 212 A, the wavelength of a 3p-3s (J=0--1) in neon-like zinc. [copyright] 1995 [ital American] [ital Institute] [ital of] [ital Physics

  2. Plant cell cultures for the production of recombinant proteins.

    PubMed

    Hellwig, Stephan; Drossard, Jürgen; Twyman, Richard M; Fischer, Rainer

    2004-11-01

    The use of whole plants for the synthesis of recombinant proteins has received a great deal of attention recently because of advantages in economy, scalability and safety compared with traditional microbial and mammalian production systems. However, production systems that use whole plants lack several of the intrinsic benefits of cultured cells, including the precise control over growth conditions, batch-to-batch product consistency, a high level of containment and the ability to produce recombinant proteins in compliance with good manufacturing practice. Plant cell cultures combine the merits of whole-plant systems with those of microbial and animal cell cultures, and already have an established track record for the production of valuable therapeutic secondary metabolites. Although no recombinant proteins have yet been produced commercially using plant cell cultures, there have been many proof-of-principle studies and several companies are investigating the commercial feasibility of such production systems. PMID:15529167

  3. Replication Origin Selection Regulates the Distribution of Meiotic Recombination

    PubMed Central

    Wu, Pei-Yun Jenny; Nurse, Paul

    2014-01-01

    Summary The program of DNA replication, defined by the temporal and spatial pattern of origin activation, is altered during development and in cancers. However, whether changes in origin usage play a role in regulating specific biological processes remains unknown. We investigated the consequences of modifying origin selection on meiosis in fission yeast. Genome-wide changes in the replication program of premeiotic S phase do not affect meiotic progression, indicating that meiosis neither activates nor requires a particular origin pattern. In contrast, local changes in origin efficiencies between different replication programs lead to changes in Rad51 recombination factor binding and recombination frequencies in these domains. We observed similar results for Rad51 when changes in efficiencies were generated by directly targeting expression of the Cdc45 replication factor. We conclude that origin selection is a key determinant for organizing meiotic recombination, providing evidence that genome-wide modifications in replication program can modulate cellular physiology. PMID:24560273

  4. Challenges and opportunities in the purification of recombinant tagged proteins.

    PubMed

    Pina, Ana Sofia; Lowe, Christopher R; Roque, Ana Cecília A

    2014-01-01

    The purification of recombinant proteins by affinity chromatography is one of the most efficient strategies due to the high recovery yields and purity achieved. However, this is dependent on the availability of specific affinity adsorbents for each particular target protein. The diversity of proteins to be purified augments the complexity and number of specific affinity adsorbents needed, and therefore generic platforms for the purification of recombinant proteins are appealing strategies. This justifies why genetically encoded affinity tags became so popular for recombinant protein purification, as these systems only require specific ligands for the capture of the fusion protein through a pre-defined affinity tag tail. There is a wide range of available affinity pairs "tag-ligand" combining biological or structural affinity ligands with the respective binding tags. This review gives a general overview of the well-established "tag-ligand" systems available for fusion protein purification and also explores current unconventional strategies under development. PMID:24334194

  5. Illegitimate replication of linear hepadnavirus DNA through nonhomologous recombination.

    PubMed Central

    Yang, W; Summers, J

    1995-01-01

    Linear hepadnavirus DNA in primary hepatocyte cultures efficiently participates in intra- and intermolecular nonhomologous recombination at its ends. The products of this recombination are (i) monomeric covalently closed circular DNAs (cccDNAs) with deletions and insertions around the site of joining and (ii) oligomeric forms in which monomers are joined near the ends in random orientation. A fraction of monomeric cccDNAs can serve as intermediates in further DNA replication through at least five generations of nonhomologous recombination in a process we call illegitimate replication. We suggest that the monomeric and oligomeric linear DNAs produced by illegitimate replication may be precursors of the integrated and other high-molecular-weight hepadnaviral DNA forms seen in chronic infection. PMID:7769660

  6. Unraveling a Region-Specific Hyper-Recombination Phenomenon: Genetic Control and Modalities of Terminal Recombination in Escherichia Coli

    PubMed Central

    Corre, J.; Cornet, F.; Patte, J.; Louarn, J. M.

    1997-01-01

    The propensity of the terminus of the Escherichia coli chromosome for recombination has been further explored, using a test based on the selectable loss of a λ prophage inserted between repeated sequences from Tn10. Terminal recombination appears region-specific and unrelated to replication termination in a strain harboring a major chromosomal rearrangement. It requires RecBC(D) activity and must therefore occur between sister chromosomes, to conserve genomic integrity in spite of DNA degradation by RecBCD. Terminal recombination is maximal in the dif region and its intensity on either side of this recombination site depends on the orientation of the repeated sequences, probably because of the single χ site present in each repeat. Additional observations support the model that the crossover is initiated by single-strand invasion between sister chromosomes followed by RecBCD action as a consequence of DNA breakage due to the initial invasion event. Crossover location within repeats inserted at dif position supports the possibility that sister chromosomes are tightly paired in the centre of the terminal recombination zone. These data reinforce the model that postreplicative reconstruction of nucleoid organization creates a localized synapsis between the termini of sister chromosomes. PMID:9383046

  7. A Network Approach to Analyzing Highly Recombinant Malaria Parasite Genes

    PubMed Central

    Larremore, Daniel B.; Clauset, Aaron; Buckee, Caroline O.

