Construct Of Adzuki Bean's Linkage Map Of Molecular Marker, Association Analyse Of Agronomic Characters And The Location Of Its QTL

Azuki bean [Vigna angularis (Willd) Ohwi & Ohashi] is one of the major food beans in China. Along with the improvement of the people's life standard, it is necessary to develop genetic and breeding research for Adzuki bean improvement, to meet the requirement of increasing food colourfulness and market demand.In the major grain crops such as rice, maize and wheat, it has been proved that DNA molecular marker assisted selection is a useful tool for raising selection efficiency. In Adzuki bean, however, the situation is relative lagging in DNA molecular marker assisted selection, except several reports on polymorphic identification among germplasms using random amplified markers. A few has been done for the development of SSR and some other molecular markers with speciese specificity. Several reports are about the construction of molecular marker linkage map and QTL identification of quantitative traits (Han et al, 2005; Kaga et al, 1996, 2000; Wang et al, 2004). They use several kinds of molecular markers, such as RAPD, SSR, AFLP and RFLP, to locate several agronomic characters. No report has been found to use RAMP marker, the most useful to Adzuki bean, to construct linkage map and identify QTL.In this study, 94 Adzuki bean varieties from 12 provinces were used as individuals in the speciese population of Adzuki bean. Association analysis was conducted based on the linkage disequilibrium between the phenotypic variation of the agronomic characters and polymorphic alleles of RAPD, AFLP and RAMP markers. The QTLs were identified and their explaination rates for the phenotypic variation of these agronomic characters were estimated.In a random block experiment with 3 replications, the 94 Adzuki bean varieties were planted and phenotyped for their plant height, node number of main stem, branch number of main stem, pod length, pod width, per plant pod number, per plant grain number, 100-grain weight and per plant grain weight. Variation analysis showed that highly significant or significant differences were detected among the 94 varieties for all the 9 agronomic characters except pod width. DNA molecular polymorphism among the 94 varieties was genotyped by PCR amplification using 10 RAPD primers, 10 AFLP primer/enzyme combinations and 42 RAMP primer combinations. 1267 polymorphic alleles were identified as a total. The average polymorphic rates of RAPD, AFLP and RAMP markers were 92.7%, 36.9% and 96.4% respectively. This result indicated that abundant polymorphism of DNA molecular markers can be found among Adzuki varieties and used for genetic diversity research. By comprehensive comparison among the three kinds of molecular markers, RAMP is believed to be a useful technique to genotype genetic diversity among Adizuki bean varieties.57540 out of 802011 linkage disequilibrium values (R~2) estimated between each two of the 1267 polymorphic alleles were significalt on the level of 5%, including 100 RAPD, 309 AFLP and 306 RAMP markers, summing to 715 polymorphic alleles as a total. A procedure for comparison and alignment of R~2 values among the alleles was designed with JAVA language, and used to located 551 of the 1267 polymorphic alleles into 16 linkage groups, including 296, 62, 55, 27, 21, 14, 13, 12, 9, 8, 7, 6, 6, 5, 5 and 5 alleles respectively.TASSEL software was used for association analysis between the phenotypic data of the eight agronomic characters, significant among the 94 varieties in variation analysis, and their polymorphic allele data of DNA molecular markers, using the Q values of population structure as covariables. The association degree was estimated between each of the agronomic characters and the polymorphic alleles. The result showed that 96 of the 1267 alleles were significantly associated with the eight agronomic characters. Fourteen, seven, eighteen, nine, six, twelve, twenty and nine alleles were associated with plant height, node number of main stem, branch number of main stem, pod length, pod width, per plant pod number, per plant grain number, 100-grain weight and per plant grain weight, and explained 90.6%, 77.33%, 97.41%, 83.34%, 30.32%, 89.79%, 87.74% and 60.44% of their phenotypic variation, respectively.From the linkage map of DNA molecular markers, the polymorphic alleles significantly associated with the agronomic characters, as well as their nearby linkage alleles, were looked for, and used to locate the relative sites of the QTLs of these characters in the likage map of DNA molecular markers. Eighty-three of the ninty-six polymorphic alleles associated with the eight characters were located into the linkage map of DNA molecular markers, implying that QTLs controlling these charaters are present around these alleles. The numbers of polymorphisc alleles associated with plant height, node number of main stem, branch number of main stem, pod length, pod width, per plant pod number, per plant grain number, 100-grain weight and per plant grain weight were 10, 6, 16, 8, 6, 11, 19 and 7, thirteen of which were associated with two or more agronomic characters. Furthermore, some characters were found to be associated with two or more polymorphic alleles, nearby in the same linkage group. From this result, it can be concluded that major QTLs controlling these agronomic characters maybe present around these polymorphic alleles.