Academic literature on the topic 'Recombinant lines'
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Journal articles on the topic "Recombinant lines"
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Sherman, J. D., L. Y. Smith, T. K. Blake, and L. E. Talbert. "Identification of barley genome segments introgressed into wheat using PCR markers." Genome 44, no. 1 (February 1, 2001): 38–44. http://dx.doi.org/10.1139/g00-092.
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Barley has several important traits that might be used in the genetic improvement of wheat. For this report, we have produced wheat-barley recombinants involving barley chromosomes 4 (4H) and 7 (5H). Wheat-barley disomic addition lines were crossed with 'Chinese Spring' wheat carrying the ph1b mutation to promote homoeologous pairing. Selection was performed using polymerase chain reaction (PCR) markers to identify lines with the barley chromosome in the ph1b background. These lines were self pollinated, and recombinants were identified using sequence-tagged-site (STS) primer sets that allowed differentiation between barley and wheat chromosomes. Several recombinant lines were isolated that involved different STS-PCR markers. Recombination was confirmed by allowing the lines to self pollinate and rescreening the progeny via STS-PCR. Progeny testing confirmed 9 recombinants involving barley chromosome 4 (4H) and 11 recombinants involving barley chromosome 7 (5H). Some recombinants were observed cytologically to eliminate the possibility of broken chromosomes. Since transmission of the recombinant chromosomes was lower than expected and since seed set was reduced in recombinant lines, the utility of producing recombinants with this method is uncertain.Key words: introgression, sequence-tagged-site, recombination.
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Biagetti, Marco, Francesca Vitellozzi, and Carla Ceoloni. "Physical mapping of wheat-Aegilops longissima breakpoints in mildew-resistant recombinant lines using FISH with highly repeated and low-copy DNA probes." Genome 42, no. 5 (October 1, 1999): 1013–19. http://dx.doi.org/10.1139/g98-172.
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Fluorescence in situ hybridization (FISH) with multiple probes, consisting of highly repeated DNA sequences (pSc119.2 and pAs1) and of a low-copy, 3BS-specific RFLP sequence (PSR907), enabled determination of the physical position of the wheat-alien breakpoints (BPs) along the 3BS and 3DS arms of common wheat recombinant lines. These lines harbour 3SlS Aegilops longissima segments containing the powdery mildew resistance gene Pm13. In all 3B recombinants, the wheat-Aegilops longissima physical BPs lie within the interval separating the two most distal of the three pSc119.2 3BS sites. In all such recombinants a telomeric segment, containing the most distal of the pSc119.2 3BS sites, was in fact replaced by a homoeologous Ae. longissima segment, marked by characteristic pSc119.2 hybridization sites. Employment of the PSR907 RFLP probe as a FISH marker allowed to resolve further the critical region in the various 3B recombinant lines. Three of them, like the control common wheat, exhibited between the two most distal pSc119.2 sites a single PSR907 FISH site, which was missing in a fourth recombinant line. The amount of alien chromatin can thus be estimated to represent around 20% of the recombinant arm in the three former lines and a maximum of 27% in the latter. A similar physical length was calculated for the alien segment contained in three 3D recombinants, all characterized by the presence of the Ae. longissima pSc119.2 sites distal to the nearly telomeric pAs1 sites of normal 3DS. Comparison between the FISH-based maps and previously developed RFLP maps of the 3BS-3SlS and 3DS-3SlS arms revealed substantial differences between physical and genetic map positions of the wheat-alien BPs and of molecular markers associated with the critical chromosomal portions.Key words: wheat-alien recombinants, chromosome engineering, fluorescence in situ hybridization, highly repeated and low-copy DNA probes, physical versus genetic maps.
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Broman, Karl W. "The Genomes of Recombinant Inbred Lines." Genetics 169, no. 2 (November 15, 2004): 1133–46. http://dx.doi.org/10.1534/genetics.104.035212.
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Broman, K. W. "The Genomes of Recombinant Inbred Lines." Genetics 173, no. 4 (August 1, 2006): 2419. http://dx.doi.org/10.1093/genetics/173.4.2419.
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Dzianott, Aleksandra, Joanna Sztuba-Solińska, and Jozef J. Bujarski. "Mutations in the Antiviral RNAi Defense Pathway Modify Brome mosaic virus RNA Recombinant Profiles." Molecular Plant-Microbe Interactions® 25, no. 1 (January 2012): 97–106. http://dx.doi.org/10.1094/mpmi-05-11-0137.
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RNA interference (RNAi) mechanism targets viral RNA for degradation. To test whether RNAi gene products contributed to viral RNA recombination, a series of Arabidopsis thaliana RNAi-defective mutants were infected with Brome mosaic virus (BMV) RNAs that have been engineered to support crossovers within the RNA3 segment. Single-cross RNA3-RNA1, RNA3-RNA2, and RNA3-RNA3 recombinants accumulated in both the wild-type (wt) and all knock-out lines at comparable frequencies. However, a reduced accumulation of novel 3′ mosaic RNA3 recombinants was observed in ago1, dcl2, dcl4, and rdr6 lines but not in wt Col-0 or the dcl3 line. A BMV replicase mutant accumulated a low level of RNA3-RNA1 single-cross recombinants in Col-0 plants while, in a dcl2 dcl4 double mutant, the formation of both RNA3-RNA1 and mosaic recombinants was at a low level. A control infection in the cpr5-2 mutant, a more susceptible BMV Arabidopsis host, generated similar-to-Col-0 profiles of both single-cross and mosaic recombinants, indicating that recombinant profiles were, to some extent, independent of a viral replication rate. Also, the relative growth experiments revealed similar selection pressure for recombinants among the host lines. Thus, the altered recombinant RNA profiles have originated at the level of recombinant formation rather than because of altered selection. In conclusion, the viral replicase and the host RNAi gene products contribute in distinct ways to BMV RNA recombination. Our studies reveal that the antiviral RNAi mechanisms are utilized by plant RNA viruses to increase their variability, reminiscent of phenomena previously demonstrated in fungi.
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Taketa, Shin, Takaya Awayama, Masahiko Ichii, Makoto Sunakawa, Tomoko Kawahara, and Koji Murai. "Molecular cytogenetic identification of nullisomy 5B induced homoeologous recombination between wheat chromosome 5D and barley chromosome 5H." Genome 48, no. 1 (February 1, 2005): 115–24. http://dx.doi.org/10.1139/g04-096.
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Chromosome 5H of Hordeum vulgare 'New Golden' (NG) carries a gene(s) that accelerates heading in a wheat background. To introduce the early heading gene(s) of NG barley into the wheat genome, we attempted to induce homoeologous recombination between wheat and NG 5H chromosomes by 5B nullisomy. A nullisomic 5B, trisomic 5A, monosomic 5H plant (2n = 42) was produced from systematic crosses between aneuploid stocks of wheat group 5 chromosomes. A total of 656 F2 plants produced by self-fertilization were screened for recombinants by a PCR assay with 3 5H-specific amplicon markers. Twelve plants (1.8%) were selected as putative wheat–barley 5H recombinants. Five of them were inviable or sterile and the remaining 7 were fertile and subjected to the progeny test. Cytological analyses using fluorescence in situ hybridization and C-banding revealed that 6 of the 7 progeny lines are true homoeologous recombinants between the long arms of chromosomes 5D and 5H, but that the other one was not a recombinant having an aberrant barley telosome. The 6 cytologically confirmed recombinant lines included only 2 types (3 lines each), which were reciprocal products derived from exchanges at the same distal interval defined by two flanking markers. One type had a small 5HL segment translocated to the 5DL terminal, and the other type had a small terminal 5DL segment translocated to the 5HL terminal. In the latter type, the physical length of translocated barley segments slightly differed among lines. Homoeologous recombinants obtained in this study should be useful for further chromosome manipulation to introgress a small interstitial 5HL chromosome segment with the early heading gene(s) to wheat. Preferential occurrence of restricted types of recombinants is discussed in relation to homoeologous relationships between wheat and barley chromosomes.Key words: genomic in situ hybridization, homoeologous pairing, Hordeum vulgare, introgression, recombinant, Triticum aestivum.