    2013-01-01

    The var genes of the human malaria parasite Plasmodium falciparum present a challenge to population geneticists due to their extreme diversity, which is generated by high rates of recombination. These genes encode a primary antigen protein called PfEMP1, which is expressed on the surface of infected red blood cells and elicits protective immune responses. Var gene sequences are characterized by pronounced mosaicism, precluding the use of traditional phylogenetic tools that require bifurcating tree-like evolutionary relationships. We present a new method that identifies highly variable regions (HVRs), and then maps each HVR to a complex network in which each sequence is a node and two nodes are linked if they share an exact match of significant length. Here, networks of var genes that recombine freely are expected to have a uniformly random structure, but constraints on recombination will produce network communities that we identify using a stochastic block model. We validate this method on synthetic data, showing that it correctly recovers populations of constrained recombination, before applying it to the Duffy Binding Like-α (DBLα) domain of var genes. We find nine HVRs whose network communities map in distinctive ways to known DBLα classifications and clinical phenotypes. We show that the recombinational constraints of some HVRs are correlated, while others are independent. These findings suggest that this micromodular structuring facilitates independent evolutionary trajectories of neighboring mosaic regions, allowing the parasite to retain protein function while generating enormous sequence diversity. Our approach therefore offers a rigorous method for analyzing evolutionary constraints in var genes, and is also flexible enough to be easily applied more generally to any highly recombinant sequences. PMID:24130474

  8. Yeast meiotic mutants proficient for the induction of ectopic recombination.

    PubMed Central

    Engebrecht, J; Masse, S; Davis, L; Rose, K; Kessel, T

    1998-01-01

    A screen was designed to identify Saccharomyces cerevisiae mutants that were defective in meiosis yet proficient for meiotic ectopic recombination in the return-to-growth protocol. Seven mutants alleles were isolated; two are important for chromosome synapsis (RED1, MEK1) and five function independently of recombination (SPO14, GSG1, SPOT8/MUM2, 3, 4). Similar to the spoT8-1 mutant, mum2 deletion strains do not undergo premeiotic DNA synthesis, arrest prior to the first meiotic division and fail to sporulate. Surprisingly, although DNA replication does not occur, mum2 mutants are induced for high levels of ectopic recombination. gsg1 diploids are reduced in their ability to complete premeiotic DNA synthesis and the meiotic divisions, and a small percentage of cells produce spores. mum3 mutants sporulate poorly and the spores produced are inviable. Finally, mum4-1 mutants produce inviable spores. The meiotic/sporulation defects of gsg1, mum2, and mum3 are not relieved by spo11 or spo13 mutations, indicating that the mutant defects are not dependent on the initiation of recombination or completion of both meiotic divisions. In contrast, the spore inviability of the mum4-1 mutant is rescued by the spo13 mutation. The mum4-1 spo13 mutant undergoes a single, predominantly equational division, suggesting that MUM4 functions at or prior to the first meiotic division. Although recombination is variably affected in the gsg1 and mum mutants, we hypothesize that these mutants define genes important for aspects of meiosis not directly related to recombination. PMID:9504908

  9. Recombinant production of mecasermin in E. coli expression system.

    PubMed

    Jafari, S; Babaeipour, V; Seyedi, H A Eslampanah; Rahaie, M; Mofid, M R; Haddad, L; Namvaran, M M; Fallah, J

    2014-01-01

    Human Insulin-like growth factor 1 (hIGF-1) consists of 70 amino acids in a single chain with three intermolecular disulfide bridges possessing valuable therapeutic effects. To date, numerous variants of specifically engineered hIGF-1 have been produced so as to improve hIGF-1 biological activity, stability and stronger binding to IGF-1 receptor. Mecasermin is one of the modified variants with one amino acid substitution near the N-terminal (T4I) approved for the treatment of growth failure diabetes, wound healing, amyotrophic lateral sclerosis and severe primary IGF-1 deficiency. No scientific report for recombinant production of mecasermin in Escherichia coli (E. coli) expression system has been sofar reported. In the present study, we therefore investigated the overexpression of mecasermin in two different E. coli strains in order to obtain higher yield of recombinant protein. To achieve this goal, mecasermin DNA encoding sequence was designed based on polypeptide sequence, optimized according to E. coli codon preference, and cloned in pET15b. Recombinant vector, pET15-mecasermin, transferred into two E. coli strains rigami B (DE3) and BL21 (DE3) and induced for expression in a small scale. Results revealed the E. coli Origami B (DE3) expression system was a preferable host for mecasermin production due to its high expression level being around twice as much as BL21 (DE3). Large scale mecasermin production was performed in batch culture and produced recombinant protein specifically confirmed by western blotting and mass spectroscopy. Since major part of recombinant mecasermin was expressed as inclusion body, isolation and refolding was accomplished through developed purification procedure, and finally recombinant protein was successfully purified by gel filtration chromatography. PMID:26339260

  10. Recombinant Rp1 genes confer necrotic or nonspecific resistance phenotypes.

    PubMed

    Smith, Shavannor M; Steinau, Martin; Trick, Harold N; Hulbert, Scot H

    2010-06-01

    Genes at the Rp1 rust resistance locus of maize confer race-specific resistance to the common rust fungus Puccinia sorghi. Three variant genes with nonspecific effects (HRp1 -Kr1N, -D*21 and -MD*19) were found to be generated by intragenic crossing over within the LRR region. The LRR region of most NBS-LRR encoding genes is quite variable and codes for one of the regions in resistance gene proteins that controls specificity. Sequence comparisons demonstrated that the Rp1-Kr1N recombinant gene was identical to the N-terminus of the rp1-kp2 gene and C-terminus of another gene from its HRp1-K grandparent. The Rp1-D*21 recombinant gene consists of the N-terminus of the rp1-dp2 gene and C-terminus of the Rp1-D gene from the parental haplotype. Similarly, a recombinant gene from the Rp1-MD*19 haplotype has the N-terminus of an rp1 gene from the HRp1-M parent and C-terminus of the rp1-D19 gene from the HRp1-D parent. The recombinant Rp1 -Kr1N, -D*21 and -MD*19 genes activated defense responses in the absence of their AVR proteins triggering HR (hypersensitive response) in the absence of the pathogen. The results indicate that the frequent intragenic recombination events that occur in the Rp1 gene cluster not only recombine the genes into novel haplotypes, but also create genes with nonspecific effects. Some of these may contribute to nonspecific quantitative resistance but others have severe consequences for the fitness of the plant. PMID:20443026

  11. Multimer Formation Explains Allelic Suppression of PRDM9 Recombination Hotspots.

    PubMed

    Baker, Christopher L; Petkova, Pavlina; Walker, Michael; Flachs, Petr; Mihola, Ondrej; Trachtulec, Zdenek; Petkov, Petko M; Paigen, Kenneth

