PCR cloning and detection of point mutations in the eburicol 14 alpha-demethylase (CYP51) gene from Erysiphe graminis f. sp. hordei, a "recalcitrant"fungus

Molecular studies of some micro-organisms are hampered by the difficulty of obtaining sufficient amounts of nucleic acids. A cloning strategy based on PCR has therefore been used to clone the eburicol 14 alpha-demethylase (CY P51) gene of the obligate fungus Erysiphe graminis f. sp. hordei (Egh) usin g minute amounts of,genomic DNA. The CYP51 gene encodes the enzymatic targe t of a major group of fungicides. Sequencing CYP51 from different Egh isola tes revealed the occurrence of two alleles for this gene.,An allele-specifi c PCR assay was developed to detect each CYP51 allele.