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SSH analysis of endosperm transcripts and characterization of heat stress regulated expressed sequence tags in bread wheat

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Title SSH analysis of endosperm transcripts and characterization of heat stress regulated expressed sequence tags in bread wheat
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Creator Suneha Goswami
Ranjeet R. Kumar
Kavita Dubey
Jyoti P. Singh
Sachidanand Tiwari
Ashok Kumar
Shuchi Smita
Dwijesh C. Mishra
Sanjeev Kumar
Monendra Grover
Jasdeep C. Padaria
Yugal K. Kala
Gyanendra P. Singh
Himanshu Pathak
Viswanathan Chinnusamy
Anil Rai
Shelly Praveen
Raj D. Rai
 
Subject Abiotic stress
Differential expression
SSH library
Terminal heat stress
Triticum aestivum
SAGs
DEGs
Differentially expressed proteins (DEPs)
 
Description Not Available
Heat stress is one of the major problems in agriculturally important cereal crops, especially wheat. Here, we have constructed a subtracted cDNA library from the endosperm of HS-treated (42°C for 2 h) wheat cv. HD2985 by suppression subtractive hybridization (SSH). We identified ~550 recombinant clones ranging from 200 to 500 bp with an average size of 300 bp. Sanger's sequencing was performed with 205 positive clones to generate the differentially expressed sequence tags (ESTs). Most of the ESTs were observed to be localized on the long arm of chromosome 2A and associated with heat stress tolerance and metabolic pathways. Identified ESTs were BLAST search using Ensemble, TriFLD, and TIGR databases and the predicted CDS were translated and aligned with the protein sequences available in pfam and InterProScan 5 databases to predict the differentially expressed proteins (DEPs). We observed eight different types of post-translational modifications (PTMs) in the DEPs corresponds to the cloned ESTs-147 sites with phosphorylation, 21 sites with sumoylation, 237 with palmitoylation, 96 sites with S-nitrosylation, 3066 calpain cleavage sites, and 103 tyrosine nitration sites, predicted to sense the heat stress and regulate the expression of stress genes. Twelve DEPs were observed to have transmembrane helixes (TMH) in their structure, predicted to play the role of sensors of HS. Quantitative Real-Time PCR of randomly selected ESTs showed very high relative expression of HSP17 under HS; up-regulation was observed more in wheat cv. HD2985 (thermotolerant), as compared to HD2329 (thermosusceptible) during grain-filling. The abundance of transcripts was further validated through northern blot analysis. The ESTs and their corresponding DEPs can be used as molecular marker for screening or targeted precision breeding program. PTMs identified in the DEPs can be used to elucidate the thermotolerance mechanism of wheat—a novel step toward the development of “climate-smart” wheat.
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Date 2018-02-22T01:56:37Z
2018-02-22T01:56:37Z
2016-08-17
 
Type Research Paper
 
Identifier Not Available
Not Available
http://krishi.icar.gov.in/jspui/handle/123456789/5842
 
Language English
 
Relation Not Available;
 
Publisher PMC, National Center for Biotechnology Information