    2015-09-01

    Genetic recombination during meiosis functions to increase genetic diversity, promotes elimination of deleterious alleles, and helps assure proper segregation of chromatids. Mammalian recombination events are concentrated at specialized sites, termed hotspots, whose locations are determined by PRDM9, a zinc finger DNA-binding histone methyltransferase. Prdm9 is highly polymorphic with most alleles activating their own set of hotspots. In populations exhibiting high frequencies of heterozygosity, questions remain about the influences different alleles have in heterozygous individuals where the two variant forms of PRDM9 typically do not activate equivalent populations of hotspots. We now find that, in addition to activating its own hotspots, the presence of one Prdm9 allele can modify the activity of hotspots activated by the other allele. PRDM9 function is also dosage sensitive; Prdm9+/- heterozygous null mice have reduced numbers and less active hotspots and increased numbers of aberrant germ cells. In mice carrying two Prdm9 alleles, there is allelic competition; the stronger Prdm9 allele can partially or entirely suppress chromatin modification and recombination at hotspots of the weaker allele. In cell cultures, PRDM9 protein variants form functional heteromeric complexes which can bind hotspots sequences. When a heteromeric complex binds at a hotspot of one PRDM9 variant, the other PRDM9 variant, which would otherwise not bind, can still methylate hotspot nucleosomes. We propose that in heterozygous individuals the underlying molecular mechanism of allelic suppression results from formation of PRDM9 heteromers, where the DNA binding activity of one protein variant dominantly directs recombination initiation towards its own hotspots, effectively titrating down recombination by the other protein variant. In natural populations with many heterozygous individuals, allelic competition will influence the recombination landscape. PMID:26368021

  12. Adaptive evolution of a lactose-consuming Saccharomyces cerevisiae recombinant.

    PubMed

    Guimarães, Pedro M R; François, Jean; Parrou, Jean Luc; Teixeira, José A; Domingues, Lucília

    2008-03-01

    The construction of Saccharomyces cerevisiae strains that ferment lactose has biotechnological interest, particularly for cheese whey fermentation. A flocculent lactose-consuming S. cerevisiae recombinant expressing the LAC12 (lactose permease) and LAC4 (beta-galactosidase) genes of Kluyveromyces lactis was constructed previously but showed poor efficiency in lactose fermentation. This strain was therefore subjected to an evolutionary engineering process (serial transfer and dilution in lactose medium), which yielded an evolved recombinant strain that consumed lactose twofold faster, producing 30% more ethanol than the original recombinant. We identified two molecular events that targeted the LAC construct in the evolved strain: a 1,593-bp deletion in the intergenic region (promoter) between LAC4 and LAC12 and a decrease of the plasmid copy number by about 10-fold compared to that in the original recombinant. The results suggest that the intact promoter was unable to mediate the induction of the transcription of LAC4 and LAC12 by lactose in the original recombinant and that the deletion established the transcriptional induction of both genes in the evolved strain. We propose that the tuning of the expression of the heterologous LAC genes in the evolved recombinant was accomplished by the interplay between the decreased copy number of both genes and the different levels of transcriptional induction for LAC4 and LAC12 resulting from the changed promoter structure. Nevertheless, our results do not exclude other possible mutations that may have contributed to the improved lactose fermentation phenotype. This study illustrates the usefulness of simple evolutionary engineering approaches in strain improvement. The evolved strain efficiently fermented threefold-concentrated cheese whey, providing an attractive alternative for the fermentation of lactose-based media. PMID:18245248

  13. Expression of Recombinant Proteins in the Methylotrophic Yeast Pichia pastoris

    PubMed Central

    Weidner, Maria; Taupp, Marcus; Hallam, Steven J.

    2010-01-01

    Protein expression in the microbial eukaryotic host Pichia pastoris offers the possibility to generate high amounts of recombinant protein in a fast and easy to use expression system. As a single-celled microorganism P. pastoris is easy to manipulate and grows rapidly on inexpensive media at high cell densities. Being a eukaryote, P. pastoris is able to perform many of the post-translational modifications performed by higher eukaryotic cells and the obtained recombinant proteins undergo protein folding, proteolytic processing, disulfide bond formation and glycosylation [1]. As a methylotrophic yeast P. pastoris is capable of metabolizing methanol as its sole carbon source. The strong promoter for alcohol oxidase, AOX1, is tightly regulated and induced by methanol and it is used for the expression of the gene of interest. Accordingly, the expression of the foreign protein can be induced by adding methanol to the growth medium [2; 3]. Another important advantage is the secretion of the recombinant protein into the growth medium, using a signal sequence to target the foreign protein to the secretory pathway of P. pastoris. With only low levels of endogenous protein secreted to the media by the yeast itself and no added proteins to the media, a heterologous protein builds the majority of the total protein in the medium and facilitates following protein purification steps [3; 4]. The vector used here (pPICZαA) contains the AOX1 promoter for tightly regulated, methanol-induced expression of the gene of interest; the α-factor secretion signal for secretion of the recombinant protein, a Zeocin resistance gene for selection in both E. coli and Pichia and a C-terminal peptide containing the c-myc epitope and a polyhistidine (6xHis) tag for detection and purification of a recombinant protein. We also show western blot analysis of the recombinant protein using the specific Anti-myc-HRP antibody recognizing the c-myc epitope on the parent vector. PMID:20186119

  14. Recombinant production of mecasermin in E. coli expression system

    PubMed Central

    Jafari, S.; Babaeipour, V.; Seyedi, H.A. Eslampanah; Rahaie, M.; Mofid, M.R.; Haddad, L.; Namvaran, M.M.; Fallah, J.

