The 5' -sequence of the isopenicillin N-synthetase gene (pcbC) from Cephalosporium acremonium directs the expression of the prokaryotic hygromycin B phosphotransferase gene (hph) in Aspergillus niger
1989
Kuck, U. | Walz, M. | Mohr, G. | Mracek, M.
We have performed a comparative transcript analysis with RNA from two different strains of Cephalosporium acremonium, using synthetic oligonucleotides as specific probes for the isopenicillin N-synthetase gene (pcbC). Strain DSM 2353 shows a considerably higher amount of the pcbC transcript than strain ATCC 14553. Subsequently, a genomic library from C. acremonium strain DSM 2353 DNA was constructed using lambda vector EMBL4. We have isolated five recombinant clones containing identical copies of the pcbC gene as was confirmed by partial DNA sequencing. The 5' region of the pcbC gene was fused with the prokaryotic gene for hygromycin B phosphotransferase (hph) using a synthetic oligonucleotide linker. The resulting plasmid pMW1 can be used for high frequency transformations of the filamentous fungus Aspergillus niger (about 10 000 transformants/microgram plasmid DNA). From Southern hybridization analysis it can be concluded that all transformants tested contain vector DNA integrated into the genomic DNA. The expression of the prokaryotic hph gene in A. niger was conclusively demonstrated with an assay specific for hygromycin B phosphotransferase.
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