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IPSILANDIS, C. G., and M. KOUTSIKA-SOTIRIOU. "The combining ability of recombinant S-lines developed from an F2 maize population." Journal of Agricultural Science 134, no. 2 (March 2000): 191–98. http://dx.doi.org/10.1017/s0021859699007406.
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Starting with the F2 generation of the single-cross commercial hybrid Lorena (PR3183), recombinant lines were developed combining half-sib/S1 evaluation on widely spaced plants in the direction of high yielding per se. Combining ability tests consisted of crosses between: (a) recombinant lines of common pedigree and (b) recombinant lines and freely available inbred lines. The highest-yielding crosses between recombinant lines reached 100% of the original F1 hybrid in a percentage of 14·2. Low heterosis was estimated owing to additive gene action of recombinant lines. Crosses between recombinant lines and freely available inbred lines outyielded significantly the commercial F1 hybrid in a percentage of 33·3. Heterosis was greater and the original F1 hybrid was outyielded significantly because of non-additive gene action. When the applied breeding procedure on a source population with high yield adaptability is adopted and where effects of intergenotypic competition masking the inherent genotypic value are controlled, population improvement may be substituted by combined half-sib/S1 selection for productivity of lines per se in low stress conditions during the very early stages.
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Mahadevappa, Mamatha, Richard A. DeScenzo, and Roger P. Wise. "Recombination of alleles conferring specific resistance to powdery mildew at the Mla locus in barley." Genome 37, no. 3 (June 1, 1994): 460–68. http://dx.doi.org/10.1139/g94-064.
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In barley (Hordeum vulgare L.), the Mla locus conditions reaction to the powdery mildew fungus Erysiphe graminis f.sp. hordei. Enrichment for genetic recombinants in the Mla region is possible by screening for recombination events between the flanking endosperm storage proteins hordeins C and B. Reciprocal crosses were made between the Franger (C.I. 16151) and Rupee (C.I. 16155) lines carrying the (Mla6 + Mla14) and Mla13 alleles, respectively. Recombinants were identified from F2 segregants by analyzing the extracted hordein polypeptides by sodium dodecyl sulphate – polyacrylamide gel electrophoresis. Two hundred and seventy-six recombinant gametes were identified from the 1800 seeds that were screened. Recombination of Mla alleles was analyzed by inoculating F4 recombinant lines with three isolates of E. graminis (A27, 5874, and CR3), which recognize specific Mla alleles. The linkage order established is Hor1–Mla6–Mla13–Mla14–Hor2. The genetic distances between Hor1–Mla6, Mla6–Mla13, and Mla13–Hor2, obtained using Mapmaker 3.0b F3 intercross analysis, are 3.9, 0.2, and 5.2 cM, respectively.Key words: recombinant, barley, powdery mildew, Mla, hordein.
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Kozub, N. A., I. A. Sozinov, A. Ya Bidnyk, N. A. Demianova, Ya B. Blume, and A. A. Sozinov. "Development of common wheat lines with the recombinant arm 1RS as a source of new combinations of disease and pest resistance genes." Interdepartmental Thematic Scientific Collection of Plant Protection and Quarantine, no. 62 (September 3, 2016): 143–50. http://dx.doi.org/10.36495/1606-9773.2016.62.143-150.
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A combination of recombinant-inbred lines of the F6 generation from the cross B-16 ќ AR 7086 between lines with two wheat-rye translocations, 1BL/1RS from the Petkus and 1AL/1RS from the rye Insave, was developed. Using gliadin and secalin loci as genetic markers we identified recombinant arm 1RS in positions 1A and 1B in about 10% of lines. The rest of lines with the rye material may also carry recombinant 1RS, which can be identified with DNA markers. Lines with recombinant arm 1RS may serve as a source of new combination of rye genes for disease and pest resistance.
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Crow, James F. "Haldane, Bailey, Taylor and Recombinant-Inbred Lines." Genetics 176, no. 2 (June 1, 2007): 729–32. http://dx.doi.org/10.1093/genetics/176.2.729.
Full textDissertations / Theses on the topic "Recombinant lines"
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Anderson, Amy D. "The genetic structure of related recombinant lines /." Thesis, Connect to this title online; UW restricted, 2003. http://hdl.handle.net/1773/8935.
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Kim, Sumee M. "Expression of a recombinant NMDA R1 cDNA in mammalian cell lines." Thesis, National Library of Canada = Bibliothèque nationale du Canada, 1997. http://www.collectionscanada.ca/obj/s4/f2/dsk2/ftp04/mq21068.pdf.
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Keith, Michelle Barbara Ann. "Screening of stably transformed insect cell lines for recombinant protein production." Thesis, National Library of Canada = Bibliothèque nationale du Canada, 1999. http://www.collectionscanada.ca/obj/s4/f2/dsk2/ftp01/MQ38634.pdf.
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Castilho, Alexandra Marina Machado Ferreira. "Molecular cytogenic analysis of recombinant chromosomes in wheat - Aegilops umbellulata lines." Thesis, University of East Anglia, 1995. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.296341.
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Guess, Adam Joseph. "QTL analysis of ray pattern in Caenorhabditis elegans recombinant inbred lines." Wright State University / OhioLINK, 2008. http://rave.ohiolink.edu/etdc/view?acc_num=wright1205197070.
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Large, Charles Henry. "Characterisation of dopamine D3 receptors in recombinant cell lines and rat brain." Thesis, University of Bristol, 1994. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.388037.
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Bello, Halima Thelma. "Phenotypic and genotypic evaluation of generations and recombinant inbred lines for response to aflatoxin." [College Station, Tex. : Texas A&M University, 2007. http://hdl.handle.net/1969.1/ETD-TAMU-1359.
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Steinmetz, Ralf Dirk. "Functional expression of recombinant N-methyl-D-aspartate (NMDA) receptors in eukaryotic cell lines." [S.l.] : [s.n.], 2000. http://deposit.ddb.de/cgi-bin/dokserv?idn=961238070.
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Patokar, Chetan. "Molecular cytogenetics and genomics of novel wheat-Thinopyrum bessarabicum recombinant lines carrying intercalary translocations." Thesis, University of Leicester, 2016. http://hdl.handle.net/2381/36195.
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The diploid wild grass Thinopyrum bessarabicum (2n = 2x = 14, JJ or EbEb) is a rich source of important genes for bread wheat (2n = 6x = 42) improvement because of its salinity tolerance and disease resistance. Development of wheat–Th. bessarabicum translocation lines by backcrossing amphiploids in the absence of the Ph1 gene (allowing intergenomic recombination) enables its practical utilization in wheat improvement. Using genomic in situ hybridization (GISH) and repetitive probes for fluorescent in situ hybridization (FISH), six novel wheat–Th. bessarabicum translocation lines involving different chromosome segments (T4BS.4BL-4JL, T6BS.6BL-6JL, T5AS.5AL-5JL, T5DL.5DS-5JS, T2BS.2BL-2JL, and the whole arm translocation T1AL.1JS) were identified and characterized in this study. No background translocations between wheat genomes were observed. The involvement of 5 of the 7 chromosomes, and small terminal segments of the Th. bessarabicum chromosome arm were important, contributing to both reduced linkage drag of the derived lines by minimizing agronomically deleterious genes from the alien species, and high stability including transmission of the alien segment. All three wheat genomes were involved in the translocations with the alien chromosome, and GISH showed the Th. bessarabicum genome was more closely related to the D genome in wheat. All the introgression lines were disomic, stable and with good morphological characters. The work also generated a high-resolution karyotype of two accessions of Th. bessarabicum using multiple repetitive DNA probes for chromosome identification. A complete CS-Th. bessarabicum amphiploid (2n=8x=56, AABBDDJJ) was used and each individual Jgenome unambiguously identified. The established karyotype will be useful for the rapid identification of potential donor chromosomes in wheat improvement programs, allowing appropriate alien-chromosome transfer. Genotyping-by-sequencing (GBS) data was collected from the wheat-Th. bessarabicum introgression lines, but the complexity of the wheat genome and need for further development of data analysis pathways limited interpretation.
