Engineering CRISPR Plasmids for Genome Editing in Schizosaccharomyces japonicus

Abstract

Schizosaccharomyces japonicus represents a dimorphic fission yeast which is able to switch between two forms: a unicellular yeast-like form and a hyphal (filamentous) form. Due to its unique biological features and close evolutionary relationship to humans, S. japonicus has proven to be a valuable model organism for studying cellular processes and mechanisms of human diseases. In this thesis research, our aim was to develop a CRISPR-Cas9 genome editing tool suitable for S. japonicus. To achieve this, pMZ379 plasmid was engineered through the incorporation of the designed sgRNA83 sequence by using a ligation-free PCR-based method. The Cas9 endonuclease successfully introduced double-stranded breaks in the target DNA of the S. japonicus genome and therefore, it was concluded that the CRISPR-Cas9 system is functional in S. japonicus cells. The development of this CRISPR-Cas9 genome editing tool provides a basis for future genetic modifications in S. japonicus.