Highly efficient genome editing in via miniature DNA nucleases IscB.

Existing CRISPR-based genome editing techniques for () are limited due to the large size of the gene. IscB, a recently reported DNA nuclease, is one-third the size of Cas9, making it a potential tool for genome editing; however, its application in remains unexplored. In this study, two IscB and enhanced IscB (enIscB)-based genome editing systems, named pBsuIscB and pBsuenIscB were established in SCK6, and their deletion efficiencies ranging from 13.3 % to 100 %. Compared to the pBsuIscB system, the pBsuenIscB system showed higher deletion efficiency, inducing the deletion of a large genomic fragment with a single ωRNA. Additionally, the pBsuenIscB system could integrate both single-copy and multi-copy genes in the SCK6 genome. Lastly, the pBsuenIscB system could efficiently conduct a second round of genome editing in SCK6. This study indicates that IscB can be used for genome editing in , enabling the efficient construction of engineered strains for large-scale biomolecule production.