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Voulgaropoulou, Frosso. "Construction and characterization of stable cell lines that generate recombinant adeno-associated virus (rAAV) /." The Ohio State University, 1996. http://rave.ohiolink.edu/etdc/view?acc_num=osu1487942476408687.
Full textBooks on the topic "Recombinant lines"
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Clark, Joanna Mary. The regulation of human phospholipase D: Studies with recombinant phospholipase D1b and myeloid leukaemic cell lines. Birmingham: University of Birmingham, 1999.
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Pines, Maya. The new human genetics: How gene splicing helps researchers fight inherited disease. [Bethesda, Md.?]: U.S. Dept. of Health and Human Services, Public Health Service, National Institutes of Health, National Institute of General Medical Sciences, 1985.
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Stephens, Mark Lee. The effect of periodic operation on mixed and recombinant bacterial populations. 1989.
Find full textBook chapters on the topic "Recombinant lines"
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Priyadarshan, P. M. "Recombinant Inbred Lines." In PLANT BREEDING: Classical to Modern, 257–68. Singapore: Springer Singapore, 2019. http://dx.doi.org/10.1007/978-981-13-7095-3_13.
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Blaas, Leander, Monica Musteanu, Beatrice Grabner, Robert Eferl, Anton Bauer, and Emilio Casanova. "The Use of Bacterial Artificial Chromosomes for Recombinant Protein Production in Mammalian Cell Lines." In Recombinant Gene Expression, 581–93. Totowa, NJ: Humana Press, 2011. http://dx.doi.org/10.1007/978-1-61779-433-9_31.
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Fischbach, Michel, and Peter Bromley. "Recombinant Cell Lines for Stress Reporter Assays." In Cell Culture Methods for In Vitro Toxicology, 131–47. Dordrecht: Springer Netherlands, 2001. http://dx.doi.org/10.1007/978-94-017-0996-5_9.
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Swiech, Kamilla, Marcela Cristina Corrêa de Freitas, Dimas Tadeu Covas, and Virgínia Picanço-Castro. "Recombinant Glycoprotein Production in Human Cell Lines." In Methods in Molecular Biology, 223–40. New York, NY: Springer New York, 2014. http://dx.doi.org/10.1007/978-1-4939-2205-5_12.
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Pollard, Daniel A. "Design and Construction of Recombinant Inbred Lines." In Methods in Molecular Biology, 31–39. Totowa, NJ: Humana Press, 2012. http://dx.doi.org/10.1007/978-1-61779-785-9_3.
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De Neve, Myriam, Helena Van Houdt, Anne-Marie Bruyns, Marc Van Montagu, and Ann Depicker. "Screening for Transgenic Lines with Stable and Suitable Accumulation Levels of a Heterologous Protein." In Recombinant Proteins from Plants, 203–27. Totowa, NJ: Humana Press, 1998. http://dx.doi.org/10.1007/978-1-60327-260-5_16.
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Spiegel, Holger, Stefan Schillberg, and Greta Nölke. "Production of Recombinant Proteins by Agrobacterium-Mediated Transient Expression." In Recombinant Proteins in Plants, 89–102. New York, NY: Springer US, 2022. http://dx.doi.org/10.1007/978-1-0716-2241-4_6.
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AbstractThe agroinfiltration of plant tissue is a robust method that allows the rapid and transient expression of recombinant proteins. Using wild-type plants as biomass, agroinfiltration exploits the ability of plants to synthesize even complex multimeric proteins that require oxidative folding and/or post-translational modifications, while avoiding the expensive and time-consuming creation of stably transformed plant lines. Here we describe a generic method for the transient expression of recombinant proteins in Nicotiana benthamiana at the small to medium laboratory scale, including appropriate binary vectors, the design and cloning of expression constructs, the transformation, selection, and cultivation of recombinant Agrobacterium tumefaciens, the infiltration of plants using a syringe or vacuum device, and finally the extraction of recombinant proteins from plant tissues.
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Steiner, Florian, Sraboni Ghose, and Urs Thomet. "Recombinant Cell Lines Stably Expressing Functional Ion Channels." In Methods in Molecular Biology, 209–21. Totowa, NJ: Humana Press, 2010. http://dx.doi.org/10.1007/978-1-60327-323-7_17.
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Reeves, Philip J. "Construction of Recombinant Cell Lines for GPCR Expression." In Methods in Molecular Biology, 43–60. New York, NY: Springer US, 2021. http://dx.doi.org/10.1007/978-1-0716-1221-7_3.
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Nakamichi, Noboru, Mamiko Ito, Yoshiko Hirota, and Toshiharu Matsumura. "Establishment of Recombinant Human Transferrin-Producing CHO Cell Lines." In Animal Cell Technology: Basic & Applied Aspects, 373–79. Dordrecht: Springer Netherlands, 1997. http://dx.doi.org/10.1007/978-94-011-5746-9_60.
Full textConference papers on the topic "Recombinant lines"
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"Molecular genetic analysis of alloplasmic recombinant lines (Triticum dicoccum) – Triticum aestivum." In Bioinformatics of Genome Regulation and Structure/ Systems Biology. institute of cytology and genetics siberian branch of the russian academy of science, Novosibirsk State University, 2020. http://dx.doi.org/10.18699/bgrs/sb-2020-218.
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Hanson, L. E., R. M. Beaudry, T. R. Goodwill, and J. M. McGrath. "RESPONSE OF SUGAR BEET RECOMBINANT INBRED LINES TO POST-HARVEST ROT FUNGI." In 37th Biennial Meeting of American Society of Sugarbeet Technologist. ASSBT, 2013. http://dx.doi.org/10.5274/assbt.2013.26.
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Iancu, Paula Iancu. "QUALITY CHARACTERISTICS OF WHOLE GRAINS FLOUR OF SOME MUTANT/RECOMBINANT WINTER WHEAT DH LINES." In 18th International Multidisciplinary Scientific GeoConference SGEM2018. Stef92 Technology, 2018. http://dx.doi.org/10.5593/sgem2018/6.2/s25.065.
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McGrath, J. Mitchell, Teresa K. Koppin, and Tim M. Duckert. "Breeding for genetics: development of Recombinant Inbred Lines (RIL's) for gene discovery and deployment." In 33rd Biennial Meeting of American Society of Sugarbeet Technologist. ASSBT, 2005. http://dx.doi.org/10.5274/assbt.2005.28.
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Волынкин, В. А., С. М. Гориславец, В. А. Володин, И. А. Васылык, Е. А. Лущай, В. В. Лиховской, and Е. К. Потокина. "PHYTOPATHOLOGICAL SPECIFICITY OF RESISTANCE OF RECOMBINANT GRAPE LINES TO ERISIPHE NECATOR AND PLASMOPARA VITICOLA." In Материалы I Всероссийской научно-практической конференции с международным участием «Геномика и современные биотехнологии в размножении, селекции и сохранении растений». Crossref, 2020. http://dx.doi.org/10.47882/genbio.2020.85.48.064.
