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Authors: Pardeep Kumar
Advisor: Yadav, Neelam R.
Title: Expression of myb gene in brassica tournefortii L.under drought stress
Publisher: CCSHAU
Language: en
Type: Thesis
Agrotags: Fruits, Genotypes, dna, Acidity, Rapd, Limes, Genetics, Confectionery, Polymorphism, Developmental stages
Abstract: Brassica tournefortii, a highly drought tolerant Brassica species was used to study myb gene expression under drought stress. Seeds of Brassica tournefortii were grown on MS medium. Then 14 days seedlings were uprooted and given drought stress by subjecting them to air drying, PEG (-2 to -8 bar) and mannitol (100 mM to 400 mM) treatments. Total RNA isolation from stressed seedlings was carried out using Trizol reagent yielding 17.52-36.8 Fg/ml of total RNA. One- step RT-PCR was carried out using total RNA as template with BjMyb-1 primer designed from conserved domain of AtMyb2 and its homologous. BjActin primers were used in RT-PCR which served as control, as actin gene is constitutively expressed in all tissues. Exposure to drought stress for 15 minutes and 30 minutes (air drying) gave no amplification showing air drying upto 30 minutes does not induce any myb expression. An amplified product of 250 bp was obtained on exposure to drought stress for 60 minutes with air drying, PEG (-2 to -8 bar) and mannitol (100 mM to 400 mM) treatments. The transcript level was found similar in all treatments irrespective of drought treatments. cDNA was eluted out from the gel and purified cDNA was transformed using pDrive cloning vector (Quigen) in XL-blue strain of E.coli using blue- white selection. Transformed clones were characterized by plasmid DNA isolation, PCR amplification of plasmid DNA with gene specific primers. Plasmid DNA from transformed clones showed higher molecular weight than untransformed plasmid DNA on agarose gel electrophoresis confirmed the insertion of DNA fragment into the plasmid. PCR amplification of plasmid DNA also confirmed the successful cloning.
Issue Date: 2009
Appears in Collections:M. Sc. Dissertations

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