A single recessive gene, rxp, controls the bacterial leaf pustule (BLP) resistance in soybean and in our previous article, it has been mapped on linkage group (LG) D2 of molecular genetic map of soybean. A total of 130 recombinant inbred lines (RILs) from a cross between BLP-resistant SS2-2 and BLP-...
A single recessive gene, rxp, controls the bacterial leaf pustule (BLP) resistance in soybean and in our previous article, it has been mapped on linkage group (LG) D2 of molecular genetic map of soybean. A total of 130 recombinant inbred lines (RILs) from a cross between BLP-resistant SS2-2 and BLP-susceptible Jangyeobkong were used to identify molecular markers linked to rxp. Fifteen simple sequence repeat (SSR) markers on LG D2 were screened to construct a genetic map of rxp locus. Only four SSR markers, Satt135, Satt372, Satt448, and Satt486, showed parental polymorphisms. Using these markers, genetic scaffold map was constructed covering 26.2cM. Based on the single analysis of variance, Satt372 among these four SSR markers was the most significantly associated with the resistance to BLP. To develop new amplified fragment length polymorphism (AFLP) marker linked to the resistance gene, bulked segregant analysis (BSA) was employed. Resistance and susceptible bulks were made by pooling equal amount of genomic DNAs from ten of each in the segregating population. A total of 192 primer combinations were used to identify specific bands to the resistance, selecting three putative AFLP markers. These AFLP markers produced the fragment present in SS2-2 and the resistant bulk, and not in Jangyeobkong and the susceptible bulk. Linkage analysis revealed that McctEact97 $(P=0.0004,\;R^2=14.67\%)$ was more significant than Satt372, previously reported as the most closely linked marker.
A single recessive gene, rxp, controls the bacterial leaf pustule (BLP) resistance in soybean and in our previous article, it has been mapped on linkage group (LG) D2 of molecular genetic map of soybean. A total of 130 recombinant inbred lines (RILs) from a cross between BLP-resistant SS2-2 and BLP-susceptible Jangyeobkong were used to identify molecular markers linked to rxp. Fifteen simple sequence repeat (SSR) markers on LG D2 were screened to construct a genetic map of rxp locus. Only four SSR markers, Satt135, Satt372, Satt448, and Satt486, showed parental polymorphisms. Using these markers, genetic scaffold map was constructed covering 26.2cM. Based on the single analysis of variance, Satt372 among these four SSR markers was the most significantly associated with the resistance to BLP. To develop new amplified fragment length polymorphism (AFLP) marker linked to the resistance gene, bulked segregant analysis (BSA) was employed. Resistance and susceptible bulks were made by pooling equal amount of genomic DNAs from ten of each in the segregating population. A total of 192 primer combinations were used to identify specific bands to the resistance, selecting three putative AFLP markers. These AFLP markers produced the fragment present in SS2-2 and the resistant bulk, and not in Jangyeobkong and the susceptible bulk. Linkage analysis revealed that McctEact97 $(P=0.0004,\;R^2=14.67\%)$ was more significant than Satt372, previously reported as the most closely linked marker.
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제안 방법
Pre-selective amplification reaction was performed using two AFLP primers with a single selective nucleotide and AmphTaq polymerase (Applied Biosystems, Foster city, CA, USA) in a PTC 100 programmable Thermal Controller (MJ Research Inc. Watertown, MA, USA). Cycling conditions consisted of a initial incubation at 72 ℃ for 2 min, followed by 30 sec of denaturation at 94 ℃, 30 sec annealing at 56 ℃, and 2 min of extension at 72 ℃ for 20 cycles, and 30 min of final incubation at 60 ℃.
The first objective of this research was to confirm the resistance gene for BLP in the resistant mutant parent, SS2- 2, and to construct a scaffold map for resistance locus using SSR markers. The second objective of this research was to develop the AFLP markers tightly linked to a resistance gene for BLP using BSA (Michelmore et al.
Amplified products from different SSR loci carrying the fluorescent label can be simultaneously analyzed in the same gel lane if allele size ranges do not overlap. The samples of combined PCR products were loaded and separated on a standard DNA sequencing gel containing 4 % polyacrylamide, 8 M urea, and 0.5X TBE at 1700 V constant power for 2 hours on an ABI 377 DNA sequencer (Applied Biosystems, Foster City, CA, USA). Genescan and Geno- typer 2.
The first objective of this research was to confirm the resistance gene for BLP in the resistant mutant parent, SS2- 2, and to construct a scaffold map for resistance locus using SSR markers. The second objective of this research was to develop the AFLP markers tightly linked to a resistance gene for BLP using BSA (Michelmore et al., 1991).
대상 데이터
Based on the SSR markers and phenotypic data, resistant and susceptible bulks were made by pooling separately with equal amount of 10 pre-selective amplified DNA obtained from resistant individuals and susceptible individuals in the population. The resistant and susceptible bulks were examined with 6 EcoRI and 32 Msel pnmers, bringing up a total of 192 different primer combinations. Primer combinations yielding candidate markers were subsequently applied to all individuals of the population in order to confirm the linkage between the AFLP markers and a resistance locus.
