A modified ctab method for efficient extraction of genomic dna in indian sweet sorghum (Sorghum bicolor. l. moench)

Krishnamurthy Sathelly, Rajani Govindannagari, Saurabh Pandey, Sai Kumar Kompelli, Visarada K. B. R. S., Tanushri Kaul*

Abstract


Isolation of high quality, intact high molecular weight genomic DNA from plants which are rich in polysaccharides, polyphenols, secondary metabolites and chemical heterogeneity is an immense problem in the field of plant biology. Several protocols have been developed for eliminating these tricky elements during the extraction of DNA, but none is found to be universally applicable. The purpose of the present study was to develop a reliable protocol for extracting high quality genomic DNA from polyphenols, polysaccharides and secondary metabolites rich plants like sweet sorghum. We made seven critical modifications to the available CTAB method to isolate genomic DNA from 25- and 60-d-old transgenic sweet sorghum leaf tissues. The yield of DNA ranged from 9.2– 10.2 µg from 200 mg of leaf tissue. An absorbance value of 1.8 at A260/A280 indicates that it’s free from RNA and protein contamination. PCR analysis using bar primers shows a consistent and reliable amplification product at 475 bp. This method is highly suitable for extracting high quality genomic DNA from plants with high levels of polysaccharides and polyphenolics without blending commercial kits. Our protocol facilitates the processing of large number of plant samples for genomic analysis, mapping and next generation sequencing. 


Keywords


Cereal; Ethanol; Genomic DNA; NaCl; Polysaccharides; Polyphenols

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