Abstract
Black gram [V. mungo (L.) Hepper] is an important legume crop extensively grown in south and south-east Asia, where it is a major source of dietary protein for its predominantly vegetarian population. However, lack of genomic information and markers has become a limitation for genetic improvement of this crop. Here, we report the transcriptome sequencing of the immature seeds of black gram cv. TU94-2, by Illumina paired end sequencing technology to generate transcriptome sequences for gene discovery and genic-SSR marker development. A total of 17.2 million paired-end reads were generated and 48,291 transcript contigs (TCS) were assembled with an average length of 443 bp. Based on sequence similarity search, 33,766 TCS showed significant similarity to known proteins. Among these, only 29,564 TCS were annotated with gene ontology (GO) functional categories. A total number of 138 unique KEGG (Kyoto Encyclopedia of Genes and Genomes) pathways were identified, of which majority of TCS are grouped into purine metabolism (678) followed by pyrimidine metabolism (263). A total of 48,291 TCS were searched for SSRs and 1,840 SSRs were identified in 1,572 TCS with an average frequency of one SSR per 11.9 kb. The tri-nucleotide repeats were most abundant (35%) followed by di-nucleotide repeats (32%). PCR primer pairs were successfully designed for 933 SSR loci. Sequences analyses indicate that about 64.4% and 35.6% of the SSR motifs were present in the coding sequences (CDS) and untranslated regions (UTRs) respectively. Tri-nucleotide repeats (57.3%) were preferentially present in the CDS. The rate of successful amplification and polymorphism were investigated using selected primers among 18 black gram accessions. Genic-SSR markers developed from the Illumina paired end sequencing of black gram immature seed transcriptome will provide a valuable resource for genetic diversity, evolution, linkage mapping, comparative genomics and marker-assisted selection in black gram.
Highlights
Black gram [V. mungo (L.) Hepper] is an important legume crop in Asia, where it is a major source of dietary protein for its predominantly vegetarian population
India is the largest producer of black gram, where about 3.26 million hectares are cultivated with a production of 1.74 million tonnes during 2010–11 [1]
transcript contigs (TCS) with length more than 500 bp accounted for 24.1% (11,632 contigs)
Summary
Black gram [V. mungo (L.) Hepper] is an important legume crop in Asia, where it is a major source of dietary protein for its predominantly vegetarian population. It is extensively grown in south and south-east Asia. The average yield of black gram is low due to its indeterminate growth habit, non-synchronous maturity, narrow genetic base, and losses due to biotic and abiotic stresses. Black gram genetic improvement programs have made relatively little progress in addressing the primary constraints to crop production, which include a range of biotic (e.g. yellow mosaic disease, powdery mildew, pod borer and bruchids) and abiotic (e.g. drought and salinity) stresses. Complete genome sequence information of black gram is not available as on date and the use of genomic resources in this crop largely depend on the sequence information available in the closely related taxa
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