Standardization of DNA Isolation and PCR Protocol for RAPD Analysis of Suaeda sp.
The present study has evaluated the genetic variability among three salt marsh plant belongs to the family of Chenopodiaceae using random amplified polymorphic DNA markers. The method involves a modified Cetyl trimethylammonium bromide extraction, using polyvinyl pyrrolidone while grinding, successive long-term Chloroform: lsoamyalcohol extractions, an overnight RNase treatment with all steps carried out at room temperature. The yield of DNA ranged from 1.5-2.5 μg μL-1 per gram of the leaf tissue. The technique is ideal for isolation of DNA from different salt marsh plant species and used for Randomly Amplified Polymorphic DNA (RAPD) analysis. RAPD protocol was optimized based on the use of lower concentrations of primer (2 μM) and Taq polymerase (2 units), 50 ng of template DNA, higher concentration of MgCl2 (2 mM) and an annealing temperature of 37°C, resulted optimal amplification. The results suggest that the optimized protocol for DNA isolation and PCR was appropriate to diversity analysis of salt marsh plants.
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