Exploring drought stress-regulated genes in senna (Cassia angustifolia Vahl.): a transcriptomic approach

Abstract

De novo assembly of reads produced by next-generation sequencing (NGS) technologies offers a rapid approach to obtain expressed gene sequences for non-model organisms. Senna (Cassia angustifolia vahl.), is drought tolerant annual under shrub of Caesalpiniaceae, a sub family of Fabaceae. There are insufficient transcriptomic and genomic data in public databases for understanding of the molecular mechanism underlying the drought tolerance of Senna. Therefore, the main purpose of this study is to know the transcriptome profile of senna, with special reference to drought stress. RNA from two different stages of leaf development was extracted and sequenced separately using the Illumina technology. A total of 200 million reads were generated and de novo assembly of processed reads in the pooled transcriptome using Trinity yielded 43,413 transcripts which were further annotated using NCBI BLAST with ‘green plant database (txid 33090)’, Swiss Prot, Kyoto encyclopedia of genes & genomes (KEGG), Cluster of orthologous gene (COG) and Gene Ontology (GO). Out of the total transcripts, 42,280 (95.0%) were annotated by BLASTX against green plant database of NCBI. Senna transcriptome showed highest similarity to Glycine max (41%), followed by Phaseolus vulgaris (16%), Cicer arietinum (15%), and Medicago trancatula (5%). Highest number of GO terms were enriched for molecular functions category of these “catalytic activity” (GO: 0003824) (25.10%) and “binding activity” (GO: 0005488) (20.10%) were most abundantly represented. We used InterProscan to see protein similarity at domain level, a total of 33,256 transcripts were annotated against the Pfam domains. The transcripts were assigned to various KEGG pathways. Coding DNA sequences (CDS) encoding various drought stress regulated pathways such as signaling factors, protein modifying/degrading enzymes, biosynthesis of phytoharmone, phytoharmone signaling, osmatically active compounds, free radical scavengers, chlorophyll metabolism, leaf cuticular wax, polyamines and protective proteins were identified through BLASTX search. Lucine rich repeat kinase family was the most abundantly found group of protein kinases. Orphan, bHLH and bZIP family TFs were the most abundantly found in senna. Six genes encoding MYC2 transcription factor, 9-cis-epoxycarotenoid dioxygenase (NCED), L-ascorbate peroxidase (APX), aminocyclopropane carboxylate oxidase (ACO), Abscisic acid 8'-hydroxylase (ABA) and WRKY transcription factor were confirmed through Reverse Transcriptase-PCR (RT-PCR) and Sanger sequencing for the first time in Senna. The potential drought stress related transcripts identified in this study provide a good start for further investigation into the drought adaptation in senna. Additionally, our transcriptome sequences are the valuable resource for accelerated genomics-assisted genetic improvement programs and facilitate a better understanding and more effective manipulation of biochemical pathways for developing drought tolerant crop plants.