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Development of genic-SSR markers by deep transcriptome sequencing in pigeonpea [Cajanus cajan (L.) Millspaugh]

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Relation http://oar.icrisat.org/12/
http://dx.doi.org/10.1186/1471-2229-11-17
 
Title Development of genic-SSR markers by deep transcriptome sequencing in pigeonpea [Cajanus cajan (L.) Millspaugh]
 
Creator Dutta, S
Kumawat, G
Singh, B P
Gupta, D K
Singh, Sangeeta
Dogra, V
Gaikwad, K
Sharma, T R
Raje, R S
Bandhopadhya, T K
Datta, S
Singh, M N
Bashasab, F
Kulwal, P
Wanjari, K B
Varshney, R K
Cook, D R
Singh, N K
 
Subject Pigeonpea
 
Description 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
 
Publisher BioMed Central
 
Date 2011
 
Type Article
PeerReviewed
 
Format application/pdf
 
Language en
 
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Identifier http://oar.icrisat.org/12/1/varsh-2011-aupay.pdf
Dutta, S and Kumawat, G and Singh, B P and Gupta, D K and Singh, Sangeeta and Dogra, V and Gaikwad, K and Sharma, T R and Raje, R S and Bandhopadhya, T K and Datta, S and Singh, M N and Bashasab, F and Kulwal, P and Wanjari, K B and Varshney, R K and Cook, D R and Singh, N K (2011) Development of genic-SSR markers by deep transcriptome sequencing in pigeonpea [Cajanus cajan (L.) Millspaugh]. BMC Plant Biology, 11 (1). pp. 17-29. ISSN 14712229