What is the best method to isolate/extract fungal genomic DNA? I have tried several methods without the addition of chitinase, but I wasn't successful in this aspect. Please suggest a better method to extraction of fungal genomic DNA for PCR amplification without addition of chitinase.
Thank you
We usually follow a protocol by which we mix a 10ml ON culture pellet with a lysis buffer with detergents, phenol/chlorophorm and glass beads, and we vortex the whole mixture thoroughly. Then, we add TE, vortex again and spin down; we precipitate the aqueous layer with ethanol, wash it and resuspend the DNA pellet in TE with RNase. Hope this is useful!
use the method described in this paper ( Bollet, C., Gevaudan, M.J., de Lamballerie, X., Zandotti, C.and de Micco, P. 1991. A simple method for the isolation of chromosomal DNA from Gram positive or acid-fast bacteria. Nucleic Acids Res, 19, 1955.) I found it useful for fungi
We have use CETAB treatment followed by standar phenol/chrophorm procedure with no problem in C. neoformans, filamentous fungi and yeast.
CETAB or washing detergent can be use. you can find any paper that explains this procedures as suggested above
I have used the protocol below with some modifications (e.g. using LN) with good results, i.e. DNA suitable for genome sequencing.
Article DNA Extraction Protocol for Plants with High Levels of Secon...
CTAB method followed by Phenol:chloriform can be used for isolation of genomic DNA from fungi
If you want to use a kit you can use the qiagen plant kit mini. At the lysis step you need to disrupt the hyphae either with a a fast prep disruption step or liquid nitrogen and a pestle and mortar
CETAB treatment followed by standar phenol/chrophorm protocol works better for fungal genomic DNA isolation
Could you please send me the CETAB protocol, my email: [email protected]
Thanks in advance
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