EST-derived genic molecular markers: development and utilization for generating an advanced transcript map of chickpea

Well-saturated linkage maps especially those based on expressed sequence tag (EST)-derived genic molecular markers (GMMs) are a pre-requisite for molecular breeding. This is especially true in important legumes such as chickpea where few simple sequence repeats (SSR) and even fewer GMM-based maps have been developed. Therefore, in this study, 2,496 ESTs were generated from chickpea seeds and utilized for the development of 487 novel EST-derived functional markers which included 125 EST-SSRs, 151 intron targeted primers (ITPs), 109 expressed sequence tag polymorphisms (ESTPs), and 102 single nucleotide polymorphisms (SNPs). Whereas EST-SSRs, ITPs, and ESTPs were developed by in silico analysis of the developed EST sequences, SNPs were identified by allele resequencing and their genotyping was performed using the Illumina GoldenGate Assay. Parental polymorphism was analyzed between C. arietinum ICC4958 and C. reticulatum PI489777, parents of the reference chickpea mapping population, using a total of 872 markers: 487 new gene-based markers developed in this study along with 385 previously published markers, of which 318 (36.5%) were found to be polymorphic and were used for genotyping. The genotypic data were integrated with the previously published data of 108 markers and an advanced linkage map was generated that contained 406 loci distributed on eight linkage groups that spanned 1,497.7 cM. The average marker density was 3.68 cM and the average number of markers per LG was 50.8. Among the mapped markers, 303 new genomic locations were defined that included 177 gene-based and 126 gSSRs (genomic SSRs) thereby producing the most advanced gene-rich map of chickpea solely based on co-dominant markers.