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Nain, Ashish, Saraswathipura L. Krishnamurthy, Parbodh C. Sharma, Bayragondlu M. Lokeshkumar, Mukesh Kumar, and Arvinder S. Warraich. "Phenotypic Evaluation of Recombinant Inbred Lines for Sodicity Tolerance at Reproductive Stage in Rice." In LAFOBA2. Basel Switzerland: MDPI, 2022. http://dx.doi.org/10.3390/environsciproc2022016047.
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Zettlmeiβl, G., H. Ragg, and H. Karges. "EXPRESSION OF BIOLOGICALLY ACTIVE HUMAN ANTITHROMBIN III IN CHINESE HAMSTER OVARY CELLS." In XIth International Congress on Thrombosis and Haemostasis. Schattauer GmbH, 1987. http://dx.doi.org/10.1055/s-0038-1643683.
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Expression of human antithrombin III (AT III) at high levels has been achieved in Chinese hamster ovary (CH0) cells by cotransfection and subsequent coamplification of the transfected sequences. Expression vectors containing the AT III cDNA gene and a dihydrofolate reductase (DHFR) cDNA gene were transfected into CH0 DHFR-deficient cells. About 20% of the DHFR+ transformants secreted recombinant human AT III into the medium. Stepwise selection of the AT III producing DHFR+ -transformants in increasing concentrations of methotrexate generated cells which had amplified the AT III géne. We determined the copy number of the AT III cDNA and the relative amounts of AT III specific mRNA at different stages of the amplification process. Transfected CH0 cell lines expressed elevated immunreactive levels of human AT III.AT III secreted from these cell lines had the same molecular weight (60 kDa), immunological properties and biological activities as AT III obtained from human plasma. In vivo data concerning the inhibition of glycosylation by different drugs suggest that recombinant AT III from CHO cells is glycosylated according to a complex type pattern.
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Vehar, G. A. "THE PRESENT STATE OF GENE TECHNOLOGY IN THE MANUFACTURE OF HUMAN COAGULATION PROTEINS." In XIth International Congress on Thrombosis and Haemostasis. Schattauer GmbH, 1987. http://dx.doi.org/10.1055/s-0038-1644755.
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The production of pharmaceuticals from human plasma that are useful in the treatment of bleeding disorders had its beginning with the development of the Cohn fractionation procedure in the 1940's. As a result of these advances, concentrates became available for the treatment of the hemophilias. Although of low purity and subject to contamination by hepatitis virus, the availability of these compounds resulted in dramatic improvements in the life expectancy and quality of life of afflicted individuals. The numerous problems associated with production of pharmaceuticals from pooled plasma made these products obvious goals for recombinant DNA technology as soon as the commercial aspects of the field became apparent. The subsequent contamination of blood products with the AIDS virus has resulted in an urgent need for a production source that is independent of human plasma. Several industrial and academic laboratories have cloned the cDNA's for human factors VIII and IX. In addition to these proteins, the utility of factor Vila in the treatment of hemophiliacs with inhibitors has shown promise. Efforts to develop a recombinant preparation of factor Vila are at a comparable stage of development as factors VIII and IX. Continuing efforts have resulted in the successful expression of these recombinant proteins in mammalian cell lines, thereby successfully completing the first steps of commercial development.Although much interest has focused upon the theoretical superiority of recombinant proteins as therapeutics, one must keep in mind that there are numerous developmental aspects of large-scale production and regulatory issues that must be addressed and solved before these drugs will be available. The coagulation proteins are complex glycoproteins that will in all probability require mammalian cell cell culture in order to produce functional proteins. The fact that these preparations will be administered over the lifetime of the patient serves to reinforce that the recombinant products be as similar to the natural proteins as possible, further supporting the concept of mammalian cell expression systems.Regulatory approval of a recombinant product are fundamentally no different than those for any other product in regards to efficacy, potency, purity, and identity. There are, however, additional considerations that must be addressed in the production of recombinant cell culture derived biologies. These relate to the possible presence in the final product of pathogenic and tumorigenic agents, and possible contamination by cell culture and cell substrate compounds. A detailed characterization of the production cell line will therefore be required, including identification and characterization of any associated viral particles. These cells must be capable of being reproducibly grown, while maintaining protein production, on ascale (tens of thousands of liters) suitable to meet the market demand of the specific protein. Apurification process must be established capable of handling the resulting large volumes of feedstock, generating a protein preparation of high purity (greater than 99% pure). Numerous assays must be developed to quantitate the purity and identity of the resulting recombinant pharmaceutical on a lot by lot basis.Studies to date have shown that recombinant forms of factors VIII and IX, produced by laboratory processes, are very similar to the plasma-derived forms as assessed by a variety of in vitro and in vivo tests. Although these results are promising, the ultimate safety and efficacy testing of these drugs will have to await the initiation of human clinical trials. Such studies will have to await the successful completion of the certain regulatory concerns. Clinical trials should begin within the near future, hopefully leading to a source of these products independent of pooled human plasma.
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Zeng, Ya-Wen, Lu-Xiang Wang, Juan Du, Xiao-Meng Yang, Xiao-Ying Pu, Li-Juan Du, Tao Yang, Jia-Zhen Yang, and Shu-Ming Yang. "Mineral Elements in Barley Grass Powder and its Grains for Recombinant Inbred Lines by ICP-AES." In 2015 International Conference on Medicine and Biopharmaceutical. WORLD SCIENTIFIC, 2016. http://dx.doi.org/10.1142/9789814719810_0151.
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"Recombinant lactaptin analogon RL2 inhibits TRAIL-induced cell death in breast cancer cell lines by mitophagy." In Bioinformatics of Genome Regulation and Structure/Systems Biology (BGRS/SB-2022) :. Institute of Cytology and Genetics, the Siberian Branch of the Russian Academy of Sciences, 2022. http://dx.doi.org/10.18699/sbb-2022-608.
Full textReports on the topic "Recombinant lines"
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Bennett, Alan B., Arthur A. Schaffer, Ilan Levin, Marina Petreikov, and Adi Doron-Faigenboim. Manipulating fruit chloroplasts as a strategy to improve fruit quality. United States Department of Agriculture, January 2013. http://dx.doi.org/10.32747/2013.7598148.bard.
