Project description:Purpose: Brassica. juncea is vulnerable to abiotic stresses at specific stages in its life cycle. However, till date no attempts have been made to elucidate the genome-wide changes in the transcriptome of B. juncea subjected to either high temperature or drought stress. Hence, to gain global insights into genes, transcription factors and kinases regulated by these stresses and to provide basic information on coding transcripts that are associated with traits of agronomic importance, we utilized a combinatorial approach of next generation sequencing and de novo assembly to discover B. juncea transcriptome associated with high temperature and drought. Results: We constructed and sequenced three transcriptome libraries namely Brassica control (BC), Brassica high temperature stress (BHS) and Brassica drought stress (BDS) from control, high temperature treated and drought treated seedlings of Brassica juncea. More than 180 million purity filtered reads were generated which were processed through quality parameters and high quality reads were assembled de-novo using SOAPde-novo assembler. A total of 77750 unique transcripts were identified out of which 69,245 (89%) were annotated with high confidence. We established a subset of 19110 transcripts, which were differentially regulated by either high temperature and/or drought stress. Furthermore, 886 and 2834 transcripts that code for transcription factors and kinases, respectively, were also identified. Investigation of identified transcription factors revealed that 92 responded to high temperature, 72 exhibited alterations in expression during drought stress, and 60 were commonly associated with both the stresses. Similarly, 217, 259 and 193 kinases were responsive to high temperature, drought or both stresses, respectively. Maximum number of up-regulated transcription factors in high temperature and drought stress belonged to heat shock factors (HSFs) and dehydration responsive element-binding (DREB) families respectively. We also identified 239 metabolic pathways, which were perturbed during high temperature and drought treatments. Analysis of gene ontologies associated with differentially regulated genes forecasted their involvement in diverse biological processes. Conclusions: Our study provides first comprehensive discovery of B. juncea transcriptome under high temperature and drought stress conditions. Transcriptome resources generated in this study will enhance our understanding on the molecular mechanisms involved in defining the response of B. juncea against two important abiotic stresses. Furthermore this information would benefit designing of efficient crop improvement strategies for tolerance against conditions of high temperature regimes and water scarcity. Total three RNA-Seq libraries were prepared and sequenced independently [B. juncea control (BC), B. juncea high temperature stressed (BHS) and B. juncea drought stressed (BDS) on Illumina GAIIx sequencer].

Project description:Purpose: Brassica. juncea is vulnerable to abiotic stresses at specific stages in its life cycle. However, till date no attempts have been made to elucidate the genome-wide changes in the transcriptome of B. juncea subjected to either high temperature or drought stress. Hence, to gain global insights into genes, transcription factors and kinases regulated by these stresses and to provide basic information on coding transcripts that are associated with traits of agronomic importance, we utilized a combinatorial approach of next generation sequencing and de novo assembly to discover B. juncea transcriptome associated with high temperature and drought. Results: We constructed and sequenced three transcriptome libraries namely Brassica control (BC), Brassica high temperature stress (BHS) and Brassica drought stress (BDS) from control, high temperature treated and drought treated seedlings of Brassica juncea. More than 180 million purity filtered reads were generated which were processed through quality parameters and high quality reads were assembled de-novo using SOAPde-novo assembler. A total of 77750 unique transcripts were identified out of which 69,245 (89%) were annotated with high confidence. We established a subset of 19110 transcripts, which were differentially regulated by either high temperature and/or drought stress. Furthermore, 886 and 2834 transcripts that code for transcription factors and kinases, respectively, were also identified. Investigation of identified transcription factors revealed that 92 responded to high temperature, 72 exhibited alterations in expression during drought stress, and 60 were commonly associated with both the stresses. Similarly, 217, 259 and 193 kinases were responsive to high temperature, drought or both stresses, respectively. Maximum number of up-regulated transcription factors in high temperature and drought stress belonged to heat shock factors (HSFs) and dehydration responsive element-binding (DREB) families respectively. We also identified 239 metabolic pathways, which were perturbed during high temperature and drought treatments. Analysis of gene ontologies associated with differentially regulated genes forecasted their involvement in diverse biological processes. Conclusions: Our study provides first comprehensive discovery of B. juncea transcriptome under high temperature and drought stress conditions. Transcriptome resources generated in this study will enhance our understanding on the molecular mechanisms involved in defining the response of B. juncea against two important abiotic stresses. Furthermore this information would benefit designing of efficient crop improvement strategies for tolerance against conditions of high temperature regimes and water scarcity.

Project description:Oilseed mustard, Brassica juncea, exhibits high levels of genetic variability for salinity tolerance. To obtain the global view of transcriptome and investigate the molecular basis of salinity tolerance in a salt-tolerant variety CS52 of B. juncea, we performed transcriptome sequencing of control and salt-stressed seedlings. De novo assembly of 184 million high-quality paired-end reads yielded 42,327 unique transcripts longer than 300 bp with RPKM ≥1. When compared with non-redundant proteins, we could annotate 67% unigenes obtained in our study. Based on the mapping to expressed sequence tags (ESTs), 52.6% unigenes are novel compared to EST data available for B. juncea and constituent genomes. Differential expression analysis revealed altered expression of 1469 unigenes in response to salinity stress. Of these, 587, mainly associated with ROS detoxification, sulfur assimilation and calcium signaling pathways, are up regulated. Notable of these is RSA1 (SHORT ROOT IN SALT MEDIUM 1) INTERACTING TRANSCRIPTION FACTOR 1 (RITF1) homolog up regulated by >100 folds in response to stress. RITF1, encoding a bHLH transcription factor, is a positive regulator of SOS1 and several key genes involved in scavenging of salt stress-induced reactive oxygen species (ROS). Further, we performed comparative expression profiling of key genes implicated in ion homeostasis and sequestration (SOS1, SOS2, SOS3, ENH1, NHX1), calcium sensing pathway (RITF1) and ROS detoxification in contrasting cultivars, B. juncea and B. nigra, for salinity tolerance. The results revealed higher transcript accumulation of most of these genes in B. juncea var. CS52 compared to salt-sensitive cultivar even under normal growth conditions. Together, these findings reveal key pathways and signaling components that contribute to salinity tolerance in B. juncea var. CS52. We report transcriptome sequencing of two-weeks-old seedlings of B. juncea var. CS52 under normal growth conditions (CTRL) and in response to salinity stress (SS) using Illumina paired-end sequencing

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