    2014-01-01

    Human Insulin-like growth factor 1 (hIGF-1) consists of 70 amino acids in a single chain with three intermolecular disulfide bridges possessing valuable therapeutic effects. To date, numerous variants of specifically engineered hIGF-1 have been produced so as to improve hIGF-1 biological activity, stability and stronger binding to IGF-1 receptor. Mecasermin is one of the modified variants with one amino acid substitution near the N-terminal (T4I) approved for the treatment of growth failure diabetes, wound healing, amyotrophic lateral sclerosis and severe primary IGF-1 deficiency. No scientific report for recombinant production of mecasermin in Escherichia coli (E. coli) expression system has been sofar reported. In the present study, we therefore investigated the overexpression of mecasermin in two different E. coli strains in order to obtain higher yield of recombinant protein. To achieve this goal, mecasermin DNA encoding sequence was designed based on polypeptide sequence, optimized according to E. coli codon preference, and cloned in pET15b. Recombinant vector, pET15-mecasermin, transferred into two E. coli strains rigami B (DE3) and BL21 (DE3) and induced for expression in a small scale. Results revealed the E. coli Origami B (DE3) expression system was a preferable host for mecasermin production due to its high expression level being around twice as much as BL21 (DE3). Large scale mecasermin production was performed in batch culture and produced recombinant protein specifically confirmed by western blotting and mass spectroscopy. Since major part of recombinant mecasermin was expressed as inclusion body, isolation and refolding was accomplished through developed purification procedure, and finally recombinant protein was successfully purified by gel filtration chromatography. PMID:26339260

  15. Oligonucleotide recombination in corynebacteria without the expression of exogenous recombinases.

    PubMed

    Krylov, Alexander A; Kolontaevsky, Egor E; Mashko, Sergey V

    2014-10-01

    Brevibacterium lactofermentum and Corynebacterium glutamicum are important biotechnology species of the genus Corynebacterium. The single-strand DNA annealing protein (SSAP)-independent oligonucleotide-mediated recombination procedure was successfully applied to the commonly used wild-type strains B. lactofermentum AJ1511 and C. glutamicum ATCC13032. When the rpsL gene was used as a target, the optimized protocol yielded up to (1.4±0.3)×10(3) and (6.7±1.3)×10(3) streptomycin-resistant colonies per 10(8) viable cells for the corresponding strains. We tested the influence of several parameters that are known to enhance the efficiency of oligonucleotide-mediated recombination in other bacterial species. Among them, increasing the concentration of oligonucleotides and targeting the lagging strand of the chromosome have proven to have positive effects on both of the tested species. No difference in the efficiency of recombination was observed between the oligonucleotides phosphorothiorated at the 5' ends and the unmodified oligonucleotides or between the oligonucleotides with four mutated nucleotides and those with one mutated nucleotide. The described approach demonstrates that during the adaptation of the recombineering technique, testing SSAP-independent oligonucleotide-mediated recombination could be a good starting point. Such testing could decrease the probability of an incorrect interpretation of the effect of exogenous protein factors (such as SSAP and/or corresponding exonucleases) due to non-optimal experimental conditions. In addition, SSAP-independent recombination itself could be useful in combination with suitable selection/enrichment methods. PMID:25087479

  16. Evidence and Consequence of Porcine Endogenous Retrovirus Recombination

    PubMed Central

    Bartosch, Birke; Stefanidis, Dimitrios; Myers, Richard; Weiss, Robin; Patience, Clive; Takeuchi, Yasuhiro

    2004-01-01

    The genetic nature and biological effects of recombination between porcine endogenous retroviruses (PERV) were studied. An infectious molecular clone was generated from a high-titer, human-tropic PERV isolate, PERV-A 14/220 (B. A. Oldmixon, et al. J. Virol. 76:3045-3048, 2002; T. A. Ericsson et al. Proc. Natl. Acad. Sci. USA 100:6759-6764, 2003). To analyze this sequence and 15 available full-length PERV nucleotide sequences, we developed a sequence comparison program, LOHATM to calculate local sequence homology between two sequences. This analysis determined that PERV-A 14/220 arose by homologous recombination of a PERV-C genome replacing an 850-bp region around the pol-env junction with that of a PERV-A sequence. This 850-bp PERV-A sequence encompasses the env receptor binding domain, thereby conferring a wide host range including human cells. In addition, we determined that multiple regions derived from PERV-C are responsible for the increased infectious titer of PERV-A 14/220. Thus, a single recombination event may be a fast and effective way to generate high-titer, potentially harmful PERV. Further, local homology and phylogenetic analyses between 16 full-length sequences revealed evidence for other recombination events in the past that give rise to other PERV genomes that possess the PERV-A, but not the PERV-B, env gene. These results indicate that PERV-A env is more prone to recombination with heterogeneous backbone genomes than PERV-B env. Such recombination events that generate more active PERV-A appear to occur in pigs rather frequently, which increases the potential risk of zoonotic PERV transmission. In this context, pigs lacking non-human-tropic PERV-C would be more suitable as donor animals for clinical xenotransplantation. PMID:15564496

  17. Phylogenetic Mapping of Recombination Hotspots in Human Immunodeficiency Virus via Spatially Smoothed Change-Point Processes

    PubMed Central

    Minin, Vladimir N.; Dorman, Karin S.; Fang, Fang; Suchard, Marc A.

    2007-01-01

    We present a Bayesian framework for inferring spatial preferences of recombination from multiple putative recombinant nucleotide sequences. Phylogenetic recombination detection has been an active area of research for the last 15 years. However, only recently attempts to summarize information from several instances of recombination have been made. We propose a hierarchical model that allows for simultaneous inference of recombination breakpoint locations and spatial variation in recombination frequency. The dual multiple change-point model for phylogenetic recombination detection resides at the lowest level of our hierarchy under the umbrella of a common prior on breakpoint locations. The hierarchical prior allows for information about spatial preferences of recombination to be shared among individual data sets. To overcome the sparseness of breakpoint data, dictated by the modest number of available recombinant sequences, we a priori impose a biologically relevant correlation structure on recombination location log odds via a Gaussian Markov random field hyperprior. To examine the capabilities of our model to recover spatial variation in recombination frequency, we simulate recombination from a predefined distribution of breakpoint locations. We then proceed with the analysis of 42 human immunodeficiency virus (HIV) intersubtype gag recombinants and identify a putative recombination hotspot. PMID:17194781

  18. Charge recombination in the muon collider cooling channel

    SciTech Connect

    Fernow, R. C.; Palmer, R. B.

