Transcriptome sequencing of lentil based on second-generation technology permits large-scale unigene assembly and SSR marker discovery

lentilLentil (Lens culinaris Medik.) is a cool-season grain legume which provides a rich source of protein for human consumption. In terms of genomic resources, lentil is relatively underdeveloped, in comparison to other Fabaceae species, with limited available data. There is hence a significant need to enhance such resources in order to identify novel genes and alleles for molecular breeding to increase crop productivity and quality.

Tissue-specific cDNA samples from six distinct lentil genotypes were sequenced using Roche 454 GS-FLX Titanium technology, generating c. 1.38 x 106 expressed sequence tags (ESTs). De novo assembly generated a total of 15,354 contigs and 68,715 singletons. The complete unigene set was sequence-analysed against genome drafts of the model legume species Medicago truncatula and Arabidopsis thaliana to identify 12,639, and 7,476 unique matches, respectively. When compared to the genome of soybean (Glycine max), a total of 20,419 unique hits were observed corresponding to c. 31% of the known gene space. A total of 25,592 lentil unigenes were subsequently annotated from GenBank. Simple sequence repeat (SSR)-containing ESTs were identified from consensus sequences and a total of 2,393 primer pairs were designed. A subset of 192 EST-SSR markers was screened for validation across a panel 12 cultivated lentil genotypes and one wild relative species. A total of 166 primer pairs obtained successful amplification, of which 47.5% detected genetic polymorphism.

A substantial collection of ESTs has been developed from sequence analysis of lentil genotypes using second-generation technology, permitting unigene definition across a broad range of functional categories. As well as providing resources for functional genomics studies, the unigene set has permitted significant enhancement of the number of publicly-available molecular genetic markers as tools for improvement of this species.

Sukhjiwan Kaur, Noel O.I. Cogan, Luke W. Pembleton, Maiko Shinozuka, Keith W. Savin, Michael Materne and John W. Forster. (2011) Transcriptome sequencing of lentil based on second-generation technology permits large-scale unigene assembly and SSR marker discovery. BMC Genomics 12, 265. [abstract]