Development of chickpea EST-SSR markers and analysis of allelic variation across related species

Abstract

Despite chickpea being the third important grain legume, there is a limited availability of genomic resources, especially of the expressed sequence tag (EST)-based markers. In this study, we generated 822 chickpea ESTs from immature seeds as well as exploited 1,309 ESTs from the chickpea database, thus utilizing a total of 2,131 EST sequences for development of functional EST-SSR markers. Two hundred and forty-six simple sequence repeat (SSR) motifs were identified from which 183 primer pairs were designed and 60 validated as functional markers. Genetic diversity analysis across 30 chickpea accessions revealed ten markers to be polymorphic producing a total of 29 alleles and an observed heterozygosity average of 0.16 thereby exhibiting low levels of intra-specific polymorphism. However, the markers exhibited high cross-species transferability ranging from 68.3 to 96.6% across the six annual Cicer species and from 29.4 to 61.7% across the seven legume genera. Sequence analysis of size variant amplicons from various species revealed that size polymorphism was due to multiple events such as copy number variation, point mutations and insertions/deletions in the microsatellite repeat as well as in the flanking regions. Interestingly, a wide prevalence of crossability-group-specific sequence variations were observed among Cicer species that were phylogenetically informative. The neighbor joining dendrogram clearly separated the chickpea cultivars from the wild Cicer and validated the proximity of C. judaicum with C. pinnatifidum. Hence, this study for the first time provides an insight into the distribution of SSRs in the chickpea transcribed regions and also demonstrates the development and utilization of genic-SSRs. In addition to proving their suitability for genetic diversity analysis, their high rates of transferability also proved their potential for comparative genomic studies and for following gene introgressions and evolution in wild species, which constitute the valuable secondary genepool in chickpea.

Developing new SSR markers from ESTs of pea (Pisum sativum L.)

“…The SSR markers from Medicago truncatula exhibited transferability to faba bean, chickpea, and pea (Gutierrez et al, 2005). The EST-SSR markers from chickpea also showed high transferability to the six Cicer species and seven legume genera (P. mungo, P. sativum, G. max, T. alexandrinum, L. esculenta, C. cajan, and M. truncatula) (Choudhary et al, 2009). The present results show that three loci of nine polymorphic P. sativum EST-SSR markers can be amplified in faba bean, which suggests that it may assist in reducing the costs of marker development, and promote genetic analysis among the bean species.…”

Cross-genera amplification of informative microsatellite markers from common bean and lentil for the assessment of genetic diversity in pigeonpea

“…Despite its importance in nutritional security, pigeonpea has not benefited much from the advances made in the field of genomic tools and molecular markers (Datta et al, 2009). As compared to large number of markers in common bean (L'taief et al, 2008;Blair et al, 2003), chickpea (Choudhary et al, 2009;Sethy et al, 2006), and lentil (Hamwieh et al, 2009;Hamwieh et al, 2005), only 109 microsatellite markers are reported so far in pigeonpea (Burns et al, 2001;Odeny et al, 2007;Odeny et al, 2009). Therefore, there is an urgent need to increase the number of polymorphic microsatellite markers in pigeonpea for diversity analysis and mapping of important traits.…”

Genomic tools and germplasm diversity for chickpea improvement

“…The neighbor joining dendrogram clearly separated the chickpea cultivars from the wild Cicer and validated the proximity of C. judaicum Choudhary et al, 2009 10 SSR markers 47 chickpea (C. arietinum) accessions including 21 induced mutation lines, 17 hybrid lines, 5 local cultigens, and 4 non-nodulating lines UPGMA and ME (minimum evolution) trees classified the accessions into 6 groups and all but 6 accessions could be clearly separated.…”