    2012-12-21

    The final stage of the ionization cooling channel for the muon collider must transversely recombine the positively and negatively charged bunches into a single beam before the muons can be accelerated. It is particularly important to minimize any emittance growth in this system since no further cooling takes place before the bunches are collided. We have found that emittance growth could be minimized by using symmetric pairs of bent solenoids and careful matching. We show that a practical design can be found that has transmission {approx}99%, emittance growth less than 0.1%, and minimal dispersion in the recombined bunches.

  19. Mechanism for radiative recombination in ZnCdO alloys

    SciTech Connect

    Buyanova, I. A.; Bergman, J. P.; Pozina, G.; Chen, W. M.; Rawal, S.; Norton, D. P.; Pearton, S. J.; Osinsky, A.; Dong, J. W.

    2007-06-25

    Temperature dependent cw- and time-resolved photoluminescence combined with absorption measurements are employed to evaluate the origin of radiative recombination in ZnCdO alloys grown by molecular-beam epitaxy. The near-band-edge emission is attributed to recombination of excitons localized within band tail states likely caused by nonuniformity in Cd distribution. Energy transfer between the tail states is argued to occur via tunneling of localized excitons. The transfer is shown to be facilitated by increasing Cd content due to a reduction of the exciton binding energy and, therefore, an increase of the exciton Bohr radius in the alloys with a high Cd content.

  20. Recombinant chromosome 18 resulting from a maternal pericentric inversion

    SciTech Connect

    Ayukawa, Hiroshi; Tsukahara, Masato; Fukuda, Masamichi; Kondoh, Osamu

    1994-05-01

    We report on a newborn girl with duplication of 18q12.2{yields}18 qter and deficiency of 18p11.2{yields}18pter which resulted from meiotic recombination of the maternal pericentric inversion, inv(18)(p11.2q12.2). Her clinical manifestations were compatible with those of partial trisomy 18q syndrome. We review the previously reported 9 cases in 8 families of rec(18) resulting from recombination of a parental pericentric inversion. 8 refs., 3 figs., 1 tab.

  1. Prominent Higher-Order Contributions to Electronic Recombination

    SciTech Connect

    Beilmann, C.; Mokler, P. H.; Bernitt, S.; Keitel, C. H.; Ullrich, J.; Lopez-Urrutia, J. R. Crespo; Harman, Z.

    2011-09-30

    Intershell higher-order (HO) electronic recombination is reported for highly charged Ar, Fe, and Kr ions, where simultaneous excitation of one K-shell electron and one or two additional L-shell electrons occurs upon resonant capture of a free electron. For the mid-Z region, HO resonance strengths grow unexpectedly strong with decreasing atomic number Z ({proportional_to}Z{sup -4}), such that, for Ar ions the 2nd-order overwhelms the 1st-order resonant recombination considerably. The experimental findings are confirmed by multiconfiguration Dirac-Fock calculations including hitherto neglected excitation pathways.

  2. Modeling of recombinant yeast cells: reduction of phase space.

    PubMed

    Birol, G; Birol, I; Kirdar, B; Onsan, Z I

    1997-01-01

    The mechanism of starch fermentation by recombinant Saccharomyces cerevisiae in batch reactor is studied. Experiments were carried in the presence and absence of oxygen, with different initial starch concentrations. A variety of data concerning biotic and abiotic phases are collected. Nonlinear data analysis techniques are used to determine the block diagram of the system under study. Data analysis and processing reported here, are believed to form a basis in further work in structured modeling of biological systems, recombinant yeast cultures in particular. PMID:9603032

  3. Enteric Immunization of Mice Against Influenza with Recombinant Vaccinia

    NASA Astrophysics Data System (ADS)

    Meitin, Catherine A.; Bender, Bradley S.; Small, Parker A., Jr.

    1994-11-01

    Intrajejunal administration to mice of a recombinant vaccinia virus containing the influenza virus hemagglutinin gene induced IgA antibody in nasal, gut, and vaginal secretions. It also induced IgG antibody in serum and cell-mediated immunity. The immunization provided significant protection against an influenza virus challenge. This work suggests that enteric-coated recombinant vaccinia could be an orally administered, inexpensive, multivalent, temperature-stable, safe, and effective vaccine for children that could be particularly useful in developing nations, where multiple injections are not easily administered. Oral administration of vaccines should also reduce children's fear of shots at the doctor's office.

  4. Recombinant protein expression in Escherichia coli: advances and challenges

    PubMed Central

    Rosano, Germán L.; Ceccarelli, Eduardo A.

    2014-01-01

    Escherichia coli is one of the organisms of choice for the production of recombinant proteins. Its use as a cell factory is well-established and it has become the most popular expression platform. For this reason, there are many molecular tools and protocols at hand for the high-level production of heterologous proteins, such as a vast catalog of expression plasmids, a great number of engineered strains and many cultivation strategies. We review the different approaches for the synthesis of recombinant proteins in E. coli and discuss recent progress in this ever-growing field. PMID:24860555

  5. A ring-shaped recombination chamber for hadron therapy dosimetry.

    PubMed

    Jakubowska, E; Zielczyński, M; Golnik, N; Gryziński, M A; Krzemiński, Ł

    2014-10-01

    An innovative recombination chamber has been designed for estimation of stray radiation doses and quality factors in hadron therapy. The chamber allows for determination of absorbed dose and recombination index of radiation quality in phantoms at small distances from simulated organs. The chamber body and electrodes are ring shaped, so the beam may be directed through the empty centre of the ring. The ionisation of the filling gas is caused by secondary or scattered radiation and can be related to the dose absorbed in the tissues close to the irradiated target volume. PMID:24430949

  6. Recombination of H(3+) and D(3+) ions with electrons

    NASA Technical Reports Server (NTRS)

    Johnsen, R.; Gougousi, T.; Golde, M. F.