데이터처리
Associations between marker genotypes and variations in phenotypic reaction were tested by single factor analysis of variance (SF-ANOVA) (Tanksley, 1989). Multiple linear regression within a single linkage group (SLG-Regr) was also applied to determine the most significant marker among those detected using SFANOVA. Two-factor analysis of vanance was also used for checking all possible pairs of significant marker.
Marker data were obtained from each segregating progeny, which were used to construct a genetic map. The segregation ratio of each marker was calculated with a chi-square test under the hypothesis of the expected 1 : 1 ratio. Linkage analysis of the entire set of markers was performed using MAPMAKER version 3.
이론/모형
, 1987). A linkage map containing SSR loci and AFLP loci was constructed using the Kosambi map function (Kosambi, 1944). For combining markers into linkage groups, a minimum likelihood of odds (LOD) of 3.
Tb obtain genomic DNA from soybean for the SSR and AFLP marker analyses, two or three young and healthy soybean leaves of each plant were harvested prior to full expansion in the greenhouse, placed in the paper bag on ice, and stored at 80℃ until use. Genomic DNA was isolated using hexadecylatn methylammonium bromide (CTAB) method described by the procedure of Keim et al. (1988).
2. Segregation of bulked segregant analysis (BSA) of 10 BLP-resistance and 10 BLP-susceptible lines using the AFLP technique. The circles indicate the potential marker (McctEact97) linked to the rxp Each line represents SS2-2, Jangyeobkong, the resistant bulk, and the susceptible bulk, from top to bottom.
The general linear model (GLM) procedure of SAS was used for statistical analyses. Associations between marker genotypes and variations in phenotypic reaction were tested by single factor analysis of variance (SF-ANOVA) (Tanksley, 1989).
참고문헌 (18)
Bernard, R. L. and M G Weiss 1973 Qualitative genetics. In B E. Caldwell. ed p 117-154. Soybean improvement, production and uses. ASA and Madison, WI
Cregan, P B , T Jarvik, A L. Bush, R. C Shoemaker, K. G Lark, A L. Kahler, T. T. Van Toai, D. G. Lohnes, J Chung, and J. E. Specht 1999 An integrated linkage map of the soybean genome. Crop Sci 39 . 1464-1490
Diwan, N and P B Cregan 1997 Automated sizing of fluorescent-labeled simple sequence repeat (SSR) marker to assay genetic variation soybean. Theor. Appl. Genet. 95 . 723-733
Feaster, C. V. 1951. Bacterial pustule disease on soybean. article inoculation, varietal response, and inheritance of resistance Missouri Agricultural Experiment Station Bulletin p 487
Hartwig, E E and S. G Lehman 1951. Inheritance of resistance to the bacterial pustule disease in soybean Agron. J 43 22-23
Keim, P., T. C. Olson, and R. C. Shoemaker. 1988. A rapid protocol forisolating soybean DNA. Soybean Genet Newsl 15 150-154
Kosambi, D. D. 1944. The estimation of map distances from recombination values. Ann. Eugen. 12 : 172-175
Lander E S , P. Green, J Abrahamson, A. Barlow, M. J. Daly, S E Lincoln, and L. Newburg. 1987. Mapmaker. an interactive computer package for constructing primary genetic maps of experimental and natural populations Genomics 1 : 174-181
Michelmore, R W, I Paran, and R. V Kessel. 1991. Identification of markers linked to disease-resistance genes by bulked segregant analysis: a rapid method to detect markers in specific genomic regions by using segregating populations. Proc. Natl. Acad Sci 88 9828-9832
Narvel, J. M., L. R. Jakkula, D. V Philips, T Wang, S-H Lee, and H. R. Boerma. 2000. Molecular mapping and pedigree analysis of the Rxp locus controlling bacterial pustule resistance in soybean. J. Hered. 92 . 267-270
Tanksley, S. D , N. D Young, A. H. Paterson, and M W Bionierbale 1989 RFLP mapping in plant breeding: new tool for an old science Bio/technology 7 257-264
Tsunematsu, H , A Yoshimura, M Yano, T Sasaki, and N. Iwata. 1996 RFLP framework map using recombinant lines in rice. Breeding Sci 46 279-284
Van, K., B -K Ha, E -Y Hwang, M Y. Kim, N.-C. Pack, S. Heu, and S.-H. Lee. 2004 SSR Mapping of genes conditioning soybean resistance to six isolate of Xanthomonas axonopodis pv glycines. Korean J. Genet. 26 . 47-54
Vos, P., R. Hogers, M Bleeker, and M. Zabeau. 1995. AFLP. A new technique for DNA fingerprinting. Nucl. Acids Res 23 4407-4414
Ziegle, J. S , Y Su, K. P. Corcoran, L. Nie, P E Mayrand, L B. Hoff, L. J. McBride, M. N. Kronick, and S. R. Kiehl 1992 Application of automated DNA sizing technology for genotyping microsatellite loci Genomics 14 . 1026-1031
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