A RABID METHOD FOR THE ISOLATION OF GENOMIC DNA FROM ASPERGILLUS-FUMIGATUS

A majority of Aspergillus induced diseases are reported to be caused b y Aspergillus fumigatus. In immunocompromized and post transplant case s it can lead to invasive aspergillosis. Due to this the molecular fin gerprinting of aspergillus isolates by RFLP analysis and development o f DNA diagnostic probes are gaining importance. Different methodologie s are being adopted for extraction of the genomic DNA from fungus. The existing procedures for isolation of DNA are time consuming and range from several hours to few days. The most difficult step in the isolat ion of DNA from aspergillus species is to disrupt the tough chitin ric h cell wall, without causing damage to genomic DNA. We report here a r apid method for extraction of genomic DNA based on the cleavage of chi tin with chitinase. The subsequent modification steps included are lys is and microwave treatment. The chromosomal DNA obtained by this proce dure is 1.5 - 2.0 mu g per mg of wet weight of mycelia and is observed to be minimally sheared. It is pure enough for restriction analysis a nd for use in the PCR to detect the gene coding for 18 kDa allergen wh ich has been identified in our laboratory using western blot analysis with human patient sera.