Bacillus methanolicus Genome sequencing and assembly

We established the first CRISPR-Cas9-mediated genome editing tool for B. methanolicus MGA3. This one-plasmid system utilizes Cas9-mediated double-strand breaks and engages two native repair pathways in B. methanolicus: directed homologous recombination for scarless genome deletions, gene replacements, and non-homologous end joining, which introduces random mutagenesis in the targeted genomic region. We employed this newly established system to investigate and confirm the activity of catalase and alanine dehydrogenase, encoded by the genes katA and ald, respectively.