Inactivation of katG was further confirmed by a phenotypic assay as the katG mutant was more resistant to Isoniazid (INH) treatment (16 μg/mL) than that of the wild-type strain (Fig. ). Because the relative position of the introduced premature stop codon in an open reading frame (ORF) can significantly affect the gene inactivation efficiency, we systematically calculated the possible targetable codons of CTBEevolve in mycobacteria with CRISPR-CBEI . As shown in Fig. , more than 75%, 60%, and 40% ORFs of the analyzed mycobacterium species (M. tuberculosis, M. smegmatis, and M. marinum) can be possibly targeted by CTBEevolve to introduce at least one premature stop codon within top 75%, 50%, and 25% of the ORF body, respectively, demonstrating that numerous genes can possibly be inactivated by CTBEevolve in mycobacteria.