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The Original Objectives were modified and two were eliminated to reflect the experimental results: Objective 1 - Identify additional genetic variability in SlGLK2 and IPin wild, traditional and heirloom tomato varieties Objective 2 - Determine carbon balance and horticultural characteristics of isogenic lines expressing functional and non-functional alleles of GLKsand IP Background: The goal of the research was to understand the unique aspects of chloroplasts and photosynthesis in green fruit and the consequences of increasing the chloroplast capacity of green fruit for ripe fruit sugars, yield, flavor and nutrient qualities. By focusing on the regulation of chloroplast formation and development solely in fruit, our integrated knowledge of photosynthetic structures/organs could be broadened and the results of the work could impact the design of manipulations to optimize quality outputs for the agricultural fruit with enhanced sugars, nutrients and flavors. The project was based on the hypothesis that photosynthetic and non-photosynthetic plastid metabolism in green tomato fruit is controlled at a basal level by light for minimal energy requirements but fruit-specific genes regulate further development of robust chloroplasts in this organ. Our BARD project goals were to characterize and quantitate the photosynthesis and chloroplast derived products impacted by expression of a tomato Golden 2- like 2 transcription factor (US activities) in a diverse set of 31 heirloom tomato lines and examine the role of another potential regulator, the product of the Intense Pigment gene (IP activities). Using tomato Golden 2-like 2 and Intense Pigment, which was an undefined locus that leads to enhanced chloroplast development in green fruit, we sought to determine the benefits and costs of extensive chloroplast development in fruit prior to ripening. Major conclusions, solutions, achievements: Single nucleotide polymorphisms in the promoter, coding and intronicSlGLK2 sequences of 20 heirloom tomato lines were identified and three SlGLK2 promoter lineages were identified; two lineages also had striped fruit variants. Lines with striped fruit but no shoulders were not identified. Green fruit chlorophyll and ripe fruit soluble sugar levels were measured in 31 heirloom varieties and fruit size correlates with ripe fruit sugars but dark shoulders does not. A combination of fine mapping, recombinant generation, RNAseq expression and SNP calling all indicated that the proposed localization of a single locus IP on chr 10 was incorrect. Rather, the IP line harbored 11 separate introgressions from the S. chmielewskiparent, scattered throughout the genome. These introgressions harbored ~3% of the wild species genome and no recombinant consistently recovered the IP parental phenotype. The 11 introgressions were dissected into small combinations in segregating recombinant populations. Based on these analyses two QTL for Brix content were identified, accounting for the effect of increased Brix in the IP line. Scientific and agricultural implications: SlGLK2 sequence variation in heirloom tomato varieties has been identified and can be used to breed for differences in SlGLK2 expression and possibly in the green striped fruit phenotype. Two QTL for Brix content have been identified in the S. chmielewskiparental line and these can be used for increasing soluble solids contents in breeding programs.
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Blum, Abraham, and Henry T. Nguyen. Molecular Tagging of Drought Resistance in Wheat: Osmotic Adjustment and Plant Productivity. United States Department of Agriculture, November 2002. http://dx.doi.org/10.32747/2002.7580672.bard.
Full textAbstract:
Drought stress is a major limitation to bread wheat (Triticumaestivum L.) productivity and its yield stability in arid and semi-arid regions of world including parts of Israel and the U.S. Currently, breeding for sustained yields under drought stress is totally dependent on the use of yield and several key physiological attributes as selection indices. The attempt to identify the optimal genotype by evaluating the phenotype is undermining progress in such breeding programs. Osmotic adjustment (OA) is an effective drought resistance mechanism in many crop plants. Evidence exists that there is a genetic variation for OA in wheat and that high OA capacity supports wheat yields under drought stress. The major objective of this research was to identify molecular markers (RFLPs, restriction fragment length polymorphisms; and AFLPs, amplified fragment length polymorph isms) linked to OA as a major attribute of drought resistance in wheat and thus to facilitate marker-assisted selection for drought resistance. We identified high and low OA lines of wheat and from their cross developed recombinant inbred lines (RILs) used in the molecular tagging of OA in relation to drought resistance in terms of plant production under stress. The significant positive co-segregation of OA, plant water status and yield under stress in this RIL population provided strong support for the important role of OA as a drought resistance mechanism sustaining wheat production under drought stress. This evidence was obtained in addition to the initial study of parental materials for constructing this RIL population, which also gave evidence for a strong correlation between OA and grain yield under stress. This research therefore provides conclusive evidence on the important role of OA in sustaining wheat yield under drought stress. The measurement of OA is difficult and the selection for drought resistance by the phenotypic expression of OA is practically impossible. This research provided information on the genetic basis of OA in wheat in relations to yield under stress. It provided the basic information to indicate that molecular marker assisted selection for OA in wheat is possible. The RIL population has been created by a cross between two agronomic spring wheat lines and the high OA recombinants in this population presented very high OA values, not commonly observed in wheat. These recombinants are therefore an immediate valuable genetic recourse for breeding well-adapted drought resistant wheat in Texas and Israel. We feel that this work taken as a whole eliminate the few previous speculated . doubts about the practical role of OA as an important mechanism of drought resistance in economic crop plants. As such it should open the way, in terms of both concept and the use of marker assisted selection, for improving drought resistance in wheat by deploying high osmotic adjustment.
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Dolja, Valerian V., Amit Gal-On, and Victor Gaba. Suppression of Potyvirus Infection by a Closterovirus Protein. United States Department of Agriculture, March 2002. http://dx.doi.org/10.32747/2002.7580682.bard.
Full textAbstract:
The plant virus family Polyviridae is the largest and most destructive of all plant viruses. Despite the continuous effort to develop resistant plant varieties, there is a desperate need for novel approaches conferring wide-range potyvirus resistance. Based on experiments with the tobacco etch potyvirus (TEV)-derived gene expression vector, we suggested approach for screening of the candidate resistance genes. This approach relies on insertion of the genes into a virus vector and evaluation of the phenotypes of the resulting recombinant viruses. The genes which suppress infection by the recombinant virus are selected as candidates for engineering transgenic resistance. Our analysis of the TEV variants expressing proteins of the beet yellows closterovirus (BYV) revealed that one of those, the leader proteinase (L-Pro), strongly and specifically interfered with the hybrid TEV infection. Since closterovirus L-Pro is evolutionary related to potyviral helper component-proteinase (HC-Pro), we suggested that the L-Pro interfered with HC-Pro function via a trans-dominant inhibitory effect. Based on these findings, we proposed to test two major hypotheses. First, we suggested that L-Pro-mediated suppression of potyvirus infection is a general phenomenon effective against a range of potyviruses. The second hypothesis stated that the suppression effect can be reproduced in transgenic plants expressing L-Pro, and can be utilized for generation of resistance to potyviruses. In accord with these hypotheses, we developed two original objectives of our proposal: A) to determine the range of the closterovirus-derived suppression of potyviral infection, and B) to try and utilize the L-Pro-mediated suppression for the development of transgenic resistance to potyviruses. In the first phase of the project, we have developed all major tools and technologies required for successful completion of the proposed research. These included TEV and ZYMV vectors engineered to express several closteroviral L-Pro variants, and generation of the large collection of transgenic plants. To our satisfaction, characterization of the infection phenotypes exhibited by chimeric TEV and ZYMV variants confirmed our first hypothesis. For instance, similar to TEV-L- Pro(BYV) chimera, ZYMV-L-Pro(LIYV) chimera was debilitated in its systemic spread. In contrast, ZYMV-GUS chimera (positive control) was competent in establishing vigorous systemic infection. These and other results with chimeric viruses indicated that several closteroviral proteinases inhibit long-distance movement of the potyviruses upon co-expression in infected plants. In order to complete the second objective, we have generated ~90 tobacco lines transformed with closteroviral L-Pro variants, as well as ~100 lines transformed with BYV Hsp70-homolog (Hsp70h; a negative control). The presence and expression of the trans gene in each line was initially confirmed using RT-PCR and RNA preparations isolated from plants. However, since detection of the trans gene-specific RNA can not guarantee production of the corresponding protein, we have also generated L-Pro- and Hsp70h-specific antisera using corresponding synthetic peptides. These antisera allowed us to confirm that the transgenic plant lines produced detectable, although highly variable levels of the closterovirus antigens. In a final phase of the project, we tested susceptibility of the transgenic lines to TEV infection. To this end, we determined that the minimal dilution of the TEV inoculum that is still capable of infecting 100% of nontransgenic plants was 1:20, and used 10 plants per line (in total, ~2,000 plants). Unfortunately, none of the lines exhibited statistically significant reduction in susceptibility. Although discouraging, this outcome prompted us to expand our experimental plan and conduct additional experiments. Our aim was to test if closteroviral proteinases are capable of functioning in trans. We have developed agroinfection protocol for BYV, and tested if co- expression of the L-Pro is capable of rescuing corresponding null-mutant. The clear-cut, negative results of these experiments demonstrated that L-Pro acts only in cis, thus explaining the lack of resistance in our transgenic plants. We have also characterized a collection of the L-Pro alanine- scanning mutants and found direct genetic evidence of the requirement for L-Pro in virus systemic spread. To conclude, our research supported by BARD confirmed one but not another of our original hypotheses. Moreover, it provided an important insight into functional specialization of the viral proteinases and generated set of tools and data with which we will be able to address the molecular mechanisms by which these proteins provide a variety of critical functions during virus life cycle.
