Genome resequencing of wild type and mutant strains for enhancing cellulase production in Pichia pastoris (syn. Komagataella phaffii)

Through five rounds of iterative ARTP (atmospheric and room temperature plasma) mutagenesis and microfluidic screening, cellulase production capacities of mutants in each round were gradually enhanced. To uncover genomic alterations accumulated during iterative ARTP mutagenesis, the ten selected mutant strains and the wild type strain were subjected to whole-genome resequencing. Comparative genome analysis identified three accumulated genomic alterations in coding region during iterative ARTP mutagenesis and screening. Intriguingly, the point mutation Rsc1(V22G) was observed in all the top-performing producers selected from each round. The other two mutations Rvs167(G9R) and Pmt6(V57M) were only found in one selected mutant. These results suggested that Rsc1 might play an important role in cellulase production, whereas Rvs167 and Pmt6 might play minor roles.