Construction by one-step gene replacement of Trichoderma reesei strains that produce the glucoamylase P of Hormoconis resinae.

Abstract

Two one-step gene replacement vectors containing either the Hormoconis resinae glucoamylase P (gamP) genomic gene or the corresponding cDNA, each under the control of the promoter of the Trichoderma reesei cellobiohydrolase 1 gene (cbh1), were constructed and used to replace the cbh1 gene in a T. reesei strain. In both vectors the cbh1 promoter is precisely fused to the gamP protein coding region. Both the gamP cDNA and the genomic gene direct the secretion of the active glucoamylase P (GAMP) enzyme from T. reesei, which indicates that the intron sequences in the genomic gamP gene are processed in T. reesei. According to the results, a T. reesei transformant strain, in which the cbh1 gene has been replaced by a single copy of the gamP genomic gene, secretes more active GAMP than does a transformant strain having three copies of the cDNA clone in tandem orientation at the cbh1 locus.