    1994-01-01

    Flowing-afterglow measurements in decaying H3(+) or D3(+) plasmas suggest that de-ionization does not occur by simple binary recombination of a single ion species. We find that vibrational excitation of the ions fails to provide an explanation for the effect, contrary to an earlier suggestion. Instead, we suggest that collisional stabilization of H3** Rydberg molecules by ambient electrons introduces an additional dependence on electron density. The proposed mechanism would permit plasma de-ionization to occur without the need for dissociative recombination by the mechanism of potential-surface crossings.

  7. Acclimation to temperature and irradiance modulates PSII charge recombination.

    PubMed

    Ivanov, A G; Sane, P V; Krol, M; Gray, G R; Balseris, A; Savitch, L V; Oquist, G; Hüner, N P A

    2006-05-15

    Acclimation of wild type and the chlorina F2 mutant of barley to either high light or low temperature results in a 2- to 3-fold increase in non-photochemical quenching which occurred independently of either energy-dependent quenching (qE), xanthophyll cycle-mediated antenna quenching or state transitions. Results of in vivo thermoluminescence measurements used to address this conundrum indicated that excitation pressure regulates the temperature gap for S(2)Q(B)(-) and S(2)Q(A)(-) charge recombinations within photosystem II reaction centers. This is discussed in terms of photoprotection through non-radiative charge recombination. PMID:16674953

  8. Radiative recombination and photon recycling in gallium arsenide solar cells

    NASA Astrophysics Data System (ADS)

    Lundstrom, M. S.; Melloch, M. R.; Lush, G. B.; Patkar, M. P.; Young, M.; Durbin, S. M.; Gray, J. L.; MacMillan, H. F.; Keyes, B. M.; Levi, D. H.; Ahrenkiel, R. K.

    1992-12-01

    This talk reviews experimental work to develop a detailed understanding of radiative recombination in n-GaAs. Photoluminescence decay studies of minority carrier lifetimes versus doping in n-GaAs are presented. We show that when the substrate is removed by etching, photon recycling is enhanced, and lifetimes increase by nearly a factor of 10. The doping-dependent absorption coefficient is measured, and detailed balance arguments are used to relate absorption and recombination. Modeling surfaces, verified by comparison with experiments, are used to examine the effects of recycling in conventional solar cells and to explore new design options.

  9. A systematic analysis of recombination activity and genotype-phenotype correlation in human recombination-activating gene 1 deficiency

    PubMed Central

    Lee, Yu Nee; Frugoni, Francesco; Dobbs, Kerry; Walter, Jolan E.; Giliani, Silvia; Gennery, Andrew R.; Al-Herz, Waleed; Haddad, Elie; LeDeist, Francoise; Bleesing, Jack H.; Henderson, Lauren A.; Pai, Sung-Yun; Nelson, Robert P.; El-Ghoneimy, Dalia H.; El-Feky, Reem A.; Reda, Shereen M.; Hossny, Elham; Soler-Palacin, Pere; Fuleihan, Ramsay L.; Patel, Niraj C.; Massaad, Michel J.; Geha, Raif S.; Puck, Jennifer M.; Palma, Paolo; Cancrini, Caterina; Chen, Karin; Vihinen, Mauno; Alt, Frederick W.; Notarangelo, Luigi D.

    2014-01-01

    Background The recombination-activating gene (RAG) 1/2 proteins play a critical role in the development of T and B cells by initiating the VDJ recombination process that leads to generation of a broad T-cell receptor (TCR) and B-cell receptor repertoire. Pathogenic mutations in the RAG1/2 genes result in various forms of primary immunodeficiency, ranging from T−B− severe combined immune deficiency to delayed-onset disease with granuloma formation, autoimmunity, or both. It is not clear what contributes to such heterogeneity of phenotypes. Objective We sought to investigate the molecular basis for phenotypic diversity presented in patients with various RAG1 mutations. Methods We have developed a flow cytometry–based assay that allows analysis of RAG recombination activity based on green fluorescent protein expression and have assessed the induction of the Ighc locus rearrangements in mouse Rag1−/− pro-B cells reconstituted with wild-type or mutant human RAG1 (hRAG1) using deep sequencing technology. Results Here we demonstrate correlation between defective recombination activity of hRAG1 mutant proteins and severity of the clinical and immunologic phenotype and provide insights on the molecular mechanisms accounting for such phenotypic diversity. Conclusions Using a sensitive assay to measure the RAG1 activity level of 79 mutations in a physiologic setting, we demonstrate correlation between recombination activity of RAG1 mutants and the severity of clinical presentation and show that RAG1 mutants can induce specific abnormalities of the VDJ recombination process. PMID:24290284

  10. RecQ promotes toxic recombination in cells lacking recombination intermediate-removal proteins.

    PubMed

    Magner, Daniel B; Blankschien, Matthew D; Lee, Jennifer A; Pennington, Jeanine M; Lupski, James R; Rosenberg, Susan M

    2007-04-27

    The RecQ-helicase family is widespread, is highly conserved, and includes human orthologs that suppress genomic instability and cancer. In vivo, some RecQ homologs promote reduction of steady-state levels of bimolecular recombination intermediates (BRIs), which block chromosome segregation if not resolved. We find that, in vivo, E. coli RecQ can promote the opposite: the net accumulation of BRIs. We report that cells lacking Ruv and UvrD BRI-resolution and -prevention proteins die and display failed chromosome segregation attributable to accumulation of BRIs. Death and segregation failure require RecA and RecF strand exchange proteins. FISH data show that replication is completed during chromosome-segregation failure/death of ruv uvrD recA(Ts) cells. Surprisingly, RecQ (and RecJ) promotes this death. The data imply that RecQ promotes the net accumulation of BRIs in vivo, indicating a second paradigm for the in vivo effect of RecQ-like proteins. The E. coli RecQ paradigm may provide a useful model for some human RecQ homologs. PMID:17466628

  11. RecQ Promotes Toxic Recombination in Cells Lacking Recombination-Intermediate-Removal Proteins

    PubMed Central

    Magner, Daniel B.; Blankschien, Matthew D.; Lee, Jennifer A.; Pennington, Jeanine M.; Lupski, James R.; Rosenberg, Susan M.

