Abstract
BackgroundIn common bean, expressed sequence tags (ESTs) are an underestimated source of gene-based markers such as insertion-deletions (Indels) or single-nucleotide polymorphisms (SNPs). However, due to the nature of these conserved sequences, detection of markers is difficult and portrays low levels of polymorphism. Therefore, development of intron-spanning EST-SNP markers can be a valuable resource for genetic experiments such as genetic mapping and association studies.ResultsIn this study, a total of 313 new gene-based markers were developed at target genes. Intronic variation was deeply explored in order to capture more polymorphism. Introns were putatively identified after comparing the common bean ESTs with the soybean genome, and the primers were designed over intron-flanking regions. The intronic regions were evaluated for parental polymorphisms using the single strand conformational polymorphism (SSCP) technique and Sequenom MassARRAY system. A total of 53 new marker loci were placed on an integrated molecular map in the DOR364 × G19833 recombinant inbred line (RIL) population. The new linkage map was used to build a consensus map, merging the linkage maps of the BAT93 × JALO EEP558 and DOR364 × BAT477 populations. A total of 1,060 markers were mapped, with a total map length of 2,041 cM across 11 linkage groups. As a second application of the generated resource, a diversity panel with 93 genotypes was evaluated with 173 SNP markers using the MassARRAY-platform and KASPar technology. These results were coupled with previous SSR evaluations and drought tolerance assays carried out on the same individuals. This agglomerative dataset was examined, in order to discover marker-trait associations, using general linear model (GLM) and mixed linear model (MLM). Some significant associations with yield components were identified, and were consistent with previous findings.ConclusionsIn short, this study illustrates the power of intron-based markers for linkage and association mapping in common bean. The utility of these markers is discussed in relation with the usefulness of microsatellites, the molecular markers by excellence in this crop.
Highlights
In common bean, expressed sequence tags (ESTs) are an underestimated source of gene-based markers such as insertion-deletions (Indels) or single-nucleotide polymorphisms (SNPs)
55 pairs of primers were designed over 33 genes involved in the nodulation process in model legumes [27,28]; 63 pairs of primers were designed over 48 transcription factors identified under phosphorus stress [29]; and 195 pairs of primers were designed over 190 putative soybean genes involved in nodule development [30]
Pilot amplification on these 313 intronic regions using the control genotypes DOR364, BAT477 and G19833 was successful in 77% of the cases
Summary
In common bean, expressed sequence tags (ESTs) are an underestimated source of gene-based markers such as insertion-deletions (Indels) or single-nucleotide polymorphisms (SNPs). Single nucleotide polymorphisms (SNPs) are the most abundant class of polymorphic sites in any genome They have become a powerful tool in genetic mapping, association studies, diversity analysis and positional cloning [1]. EST libraries of the Mesoamerican genotype Negro Jamapa 81 and the Andean genotype G19833 were used to establish the first consolidated resource of SNP markers [9]. Some of these SNPs were mapped in the population DOR364 × G19833 using mismatch cleavage nuclease CEL I [10] and single strand conformational polymorphism (SSCP) [7]. This constraint can be avoided by means of a deeper exploration of the intronic regions
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