We have previously isolated and cloned the gene for pyruvate decarboxylase, pdcA, in Aspergillus nidulans (Lockington et al. 1977 Gene, in press) using PCR. Here we identify the genetic and physical map position of the gene.
The ordered cosmid library consisting of all clones from the pWE15 and pLORIST2 libraries showing hybridization to chromosome-specific probes (Brody et al. 1991 Nucl. Acids Res. 19:305-3109) was screened, initially by PCR analysis of DNA from cosmid pools and finally by colony hybridization. The cosmid SW16B11 was shown to contain the pdcA gene. This cosmid is part of a small (250 kilobase) contig on chromosome III of A. nidulans. All the cosmids in this contig were probed with the PCR probe, and no other cosmids hybridized to the pdcA probe.
A gene disruption of the pdaA gene was obtained with a plasmid construct in which the riboB gene from pPL3 (Oakley et al. 1987 Gene 53:293-298) replaced a portion of the pdcA gene. The disruption was obtained by screening diploid transformants for integration events on chromosome III and confirmed by Southern transfer analysis of haploid segregants. A strain containing the pdcA gene disruption marked by the riboB gene was used to map the position of pdcA on chromosome III genetically. Strong linkage of pdcA was found to the sorB and cnxH genes in the progeny of a cross between a strain of genotype yA2;sorB11cnxH14;pyroA4;riboB2 and a strain of genotype yA1 adE20suA1 adE20; argB2 pdcA ::riboB+;aldA67 riboB2. From 96 progeny the following linkage distances and orders were obtained for the pdcA, chxH, and sorB genes.
No recombinants in the two classes requiring a double crossover event were obtained. This result thus positions the pdcA gene on the A. nidulans linkage map and also positions the cosmid containing SW16B11. No other genes are at present mapped to this small contig.
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