Improving the yield and quality of DNA isolated from white-rot fungi.
A new simple method used to eliminate polysaccharides that cause problems during DNA isolation was established for 6 different white-rot fungi using 1% hexadecyltrimethylammonium bromide (CTAB) as wash buffer and followed by centrifugation. Variation in the DNA yield and quality was ascertained using precipitating agents, detergents and cell-wall-hydrolyzing chitinase. Considerable amount of exopolysaccharides from fungal biomass was removed with the use of 1% CTAB wash buffer followed by centrifugation. The DNA varied in terms of yield and quality. For the DNA extraction use of 2% SDS in extraction buffer worked best for Pycnoporus cinnabarinus, Cyathus bulleri, Cyathus striatus and Cyathus stercoreus, while 2% CTAB worked best for Phanerochaete chrysosporium and Pleurotus ostreatus. Elimination of phenol and use of absolute ethanol for precipitating DNA resulted in good yield and quality of DNA. This DNA was amenable to restriction endonuclease digestion.
Kuhad
R C
RC
Department of Microbiology, University of Delhi South Campus, New Delhi-110 021, India. kuhad@hotmail.com
Kapoor
R K
RK
Lal
R
R
D001487:Basidiomycota / Q000737:chemistry*; D000077286:Cetrimonium; D002593:Cetrimonium Compounds / Q000737:chemistry; D002688:Chitinases / Q000378:metabolism; D004262:DNA Restriction Enzymes / Q000378:metabolism; D004271:DNA, Fungal / Q000032:analysis / Q000302:isolation & purification* / Q000378:metabolism; D003902:Detergents; D005590:Fractional Precipitation; D009172:Mycology / Q000379:methods*; D020075:Phanerochaete / Q000737:chemistry; D019800:Phenol / Q000737:chemistry; D020076:Pleurotus / Q000737:chemistry; D011134:Polysaccharides / Q000737:chemistry
D002593:Cetrimonium Compounds; D004271:DNA, Fungal; D003902:Detergents; D011134:Polysaccharides; D019800:Phenol; D004262:DNA Restriction Enzymes; D002688:Chitinases; D000077286:Cetrimonium