Project description:In low rainfall regions soils are naturally conditioned with frequent co-occurrence of salinity and alkalinity. Plant salinity responses both at physiological and molecular level have been extensively researched. However, effects of the combined treatment of alkaline salinity that could greatly reduce plant growth and the mechanisms responsible for tolerance remain indeterminate. In Brassica juncea, large reductions in biomass and increased leaf Na+ concentration under alkaline salinity indicates that the combined treatment had greater negative effect than salinity on both growth and the physiological responses of the plant. To determine molecular mechanisms potentially controlling adaptive tolerance responses to salinity and alkaline salinity, the moderately tolerant genotype NDR 8501 was further investigated using microarray analysis. The transcripts of treated leaf tissues verses those of the untreated control sample were analysed after prolonged stress of four weeks. In total, 528 salinity responsive and 1245 alkaline salinity responsive genes were indentified and only 101 genes were expressed jointly in either of the two treatments. Transcription of 37% more genes involved in response to alkaline salinity than salinity alone, which suggests the increased impact and severity of the combined stress on the plant, indicating the transcription of a far greater number of genes likely involved in mitigation and damage control. Transcription of KUP2 and KUP7 genes involved in potassium homeostasis under salinity alone and NHX1 and ENH1 genes for ion (K+ and Na+) homeostasis under alkaline salinity, clearly demonstrated that different genes and genetic pathways are involved in response to each stress. They further provide supporting evidence for the physiological responses that occur in the plant, with massive reprogramming of the transcriptome leading to partial ion exclusion, shuttling and compartmentation. Salinity and alkaline salinity induced gene expression in Brassica juncea leaf was measured at 4 weeks of prolonged treatment of 50 mM NaCl alone and combined with 2.5 mM HCO3- versus non-stressed control. A single experiment was conducted using Brassica juncea genotype NDR 8501 at a single time point (fours weeks after treatment) with three replications per treatment.

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:Purpose: To identify high temperature, sailinity and drought-responsive miRNAs at genome wide level in B. juncea var varuna. Results: In this study four small RNA libraries viz. B. juncea control (BJC), B. juncea high temperature stressed (BJH), B. juncea salinity stressed (BJS) and B. juncea drought stressed (BJD) were prepared and sequenced. With the help of UEA sRNA workbench software package 51 conserved miRNAs belonging to 30 miRNA families were identified. As there was limited genomic information available for B. juncea, we generated and assembled its genome sequence at a very low coverage. Using the generated sequence and other publically available Brassica genomic/transcriptomic resources as mapping reference, 126 novel (not reported so far in any plant species) were discovered for the first time in B. juncea. Further analysis also revealed existence of 32 and 37 star sequences for conserved and novel miRNAs, respectively. The expression of a few selected conserved and novel miRNAs under conditions of different abiotic stresses was revalidated through universal TaqMan based real time PCR. Putative targets of identified conserved and novel miRNAs were predicted in B. rapa to gain insights into functional roles manifested by B. juncea miRNAs. Furthermore, SPL2-like, ARF17-like and a NAC domain containing protein were experimentally validated as targets of miR156, miR160 and miR164 respectively. Investigation of gene ontologies linked with targets of known and novel miRNAs forecasted their involvement in various biological functions. Conclusions: We have generated in this study, the first comprehensive abiotic stress influenced small RNA dataset in B. juncea. The combinatorial approach of NGS and computational methods led to the discovery of 51 conserved and 126 novel miRNAs. The present study provides a holistic view of B. juncea miRNAome under conditions of high temperature, salinity and drought. The catalogue of miRNA sequences, their expression and putative targets, generated in this study can be utilized to design crop improvement strategies in B. juncea and related species.

Project description:Purpose: To identify high temperature, sailinity and drought-responsive miRNAs at genome wide level in B. juncea var varuna. Results: In this study four small RNA libraries viz. B. juncea control (BJC), B. juncea high temperature stressed (BJH), B. juncea salinity stressed (BJS) and B. juncea drought stressed (BJD) were prepared and sequenced. With the help of UEA sRNA workbench software package 51 conserved miRNAs belonging to 30 miRNA families were identified. As there was limited genomic information available for B. juncea, we generated and assembled its genome sequence at a very low coverage. Using the generated sequence and other publically available Brassica genomic/transcriptomic resources as mapping reference, 126 novel (not reported so far in any plant species) were discovered for the first time in B. juncea. Further analysis also revealed existence of 32 and 37 star sequences for conserved and novel miRNAs, respectively. The expression of a few selected conserved and novel miRNAs under conditions of different abiotic stresses was revalidated through universal TaqMan based real time PCR. Putative targets of identified conserved and novel miRNAs were predicted in B. rapa to gain insights into functional roles manifested by B. juncea miRNAs. Furthermore, SPL2-like, ARF17-like and a NAC domain containing protein were experimentally validated as targets of miR156, miR160 and miR164 respectively. Investigation of gene ontologies linked with targets of known and novel miRNAs forecasted their involvement in various biological functions. Conclusions: We have generated in this study, the first comprehensive abiotic stress influenced small RNA dataset in B. juncea. The combinatorial approach of NGS and computational methods led to the discovery of 51 conserved and 126 novel miRNAs. The present study provides a holistic view of B. juncea miRNAome under conditions of high temperature, salinity and drought. The catalogue of miRNA sequences, their expression and putative targets, generated in this study can be utilized to design crop improvement strategies in B. juncea and related species. Total four small RNA libraries were prepared and sequenced independently [B. juncea control (BJC), B. juncea high temperature stressed (BJH), B. juncea salinity stressed (BJS) and B. juncea drought stressed (BJD)] on Illumina GAIIx

			Action	DRS
	SRR5469110_1.fastq.gz	Fastqsanger.gz
	SRR5469110_2.fastq.gz	Fastqsanger.gz
	SRR5469120_1.fastq.gz	Fastqsanger.gz
	SRR5469120_2.fastq.gz	Fastqsanger.gz
	SRR5469130_1.fastq.gz	Fastqsanger.gz

Dataset Information

Brassica juncea cultivar:Pusa Bold and Pusa Jai Kisan

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