Targeted mining of drought stress-responsive genes from EST resources in Cleistogenes songorica

► Our findings illustrate the potential for targeted functional genomic studies in non-model, non-crop plant species, particularly in extremophiles, to identify novel or enhanced stress tolerance mechanisms. ► A gene discovery and transcript abundance profiling study has been undertaken in Cleistogenes songorica, a grass that thrives in severely water depleted environments in Inner Mongolia. ► Amongst an EST collection from drought stressed plants are many sequences with roles in response to drought stress. ► Transcript abundance profiling revealed that the majority of genes tested were regulated by drought. ► These findings offer a functional insight into the molecular mechanisms used by C. songorica to cope with moisture limitation, and identify potential resources for conferring drought tolerance in sensitive plants. Cleistogenes songorica is an important perennial grass found in the pastoral steppe of Inner Mongolia. C. songorica flourishes in drought prone environments, and therefore provides an ideal candidate plant system for the identification of drought-tolerance conferring genes. We constructed cDNA libraries from leaves and roots of drought-stressed C. songorica seedlings. Expressed sequence tag (EST) sequencing of 5664 random cDNA clones produced 3579 high quality, trimmed sequences. The average read length of trimmed ESTs was 613 bp. Clustering and assembly identified a non-redundant set of 1499 contigs, including 805 singleton unigenes and 694 multi-member unigenes. The resulting unigenes were functionally categorized according to the Gene Ontology (GO) hierarchy using the in house Bioinformatic Advanced Scientific Computing (BASC) annotation pipeline. Among the total 2.2 Mbp of EST sequence data, 161 putative SSRs were found, a frequency similar to that previously observed in oat and Arabidopsis ESTs. Sixty-three unigenes were functionally annotated as being stress responsive, of which 22 were similar to genes implicated in drought stress response. Using quantitative real time RT-PCR, transcripts of 13 of these 22 genes were shown to be at least three fold more, or less abundant in drought-stressed leaves or roots, with 8 increased and 5 decreased in relative transcript abundance. The C. songorica EST and cDNA collections generated in this study are a valuable resource for microarray-based expression profiling, and functional genomics in order to elucidate their role, and to understand the underlying mechanisms of drought-tolerance in C. son