ISOLATION AND CHARACTERIZATIN OF A GENE SEQUENCE ENCODING A TRANSCRIPTION FACTOR INVOLVED IN ABA DEPENDENT SIGNAL TRANSDUCTION
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Date
2009
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IARI, DIVISION OF BIOCHEMISTRY
Abstract
Abscisic acid (ABA), a plant stress hormone, plays a key role in regulation of
sub-set of stress responsive genes. These genes respond to ABA through specific
transcription factors which bind to cis-regulatory elements present in their promoters. In
this study, ABA was quantified at different developmental stages of Oryza sativa
cultivar N22 from seeds as well as leaves. ABA level in seeds at 30 DAF was
approximately double as compared to the seeds at 20 DAF which shows that ABA is
accumulated in the developing seeds at a very fast rate. Increased levels of ABA were
observed in the leaf tissues under water deficit stress conditions as compared to control
at three different developmental stages namely 10, 25 and 40 DAP. Specific primers of
three different transcription factor families namely WRKY, bZIP and MYB were used for
PCR amplification of genomic DNA. The size of amplified products of WRKY, bZIP and
MYB genes were ~280bp, ~800bp and ~1050bp respectively. cDNA prepared from total
RNA of drought stressed leaf tissues was amplified using WRKY, bZIP and MYB gene
primers. A specific ~280 bp fragment was obtained with WRKY specific gene primers
only. The same size of amplicon obtained from genomic DNA and cDNA suggests the
absence of intron in the WRKY transcription factor gene. The amplified product was
cloned in pGEMT vector and used for transforming DH5α cells. Sequence analysis of
positive clones predicted that the gene contains an ORF of 186 bp with 5’ and 3’ UTR of
49 and 43 nucleotides respectively and encodes a protein of 61 amino acid residues. RTPCR
analysis revealed an upregulated expression of WRKY transcription factor gene in
ABA treated plants as compared to control. Hence, it can be predicted that the
expression of this gene is induced by ABA
Description
T-8121