Incorporation of a bzip transcription factor specific sgRNA into pMZ379 plasmid.

Abstract

This thesis outlines the successful incorporation of a Schizosaccharomyces japonicus-specific sgRNA targeting the SJAG_02569 gene into the CRISPR-Cas9 plasmid vector pMZ379. The study highlights the use of CRISPR-Cas9 gene editing to expand molecular tools for S. japonicus, a promising but underutilized model organism. The procedure involved plasmid extraction, PCR amplification using sgRNA-carrying primers, transformation into E. coli DH5α, and verification via colony PCR. Despite technical challenges, including low transformation efficiency and potential template contamination, the study confirmed the correct construction of a recombinant plasmid carrying the desired sgRNA. This newly engineered plasmid will facilitate future genome editing experiments in S. japonicus, addressing the current lack of a CRISPR-Cas9 system for this species.