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Lillehoj, Hyun, Dan Heller, and Mark Jenkins. Cellular and molecular identification of Eimeria Acervulina Merozoite Antigens eliciting protective immunity. United States Department of Agriculture, November 1992. http://dx.doi.org/10.32747/1992.7561056.bard.
Full textAbstract:
Coccidiosis, ubiquitous diseases of poultry, seriously impair the growth and feed utilization of livestock and poultry. Coccidiosis causes over $600 million annual losses world-wide and no vaccine is currently available. The goal of this study was to investigate the cellular and molecular mechanisms controlling protective immune responses to coccidia parasites in order to develop immunological control strategy against coccidiosis. The major findings of this study were: 1) cell-mediated immunity plays a major role in protection against coccidiosis, 2) when different genetic lines showing different levels of disease susceptibility were compared, higher T-cell response was seen in the strains of chickens showing higher disease resistance, 3) early interferon secretion was observed in more coccidia-resistant chicken strains, 4) both sporozoite and merozoite antigens were able to induce interferon production, and 5) chicken monoclonal antibodies which detect immunogenic coccidia proteins have been developed. This study provided a good background work for future studies toward the development of recombinant coccidial vaccine. Availability of chicken monoclonal antibodies which detect immunogenic coccidia proteins will enhance our ability to identify potential coccidial vaccine antigens.
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Fridman, Eyal, Jianming Yu, and Rivka Elbaum. Combining diversity within Sorghum bicolor for genomic and fine mapping of intra-allelic interactions underlying heterosis. United States Department of Agriculture, January 2012. http://dx.doi.org/10.32747/2012.7597925.bard.
Full textAbstract:
Heterosis, the enigmatic phenomenon in which whole genome heterozygous hybrids demonstrate superior fitness compared to their homozygous parents, is the main cornerstone of modern crop plant breeding. One explanation for this non-additive inheritance of hybrids is interaction of alleles within the same locus. This proposal aims at screening, identifying and investigating heterosis trait loci (HTL) for different yield traits by implementing a novel integrated mapping approach in Sorghum bicolor as a model for other crop plants. Originally, the general goal of this research was to perform a genetic dissection of heterosis in a diallel built from a set of Sorghum bicolor inbred lines. This was conducted by implementing a novel computational algorithm which aims at associating between specific heterozygosity found among hybrids with heterotic variation for different agronomic traits. The initial goals of the research are: (i) Perform genotype by sequencing (GBS) of the founder lines (ii) To evaluate the heterotic variation found in the diallel by performing field trails and measurements in the field (iii) To perform QTL analysis for identifying heterotic trait loci (HTL) (iv) to validate candidate HTL by testing the quantitative mode of inheritance in F2 populations, and (v) To identify candidate HTL in NAM founder lines and fine map these loci by test-cross selected RIL derived from these founders. The genetic mapping was initially achieved with app. 100 SSR markers, and later the founder lines were genotyped by sequencing. In addition to the original proposed research we have added two additional populations that were utilized to further develop the HTL mapping approach; (1) A diallel of budding yeast (Saccharomyces cerevisiae) that was tested for heterosis of doubling time, and (2) a recombinant inbred line population of Sorghum bicolor that allowed testing in the field and in more depth the contribution of heterosis to plant height, as well as to achieve novel simulation for predicting dominant and additive effects in tightly linked loci on pseudooverdominance. There are several conclusions relevant to crop plants in general and to sorghum breeding and biology in particular: (i) heterosis for reproductive (1), vegetative (2) and metabolic phenotypes is predominantly achieved via dominance complementation. (ii) most loci that seems to be inherited as overdominant are in fact achieving superior phenotype of the heterozygous due to linkage in repulsion, namely by pseudooverdominant mechanism. Our computer simulations show that such repulsion linkage could influence QTL detection and estimation of effect in segregating populations. (iii) A new height QTL (qHT7.1) was identified near the genomic region harboring the known auxin transporter Dw3 in sorghum, and its genetic dissection in RIL population demonstrated that it affects both the upper and lower parts of the plant, whereas Dw3 affects only the part below the flag leaf. (iv) HTL mapping for grain nitrogen content in sorghum grains has identified several candidate genes that regulate this trait, including several putative nitrate transporters and a transcription factor belonging to the no-apical meristem (NAC)-like large gene family. This activity was combined with another BARD-funded project in which several de-novo mutants in this gene were identified for functional analysis.
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Aharoni, Asaph, Zhangjun Fei, Efraim Lewinsohn, Arthur Schaffer, and Yaakov Tadmor. System Approach to Understanding the Metabolic Diversity in Melon. United States Department of Agriculture, July 2013. http://dx.doi.org/10.32747/2013.7593400.bard.
Full textAbstract:
Fruit quality is determined by numerous genetic factors that affect taste, aroma, color, texture, nutritional value and shelf life. To unravel the genetic components involved in the metabolic pathways behind these traits, the major goal of the project was to identify novel genes that are involved in, or that regulate, these pathways using correlation analysis between genotype, metabolite and gene expression data. The original and specific research objectives were: (1) Collection of replicated fruit from a population of 96 RI lines derived from parents distinguished by great diversity in fruit development and quality phenotypes, (2) Phenotypic and metabolic profiling of mature fruit from all 96 RI lines and their parents, (3) 454 pyrosequencing of cDNA representing mRNA of mature fruit from each line to facilitate gene expression analysis based on relative EST abundance, (4) Development of a database modeled after an existing database developed for tomato introgression lines (ILs) to facilitate online data analysis by members of this project and by researchers around the world. The main functions of the database will be to store and present metabolite and gene expression data so that correlations can be drawn between variation in target traits or metabolites across the RI population members and variation in gene expression to identify candidate genes which may impact phenotypic and chemical traits of interest, (5) Selection of RI lines for segregation and/or hybridization (crosses) analysis to ascertain whether or not genes associated with traits through gene expression/metabolite correlation analysis are indeed contributors to said traits. The overall research strategy was to utilize an available recombinant inbred population of melon (Cucumis melo L.) derived from phenotypically diverse parents and for which over 800 molecular markers have been mapped for the association of metabolic trait and gene expression QTLs. Transcriptomic data were obtained by high throughput sequencing using the Illumina platform instead of the originally planned 454 platform. The change was due to the fast advancement and proven advantages of the Illumina platform, as explained in the first annual scientific report. Metabolic data were collected using both targeted (sugars, organic acids, carotenoids) and non-targeted metabolomics analysis methodologies. Genes whose expression patterns were associated with variation of particular metabolites or fruit quality traits represent candidates for the molecular mechanisms that underlie them. Candidate genes that may encode enzymes catalyzingbiosynthetic steps in the production of volatile compounds of interest, downstream catabolic processes of aromatic amino acids and regulatory genes were selected and are in the process of functional analyses. Several of these are genes represent unanticipated effectors of compound accumulation that could not be identified using traditional approaches. According to the original plan, the Cucurbit Genomics Network (http://www.icugi.org/), developed through an earlier BARD project (IS-3333-02), was expanded to serve as a public portal for the extensive metabolomics and transcriptomic data resulting from the current project. Importantly, this database was also expanded to include genomic and metabolomic resources of all the cucurbit crops, including genomes of cucumber and watermelon, EST collections, genetic maps, metabolite data and additional information. In addition, the database provides tools enabling researchers to identify genes, the expression patterns of which correlate with traits of interest. The project has significantly expanded the existing EST resource for melon and provides new molecular tools for marker-assisted selection. This information will be opened to the public by the end of 2013, upon the first publication describing the transcriptomic and metabolomics resources developed through the project. In addition, well-characterized RI lines are available to enable targeted breeding for genes of interest. Segregation of the RI lines for specific metabolites of interest has been shown, demonstrating the utility in these lines and our new molecular and metabolic data as a basis for selection targeting specific flavor, quality, nutritional and/or defensive compounds. To summarize, all the specific goals of the project have been achieved and in many cases exceeded. Large scale trascriptomic and metabolomic resources have been developed for melon and will soon become available to the community. The usefulness of these has been validated. A number of novel genes involved in fruit ripening have been selected and are currently being functionally analyzed. We thus fully addressed our obligations to the project. In our view, however, the potential value of the project outcomes as ultimately manifested may be far greater than originally anticipated. The resources developed and expanded under this project, and the tools created for using them will enable us, and others, to continue to employ resulting data and discoveries in future studies with benefits both in basic and applied agricultural - scientific research.
