Chemistry: molecular biology and microbiology – Measuring or testing process involving enzymes or... – Involving nucleic acid
Patent
1993-08-16
1997-01-07
Wax, Robert A.
Chemistry: molecular biology and microbiology
Measuring or testing process involving enzymes or...
Involving nucleic acid
435 25, 435 29, 435 32, 43525421, 4352544, 4353201, C12N 1563, C12Q 102, C12Q 126, C12Q 168
Patent
active
055915760
ABSTRACT:
A binary assay identifies agents that inhibit sterol .DELTA.14 reductase involved in ergosterol biosynthesis. In the primary screen, sterol .DELTA.14 reductase inhibition by a test sample is assayed by adding the test sample to a culture of Neurospora crassa having an erg-3 mutation and also to a culture of a strain having an erg-1 mutation, comparing the extent of growth inhibition after incubation in the two cultures, and identifying as positives those samples that show growth inhibition in the erg-3 culture exceeding that in the erg-1 culture. In the secondary screen, samples that test positive in the primary screen are reassayed by adding the test sample to a culture of a Saccharomyces cerevisiae strain into which has been introduced multiple copies of a gene encoding sterol .DELTA.14 reductase and also to a strain of S. cerevisiae that does not have the introduced gene; positive samples are identified after incubation by observation that growth inhibition in the culture having no introduced reductase gene exceeds growth inhibition in the culture having the introduced reductase gene. In preferred embodiments, a known inhibitor of sterol .DELTA.14 reductase is employed in solidified media in both the primary and the secondary screens, resulting in an assay that is highly sensitive and specific for the detection of sterol .DELTA.14 reductase inhibitors.
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Bard Martin
Kirsch Donald R.
Lai Margaret H. K.
American Cyanamid Company
Grimes Eric
Krinsky Mary
Wax Robert A.
Webster Darryl L.
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