The Effect of the TacA Knockout Transcription Factor on the cbhI Gene Transcription and Expression in the Filamentous Fungus Strain Penicillium verruculosum

Abstract

The tacA gene encoding the TacA repressor protein is cloned by the method for “walking” in uncloned DNA from the genomic DNA of the fungus Penicillium verruculosum B1-221-151. Knockout of the tacA and niaD genes by the CRISPR/CAS9 method leads to the production of a new recipient strain P. verruculosum ΔniaDΔtacA, characterized by a higher rate of extracellular protein biosynthesis. Analysis of the cbhI gene transcription and expression in the original P. verruculosum B1-221-151 strain and in the P. verruculosum ΔniaDΔtacA strain shows a sharp increase in the level of the cbhI gene transcription as early as 2 h after the start of induction with cellobiose, cellotriose, gentiobiose, and a mixture of di- and trisaccharides compared with transcription of the cbhI gene in the original strain. The specific activity of cellobiohydrolase I, the main enzyme of the cellulolytic complex of the P. verruculosum fungus, tripled after 96 h of fermentation of the ΔtacA strain compared with the initial strain.