%0 Journal Article
%@ 14712229
%A Dutta, S
%A Kumawat, G
%A Singh, B P
%A Gupta, D K
%A Singh, Sangeeta
%A Dogra, V
%A Gaikwad, K
%A Sharma, T R
%A Raje, R S
%A Bandhopadhya, T K
%A Datta, S
%A Singh, M N
%A Bashasab, F
%A Kulwal, P
%A Wanjari, K B
%A Varshney, R K
%A Cook, D R
%A Singh, N K
%D 2011
%F icrisat:12
%I BioMed Central
%J BMC Plant Biology
%N 1
%P 17-29
%T Development of genic-SSR markers by deep transcriptome sequencing in pigeonpea [Cajanus cajan (L.) Millspaugh]
%U http://oar.icrisat.org/12/
%V 11
%X Background: Pigeonpea [Cajanus cajan (L.) Millspaugh], one of the most important food legumes of semi-arid  tropical and subtropical regions, has limited genomic resources, particularly expressed sequence based (genic)  markers. We report a comprehensive set of validated genic simple sequence repeat (SSR) markers using deep  transcriptome sequencing, and its application in genetic diversity analysis and mapping.  Results: In this study, 43,324 transcriptome shotgun assembly unigene contigs were assembled from 1.696 million  454 GS-FLX sequence reads of separate pooled cDNA libraries prepared from leaf, root, stem and immature seed of  two pigeonpea varieties, Asha and UPAS 120. A total of 3,771 genic-SSR loci, excluding homopolymeric and  compound repeats, were identified; of which 2,877 PCR primer pairs were designed for marker development.  Dinucleotide was the most common repeat motif with a frequency of 60.41%, followed by tri- (34.52%), hexa-  (2.62%), tetra- (1.67%) and pentanucleotide (0.76%) repeat motifs. Primers were synthesized and tested for 772 of  these loci with repeat lengths of ≥18 bp. Of these, 550 markers were validated for consistent amplification in eight  diverse pigeonpea varieties; 71 were found to be polymorphic on agarose gel electrophoresis. Genetic diversity  analysis was done on 22 pigeonpea varieties and eight wild species using 20 highly polymorphic genic-SSR  markers. The number of alleles at these loci ranged from 4-10 and the polymorphism information content values  ranged from 0.46 to 0.72. Neighbor-joining dendrogram showed distinct separation of the different groups of  pigeonpea cultivars and wild species. Deep transcriptome sequencing of the two parental lines helped in silico  identification of polymorphic genic-SSR loci to facilitate the rapid development of an intra-species reference  genetic map, a subset of which was validated for expected allelic segregation in the reference mapping  population.  Conclusion: We developed 550 validated genic-SSR markers in pigeonpea using deep transcriptome sequencing.  From these, 20 highly polymorphic markers were used to evaluate the genetic relationship among species of the  genus Cajanus. A comprehensive set of genic-SSR markers
%Z This study was financially supported by the Pigeonpea Genomics Initiative  (PGI) of the Indian Council of Agricultural Research (ICAR), New Delhi under  the framework of Indo-US Agricultural Knowledge Initiative (AKI).  Contribution of Doug Cook was supported by the National Science  Foundation (NSF), USA. GK acknowledges fellowship support from  Department of Biotechnology, Government of India.