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Lers, Amnon, Majid R. Foolad, and Haya Friedman. genetic basis for postharvest chilling tolerance in tomato fruit. United States Department of Agriculture, January 2014. http://dx.doi.org/10.32747/2014.7600014.bard.
Full textAbstract:
ABSTRACT Postharvest losses of fresh produce are estimated globally to be around 30%. Reducing these losses is considered a major solution to ensure global food security. Storage at low temperatures is an efficient practice to prolong postharvest performance of crops with minimal negative impact on produce quality or human health and the environment. However, many fresh produce commodities are susceptible to chilling temperatures, and the application of cold storage is limited as it would cause physiological chilling injury (CI) leading to reduced produce quality. Further, the primary CI becomes a preferred site for pathogens leading to decay and massive produce losses. Thus, chilling sensitive crops should be stored at higher minimal temperatures, which curtails their marketing life and in some cases necessitates the use of other storage strategies. Development of new knowledge about the biological basis for chilling tolerance in fruits and vegetables should allow development of both new varieties more tolerant to cold, and more efficient postharvest storage treatments and storage conditions. In order to improve the agricultural performance of modern crop varieties, including tomato, there is great potential in introgression of marker-defined genomic regions from wild species onto the background of elite breeding lines. To exploit this potential for improving tomato fruit chilling tolerance during postharvest storage, we have used in this research a recombinant inbred line (RIL) population derived from a cross between the red-fruited tomato wild species SolanumpimpinellifoliumL. accession LA2093 and an advanced Solanum lycopersicumL. tomato breeding line NCEBR-1, developed in the laboratory of the US co-PI. The original specific objectives were: 1) Screening of RIL population resulting from the cross NCEBR1 X LA2093 for fruit chilling response during postharvest storage and estimation of its heritability; 2) Perform a transcriptopmic and bioinformatics analysis for the two parental lines following exposure to chilling storage. During the course of the project, we learned that we could measure greater differences in chilling responses among specific RILs compared to that observed between the two parental lines, and thus we decided not to perform transcriptomic analysis and instead invest our efforts more on characterization of the RILs. Performing the transcriptomic analysis for several RILs, which significantly differ in their chilling tolerance/sensitivity, at a later stage could result with more significant insights. The RIL population, (172 lines), was used in field experiment in which fruits were examined for chilling sensitivity by determining CI severity. Following the field experiments, including 4 harvest days and CI measurements, two extreme tails of the response distribution, each consisting of 11 RILs exhibiting either high sensitivity or tolerance to chilling stress, were identified and were further examined for chilling response in greenhouse experiments. Across the RILs, we found significant (P < 0.01) correlation between field and greenhouse grown plants in fruit CI. Two groups of 5 RILs, whose fruits exhibited reproducible chilling tolerant/sensitive phenotypes in both field and greenhouse experiments, were selected for further analyses. Numerous genetic, physiological, biochemical and molecular variations were investigated in response to postharvest chilling stress in the selected RILs. We confirmed the differential response of the parental lines of the RIL population to chilling stress, and examined the extent of variation in the RIL population in response to chilling treatment. We determined parameters which would be useful for further characterization of chilling response in the RIL population. These included chlorophyll fluorescence Fv/Fm, water loss, total non-enzymatic potential of antioxidant activity, ascorbate and proline content, and expression of LeCBF1 gene, known to be associated with cold acclimation. These parameters could be used in continuation studies for the identification and genetic mapping of loci contributing to chilling tolerance in this population, and identifying genetic markers associated with chilling tolerance in tomato. Once genetic markers associated with chilling tolerance are identified, the trait could be transferred to different genetic background via marker-assisted selection (MAS) and breeding. The collaborative research established in this program has resulted in new information and insights in this area of research and the collaboration will be continued to obtain further insights into the genetic, molecular biology and physiology of postharvest chilling tolerance in tomato fruit. The US Co-PI, developed the RIL population that was used for screening and measurement of the relevant chilling stress responses and conducted statistical analyses of the data. Because we were not able to grow the RIL population under field conditions in two successive generations, we could not estimate heritability of response to chilling temperatures. However, we plan to continue the research, grow the RIL progeny in the field again, and determine heritability of chilling tolerance in a near future. The IS and US investigators interacted regularly and plan to continue and expand on this study, since combing the expertise of the Co-PI in genetics and breeding with that of the PI in postharvest physiology and molecular biology will have great impact on this line of research, given the significant findings of this one-year feasibility project.
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Dubcovsky, Jorge, Tzion Fahima, and Ann Blechl. Positional cloning of a gene responsible for high grain protein content in tetraploid wheat. United States Department of Agriculture, September 2003. http://dx.doi.org/10.32747/2003.7695875.bard.
Full textAbstract:
High Grain Protein Content (GPC) is a desirable trait in breadmaking and pasta wheat varieties because of its positive effects on quality and nutritional value. However, selection for GPC is limited by our poor understanding of the genes involved in the accumulation of protein in the grain. The long-term goal of this project is to provide a better understanding of the genes controlling GPC in wheat. The specific objectives of this project were: a) to develop a high-density genetic map of the GPC gene in tetraploid wheat, b) to construct a T. turgidum Bacterial Artificial Chromosome (BAC) library, c) to construct a physical map of the GPC gene and identify a candidate for the GPC gene. A gene with a large effect on GPC was detected in Triticum turgidum var. dicoccoides and was previously mapped in the short arm of chromosome 6B. To define better the position of the Gpc-B1 locus we developed homozygous recombinant lines with recombination events within the QTL region. Except for the 30-cM region of the QTL these RSLs were isogenic for the rest of the genome minimizing the genetic variability. To minimize the environmental variability the RSLs were characterized using 10 replications in field experiments organized in a Randomized Complete Block Design, which were repeated three times. Using this strategy, we were able to map this QTL as a single Mendelian locus (Gpc-B1) on a 2.6-cM region flanked by RFLP markers Xcdo365 and Xucw67. All three experiments showed that the lines carrying the DIC allele had an average absolute increase in GPC of 14 g/kg. Using the RFLP flanking markers, we established the microcolinearity between a 2.l-cM region including the Gpc-B1 gene in wheat chromosome 6BS and a 350-kb region on rice chromosome 2. Rice genes from this region were used to screen the Triticeae EST collection, and these ESTs were used to saturate the Gpc-B1 region with molecular markers. With these new markers we were able to map the Gpc-B1 locus within a 0.3-cM region flanked by PCR markers Xucw83 and Xucw71. These flanking markers defined a 36-kb colinear region with rice, including one gene that is a potential candidate for the Gpc-B1 gene. To develop a physical map of the Gpc-B1 region in wheat we first constructed a BAC library of tetraploid wheat, from RSL#65 including the high Gpc-B1 allele. We generated half- million clones with an average size of l3l-kb (5.1 X genome equivalents for each of the two genomes). This coverage provides a 99.4% probability of recovering any gene from durum wheat. We used the Gpc-BI flanking markers to screen this BAC library and then completed the physical map by chromosome walking. The physical map included two overlapping BACs covering a region of approximately 250-kb, including two flanking markers and the Gpc-B1 gene. Efforts are underway to sequence these two BACs to determine if additional wheat genes are present in this region. Weare also developing new RSLs to further dissect this region. We developed PCR markers for flanking loci Xucw79andXucw71 to facilitate the introgression of this gene in commercial varieties by marker assisted selection (httQ://maswheat.ucdavis.edu/ orotocols/HGPC/index.hlm). Using these markers we introgressed the Gpc-B1 gene in numerous pasta and common wheat breeding lines.
