The application of PCR for the isolation of a lipase gene from the genomic DNA of an Antarctic microfungus.
We successfully isolated a lipase gene (designated lipPA) directly from the genomic DNA of an Antarctic isolate of Penicillium allii using PCR and a suite of degenerate primers specifically designed to target two conserved regions of fungal lipase genes. We applied the biolistic transformation system to successfully integrate the lipPA gene into a heterologous fungal host, Trichoderma reesei, one of the most powerful secretors of extracellular proteins, and induced the transformant to secrete an active lipase into the growth medium. The recombinant lipase had a temperature optimum of 25 degrees C at pH 7.9 and retained greater than 50% of the maximum activity from 10 degrees C to 35 degrees C and over a pH range from 4.0 to 8.5.
D000595:Amino Acid Sequence; D000864:Antarctic Regions; D001483:Base Sequence; D019470:Biolistics; D017931:DNA Primers; D040461:Gene Components; D015870:Gene Expression; D006863:Hydrogen-Ion Concentration; D008049:Lipase / Q000235:genetics* / Q000378:metabolism; D008969:Molecular Sequence Data; D010407:Penicillium / Q000235:genetics*; D010957:Plasmids / Q000235:genetics; D016133:Polymerase Chain Reaction / Q000379:methods; D017422:Sequence Analysis, DNA; D013696:Temperature; D014242:Trichoderma / Q000235:genetics
D017931:DNA Primers; D008049:Lipase