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|Title:||Evaluation and optimization of DNA extraction method for Dalbergia sissoo leaf|
|Authors:||Ginwal, H S|
Maurya, Shalini Singh
|Abstract:||A modified protocol for Dalbergia sissoo genomic DNA isolation has been optimized based on a cetyl trimethyl ammonium bromide (CTAB) method, described for other forest species. Leaves obtained from macro-propagated clones and mature trees of D. sissoo were tested. The method involves mortar grinding of tissue, a modified CTAB extraction buffer incorporating high salt concentrations, polyvinyl pyrrolidone and successive isoamyl alcohol-chloroform extractions with modified temperature conditions. The modification involved the use of doubled concentration of polyvinyl pyrrolidone (4% instead of 2%), increased incubation time with extraction buffer (40 min instead of 30 min), use of freshly prepared CTAB buffer and increased timing of washing of DNA pellet with wash buffer (45 min instead of 30 min). The yield was approximately 100 to 400 µg DNA per 100 mg of leaf tissue. The genomic DNA obtained by this method was suitable to be used in RAPD and ISSR analysis. This extraction method would allow the molecular analysis of DNA from different clones of D. sissoo.|
|ISSN:||0975-0967 (Online); 0972-5849 (Print)|
|Appears in Collections:||IJBT Vol.09(1) [January 2010]|
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