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Feldman, Moshe, Eitan Millet, Calvin O. Qualset, and Patrick E. McGuire. Mapping and Tagging by DNA Markers of Wild Emmer Alleles that Improve Quantitative Traits in Common Wheat. United States Department of Agriculture, February 2001. http://dx.doi.org/10.32747/2001.7573081.bard.
Full textAbstract:
The general goal was to identify, map, and tag, with DNA markers, segments of chromosomes of a wild species (wild emmer wheat, the progenitor of cultivated wheat) determining the number, chromosomal locations, interactions, and effects of genes that control quantitative traits when transferred to a cultivated plant (bread wheat). Slight modifications were introduced and not all objectives could be completed within the human and financial resources available, as noted with the specific objectives listed below: 1. To identify the genetic contribution of each of the available wild emmer chromosome-arm substitution lines (CASLs) in the bread wheat cultivar Bethlehem for quantitative traits, including grain yield and its components and grain protein concentration and yield, and the effect of major loci affecting the quality of end-use products. [The quality of end-use products was not analyzed.] 2. To determine the extent and nature of genetic interactions (epistatic effects) between and within homoeologous groups 1 and 7 for the chromosome arms carrying "wild" and "cultivated" alleles as expressed in grain and protein yields and other quantitative traits. [Two experiments were successful, grain protein concentration could not be measured; data are partially analyzed.] 3. To derive recombinant substitution lines (RSLs) for the chromosome arms of homoeologous groups 1 and 7 that were found previously to promote grain and protein yields of cultivated wheat. [The selection of groups 1 and 7 tons based on grain yield in pot experiments. After project began, it was decided also to derive RSLs for the available arms of homoeologous group 4 (4AS and 4BL), based on the apparent importance of chromosome group 4, based on early field trials of the CASLs.] 4. To characterize the RSLs for quantitative traits as in objective 1 and map and tag chromosome segments producing significant effects (quantitative trait loci, QTLs by RFLP markers. [Producing a large population of RSLs for each chromosome arm and mapping them proved more difficult than anticipated, low numbers of RSLs were obtained for two of the chromosome arms.] 5. To construct recombination genetic maps of chromosomes of homoeologous groups 1 and 7 and to compare them to existing maps of wheat and other cereals [Genetic maps are not complete for homoeologous groups 4 and 7.] The rationale for this project is that wild species have characteristics that would be valuable if transferred to a crop plant. We demonstrated the sequence of chromosome manipulations and genetic tests needed to confirm this potential value and enhance transfer. This research has shown that a wild tetraploid species harbors genetic variability for quantitative traits that is interactive and not simply additive when introduced into a common genetic background. Chromosomal segments from several chromosome arms improve yield and protein in wheat but their effect is presumably enhanced when combination of genes from several segments are integrated into a single genotype in order to achieve the benefits of genes from the wild species. The interaction between these genes and those in the recipient species must be accounted for. The results of this study provide a scientific basis for some of the disappointing results that have historically obtained when using wild species as donors for crop improvement and provide a strategy for further successes.
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Jordan, Ramon L., Abed Gera, Hei-Ti Hsu, Andre Franck, and Gad Loebenstein. Detection and Diagnosis of Virus Diseases of Pelargonium. United States Department of Agriculture, July 1994. http://dx.doi.org/10.32747/1994.7568793.bard.
Full textAbstract:
Pelargonium (Geranium) is the number one pot plant in many areas of the United States and Europe. Israel and the U.S. send to Europe rooted cuttings, foundation stocks and finished plants to supply a certain share of the market. Geraniums are propagated mainly vegetatively from cuttings. Consequently, viral diseases have been and remain a major threat to the production and quality of the crop. Among the viruses isolated from naturally infected geraniums, 11 are not specific to Pelargonium and occur in other crops while 6 other viruses seem to be limited to geranium. However, several of these viruses are not sufficiently characterized to conclude that they are distinct agents and their nomenclature and taxonomy are confusing. The ability to separate, distinguish and detect the different viruses in geranium will overcome obstacles te developing effective detection and certification schemes. Our focus was to further characterize some of these viruses and develop better methods for their detection and control. These viruses include: isolates of pelargonium line pattern virus (PLPV), pelargonium ringspot virus (PelRSV), pelargonium flower break virus (PFBV), pelargonium leaf curl (PLCV), and tomato ringspot virus (TomRSV). Twelve hybridoma cell lines secreting monoclonal antibodies specific to a geranium isolate of TomRSV were produced. These antibodies are currently being characterized and will be tested for the ability to detect TomRSV in infected geraniums. The biological, biochemical and serological properties of four isometric viruses - PLPV, PelRSV, and PFBV (and a PelRSV-like isolate from Italy called GR57) isolated from geraniums exhibiting line and ring pattern or flower break symptoms - and an isolate ol elderbeny latent virus (ELV; which the literature indicates is the same as PelRSV) have been determined Cloned cDNA copies of the genomic RNAs of these viruses were sequenced and the sizes and locations of predicted viral proteins deduced. A portion of the putative replicase genes was also sequenced from cloned RT-PCR fragments. We have shown that, when compared to the published biochemical and serological properties, and sequences and genome organizations of other small isometric plant viruses, all of these viruses should each be considered new, distinct members of the Carmovirus group of the family Tombusviridae. Hybridization assays using recombinant DNA probes also demonstrated that PLPV, PelRSV, and ELV produce only one subgenomic RNA in infected plants. This unusual property of the gene expression of these three viruses suggests that they are unique among the Carmoviruses. The development of new technologies for the detection of these viruses in geranium was also demonstrated. Hybridization probes developed to PFBV (radioactively-labeled cRNA riboprobes) and to PLPV (non-radioactive digoxigenin-labeled cDNAs) were generally shown to be no more sensitive for the detection of virus in infected plants than the standard ELISA serology-based assays. However, a reverse transcriptase-polymerase chain reaction assay was shown to be over 1000 times more sensitive in detecting PFBV in leaf extracts of infected geranium than was ELISA. This research has lead to a better understanding of the identity of the viruses infecting pelargonium and to the development of new tools that can be used in an improved scheme of providing virus-indexed pelargonium plants. The sequence information, and the serological and cloned DNA probes generated from this work, will allow the application of these new tools for virus detection, which will be useful in domestic and international indexing programs which are essential for the production of virus-free germplasm both for domestic markets and the international exchange of plant material.