Construction and analysis of a random mutation library of △6-desaturase gene in Mucor sp. EIM-10

Abstract:

△6-desaturase plays an important role in the biosynthesis of γ-linolenic acid in Mucor sp. EIM-10. To improve enzymatic activity and confirm the effect of primary structure on the △6-desaturase activity, a random mutation library was constructed. The △6-desaturase gene (mcd6, GenBank accession No. EU717846) was mutated by error-prone PCR strategy. The PCR products were ligated into the yeast expression vector PYMD6PMCD6. The resulting plasmid was named PYTBMCD6 and transformed into Saccharomyces cerevisiae strain INVSc1. A random △6-desaturase gene mutation library was constructed in Saccharomyces cerevisiae. The titers of random mutation library were up to 4.6×104 CFU. The random mutation library of △6-desaturase gene could be used for screening the mutants with change in catalytic activity and the subsequent site-directed mutagenesis.