    2010-01-01

    Summary The RecQ-helicase family is widespread, highly conserved, and includes human orthologues that suppress genomic instability and cancer. In vivo, some RecQ homologues promote reduction of steady-state levels of bimolecular recombination intermediates (BRIs), which block chromosome segregation if not resolved. We find that in vivo, E. coli RecQ can promote the opposite: the net accumulation of BRIs. We report that cells lacking Ruv and UvrD BRI-resolution and -prevention proteins die and display failed chromosome segregation attributable to accumulation of BRIs. Death and segregation failure require RecA and RecF strand-exchange proteins. FISH data show that replication is completed during chromosome-segregation failure/death of ruv uvrD recA(Ts) cells. Surprisingly, RecQ (and RecJ) promote this death. The data imply that RecQ promotes the net accumulation of BRIs in vivo, indicating a second paradigm for the in-vivo effect of RecQ-like proteins. The E. coli RecQ paradigm may provide a useful model for some human RecQ homologues. PMID:17466628

  12. High-Resolution Mapping of Homologous Recombination Events in rad3 Hyper-Recombination Mutants in Yeast

    PubMed Central

    Dominska, Margaret; Moriel-Carretero, María; Herrera-Moyano, Emilia; Aguilera, Andrés; Petes, Thomas D.

    2016-01-01

    The Saccharomyces cerevisae RAD3 gene is the homolog of human XPD, an essential gene encoding a DNA helicase of the TFIIH complex involved in both nucleotide excision repair (NER) and transcription. Some mutant alleles of RAD3 (rad3-101 and rad3-102) have partial defects in DNA repair and a strong hyper-recombination (hyper-Rec) phenotype. Previous studies showed that the hyper-Rec phenotype associated with rad3-101 and rad3-102 can be explained as a consequence of persistent single-stranded DNA gaps that are converted to recombinogenic double-strand breaks (DSBs) by replication. The systems previously used to characterize the hyper-Rec phenotype of rad3 strains do not detect the reciprocal products of mitotic recombination. We have further characterized these events using a system in which the reciprocal products of mitotic recombination are recovered. Both rad3-101 and rad3-102 elevate the frequency of reciprocal crossovers about 100-fold. Mapping of these events shows that three-quarters of these crossovers reflect DSBs formed at the same positions in both sister chromatids (double sister-chromatid breaks, DSCBs). The remainder reflects DSBs formed in single chromatids (single chromatid breaks, SCBs). The ratio of DSCBs to SCBs is similar to that observed for spontaneous recombination events in wild-type cells. We mapped 216 unselected genomic alterations throughout the genome including crossovers, gene conversions, deletions, and duplications. We found a significant association between the location of these recombination events and regions with elevated gamma-H2AX. In addition, there was a hotspot for deletions and duplications at the IMA2 and HXT11 genes near the left end of chromosome XV. A comparison of these data with our previous analysis of spontaneous mitotic recombination events suggests that a sub-set of spontaneous events in wild-type cells may be initiated by incomplete NER reactions, and that DSCBs, which cannot be repaired by sister

  13. Recombinational landscape of porcine X chromosome and individual variation in female meiotic recombination associated with haplotypes of Chinese pigs

    PubMed Central

    2010-01-01

    Background Variations in recombination fraction (θ) among chromosomal regions, individuals and families have been observed and have an important impact on quantitative trait loci (QTL) mapping studies. Such variations on porcine chromosome X (SSC-X) and on other mammalian chromosome X are rarely explored. The emerging assembly of pig sequence provides exact physical location of many markers, facilitating the study of a fine-scale recombination landscape of the pig genome by comparing a clone-based physical map to a genetic map. Using large offspring of F1 females from two large-scale resource populations (Large White ♂ × Chinese Meishan ♀, and White Duroc ♂ × Chinese Erhualian ♀), we were able to evaluate the heterogeneity in θ for a specific interval among individual F1 females. Results Alignments between the cytogenetic map, radiation hybrid (RH) map, genetic maps and clone map of SSC-X with the physical map of human chromosome X (HSA-X) are presented. The most likely order of 60 markers on SSC-X is inferred. The average recombination rate across SSC-X is of ~1.27 cM/Mb. However, almost no recombination occurred in a large region of ~31 Mb extending from the centromere to Xq21, whereas in the surrounding regions and in the Xq telomeric region a recombination rate of 2.8-3.3 cM/Mb was observed, more than twice the chromosome-wide average rate. Significant differences in θ among F1 females within each population were observed for several chromosomal intervals. The largest variation was observed in both populations in the interval UMNP71-SW1943, or more precisely in the subinterval UMNP891-UMNP93. The individual variation in θ over this subinterval was found associated with F1 females' maternal haplotypes (Chinese pig haplotypes) and independent of paternal haplotype (European pig haplotypes). The θ between UMNP891 and UMNP93 for haplotype 1122 and 4311 differed by more than fourteen-fold (10.3% vs. 0.7%). Conclusions This study reveals marked

  14. Primordial helium recombination. II. Two-photon processes

    SciTech Connect

    Hirata, Christopher M.; Switzer, Eric R.

    2008-04-15

    Interpretation of precision measurements of the cosmic microwave background (CMB) will require a detailed understanding of the recombination era, which determines such quantities as the acoustic oscillation scale and the Silk damping scale. This paper is the second in a series devoted to the subject of helium recombination, with a focus on two-photon processes in He i. The standard treatment of these processes includes only the spontaneous two-photon decay from the 2{sup 1}S level. We extend this treatment by including five additional effects, some of which have been suggested in recent papers but whose impact on He i recombination has not been fully quantified. These are: (i) stimulated two-photon decays; (ii) two-photon absorption of redshifted He i line radiation; (iii) two-photon decays from highly excited levels in He i (n{sup 1}S and n{sup 1}D, with n{>=}3); (iv) Raman scattering; and (v) the finite width of the 2{sup 1}P{sup o} resonance. We find that effect (iii) is highly suppressed when one takes into account destructive interference between different intermediate states contributing to the two-photon decay amplitude. Overall, these effects are found to be insignificant: they modify the recombination history at the level of several parts in 10{sup 4}.

