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|Title:||A simplified high yielding miniprep genomic DNA extraction protocol for three chemotypically different plant species|
Chattopadhyay, S K
Ghosh, P D
|Abstract:||The presence of metabolites has been observed to create interference with DNA isolation procedures and downstream reactions, such as, DNA amplification, restriction and cloning. Moreover, the chemotypic heterogeneity among species may cause hindrance in optimal DNA yields with a single protocol. Thus, even closely related species may require different isolation protocols. A rapid and high yielding DNA extraction procedure from heterogeneous plant tissues of 3 chemotypic different plants has been presented here. This modified method required grinding of plant tissues in DNA extraction buffer [200 mM Tris-HCL (pH 8.0), 200 mM NaCl and 25 mM EDTA, and 1% PVP], followed by phenol:chloroform:isoamyl alcohol (25:24:1, v/v/v) treatment. Ethanol in the presence of high salt concentration was then used to precipitate the DNA and for removal of polysaccharides. The average yield of DNA ranged from 1.68 to 5.37 mg g-1 of plant tissue and the purity was 1.6 to 1.9. The DNA was quite suitable for PCR using RAPD and also for restriction digestion. This method did not require liquid nitrogen for fixation, RNase treatment or storage at –80°C, making it advantageous over other common protocols.|
|ISSN:||0975-0967 (Online); 0972-5849 (Print)|
|Appears in Collections:||IJBT Vol.11(3) [July 2012]|
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