Seed quality traits (especially hardseededness and seed size) and resistance to disease (in particular powdery mildew caused by Erysiphe polygoni DC and charcoal rot caused by Macrophomina phaseolina (Tassi) Goid.) are key factors for the success and expansion of the mungbean (Vigna radiata (L.) Wilczek) industry in Australia. Understanding the inheritance of these characters will undoubtedly increase the efficiency of mungbean breeding programs and increase the potential to produce more desirable cultivars for both the local and overseas markets. Map based QTL analysis has recently become a powerful tool in the identification and manipulation of genes underlying important traits where little is known about them. With this in mind, two recombinant inbred (RI) populations segregating for seed characters and powdery mildew resistance were produced and regions of the mungbean genome conditioning these important traits were identified using QTL analysis.
A genetic linkage map of mungbean consisting of 255 RFLP loci was developed using a RI population of 80 individuals. The population was derived from an inter-subspecific cross between the large, softseeded cultivar Berken (V. radiata subsp. radiata) and the completely hardseeded wild accession, ACC 41 (V. radiata subsp. Sublobata). The map, which comprised 13 linkage groups, spanned 738 cM with an average distance between markers of 3.0 cM and a maximum distance between linked markers of 15.4 cM. This map was compared to a previously published map of lablab (Lablab purpureus L.) using a common set of 65 RFLP probes. In contrast to some other comparative mapping studies among members of the Fabaceae, where a high level of chromosomal rearrangement has been observed, marker order between mungbean and lablab was found to be highly conserved. However, there was evidence that the two genomes had accumulated a large number of duplications/deletions after they diverged.
To analyse seed characters, a framework set of 64 of the 255 RFLP clones and one morphological marker (leaf lobing) were selected to construct a linkage map using a population of 227 RI individuals from the same inter-subspecific cross. These markers were chosen to be located approximately 15 cM apart and/or at the distal regions of existing linkage groups. Using this population, a linkage map consisting of 79 loci was generated over 13 linkage groups, spanning a total distance of 685 cMwith an average distance between markers of 10.4 cM and a maximum distance of 20.7 cM. QTL analysis revealed four regions associated with hardseededness, with individual loci explaining between 4.9% and 23.2% of the variation. For seed size, 11 regions were identified, with individual loci explaining between 4.2% and 18.8% of the variation. Two of the loci involved in hardseededness co-located with those conditioning seed size, indicating that there may be a physiological effect of seed size on hardseededness in this population.
A major locus that conferred resistance to powdery mildew in mungbean was also identified using a second population of 147 RI individuals derived from an intraspecific cross between Berken and the introduction line ATF 3640. Berken was susceptible and ATF 3640 highly resistant to the isolates of powdery mildew used in this study. To generate a linkage map for this population, 51 RFLP clones identified as being polymorphic from parental screening for the Berken x ACC 41 linkage map were selected to screen the Berken x ATF 3640 population. The 51 probes generated 52 mapped loci, which were used to construct a linkage map spanning 350 cM of the mungbean genome over 10 linkage groups. Using these markers, a major locus was identified that explained up to 88.3% of the total variation in the resistance response to the pathogen.
To demonstrate the feasibility of developing user-friendly markers linked to important traits in mungbean, a marker closely linked to the powdery mildew resistance locus was then used to screen a BAC (bacterial artificial chromosome) library. Subclones of positive BAC clones were probed with oligonucleotides containing both AT and ATT repeats to identify those that contained putative microsatellites. Sequence analysis of positive subclones identified two unique sequences containing (AT)n repeats, one unique sequence containing an (ATT)n repeat and one unique sequence containing a compound repeat. A polymorphic SSR (simple sequence repeat) marker closely linked to the major locus conditioning powdery mildew resistance was generated from one of these. This marker could be used in breeding programs aimed at generating powdery mildew resistant cultivars. This approach could be used to develop more user-friendly markers for loci conditioning other important agronomic traits in mungbean.