  15. From gateway to multisite gateway in one recombination event

    Technology Transfer Automated Retrieval System (TEKTRAN)

    We created a construct, pDONR-R4-R3, to easily convert any available Gateway destination vector to a MultiSite Gateway vector in a single recombination reaction. In addition, we designed pDONR-R4-R3 so that DNA fragments already cloned upstream or downstream of the Gateway cassette in the original d...

  16. Phylogenomic evidence for recombination of adenoviruses in wild gorillas.

    PubMed

    Hoppe, Eileen; Pauly, Maude; Robbins, Martha; Gray, Maryke; Kujirakwinja, Deo; Nishuli, Radar; Boji Mungu-Akonkwa, Dieu-Donné; Leendertz, Fabian H; Ehlers, Bernhard

    2015-10-01

    Human adenoviruses (HAdVs) of species Human mastadenovirus B (HAdV-B) are genetically highly diverse and comprise several pathogenic types. AdVs closely related to members of HAdV-B infect African great apes and the evolutionary origin of HAdV-B has recently been determined in ancient gorillas. Genetic evidence for intra- and inter-species recombination has been obtained for AdVs of humans and captive great apes, but evidence from wild great apes is lacking. In this study, potential HAdV-B members of wild Eastern gorillas were analysed for evidence of recombination. One near-complete genome was amplified from primary sample material and sequenced, and from another six individuals genome fragments were obtained. In phylogenomic analysis, their penton base, pVII-pVI, hexon and fiber genes were compared with those of all publicly available HAdV-B full-genome sequences of humans and captive great apes. Evidence for intra-species recombination between different HAdV-B members of wild gorillas as well as between HAdV-B members of chimpanzees and gorillas was obtained. Since zoonotic AdVs have been reported to cause respiratory outbreaks in both humans and monkeys, and humans in West and Central Africa frequently hunt and butcher primates thereby increasing the chance of zoonotic transmission, such HAdV-B recombinants might widen the pool of potential human pathogens. PMID:26219820

  17. Spin-dependent recombination probed through the dielectric polarizability

    PubMed Central

    Bayliss, Sam L.; Greenham, Neil C.; Friend, Richard H.; Bouchiat, Hélène; Chepelianskii, Alexei D

    2015-01-01

    Despite residing in an energetically and structurally disordered landscape, the spin degree of freedom remains a robust quantity in organic semiconductor materials due to the weak coupling of spin and orbital states. This enforces spin-selectivity in recombination processes which plays a crucial role in optoelectronic devices, for example, in the spin-dependent recombination of weakly bound electron-hole pairs, or charge-transfer states, which form in a photovoltaic blend. Here, we implement a detection scheme to probe the spin-selective recombination of these states through changes in their dielectric polarizability under magnetic resonance. Using this technique, we access a regime in which the usual mixing of spin-singlet and spin-triplet states due to hyperfine fields is suppressed by microwave driving. We present a quantitative model for this behaviour which allows us to estimate the spin-dependent recombination rate, and draw parallels with the Majorana–Brossel resonances observed in atomic physics experiments. PMID:26439933

  18. Doping in the recombinant era: strategies and counterstrategies.

    PubMed

    Azzazy, Hassan M E; Mansour, Mai M H; Christenson, Robert H

    2005-11-01

    Advances in recombinant DNA technology have created one of the most powerful weapons in the current doping arsenal: recombinant proteins [Sweeney HL. Gene doping. Sci Am 2004;291:62-9; Unal M, Ozer Unal D. Gene doping in sports. Sports Med 2004;34:357-62]. Recombinant erythropoietin (EPO) and human growth hormone (hGH) are currently being abused but are fortunately detectable either directly by employing isoelectric focusing and immunoassays or indirectly by assessing changes in selected hematopoietic parameters. The detection is technically demanding due to the extent of similarity between the recombinant proteins and their endogenous counterparts. Another issue facing detection efforts is the speed and conditions at which blood samples are collected and analyzed in a sports setting. Recently, gene doping, which stemmed out of legitimate gene therapy trials, has emerged as the next level of doping. Erythropoietin (EPO), human growth hormone (hGH), insulin-like growth factor-1 (IGF-1), peroxisome proliferator-activated receptor-delta (PPAR delta), and myostatin inhibitor genes have been identified as primary targets for doping. Sports clinical scientists today are racing against the clock because assuring the continued integrity of sports competition depends on their ability to outpace the efforts of dopers by developing new detection strategies. PMID:16286094

  19. Detecting Recombination Hotspots from Patterns of Linkage Disequilibrium

    PubMed Central

    Wall, Jeffrey D.; Stevison, Laurie S.

    2016-01-01

    With recent advances in DNA sequencing technologies, it has become increasingly easy to use whole-genome sequencing of unrelated individuals to assay patterns of linkage disequilibrium (LD) across the genome. One type of analysis that is commonly performed is to estimate local recombination rates and identify recombination hotspots from patterns of LD. One method for detecting recombination hotspots, LDhot, has been used in a handful of species to further our understanding of the basic biology of recombination. For the most part, the effectiveness of this method (e.g., power and false positive rate) is unknown. In this study, we run extensive simulations to compare the effectiveness of three different implementations of LDhot. We find large differences in the power and false positive rates of these different approaches, as well as a strong sensitivity to the window size used (with smaller window sizes leading to more accurate estimation of hotspot locations). We also compared our LDhot simulation results with comparable simulation results obtained from a Bayesian maximum-likelihood approach for identifying hotspots. Surprisingly, we found that the latter computationally intensive approach had substantially lower power over the parameter values considered in our simulations. PMID:27226166

  20. Meiotic Segregation and Male Recombination in Interspecific Hybrids of Drosophila

    PubMed Central

    Coyne, Jerry A.

    1986-01-01

    Male hybrids between three pairs of Drosophila species show no substantial distortion of Mendelian segregation and no appreciable male recombination. These results do not support the theories that meiotic drive alleles of large effect are often fixed within species and that transposable genetic elements cause speciation. PMID:3021573