Figure 7: Anti-tag inactivation of Cas13 prevents self-targeting of antisense CRISPR transcripts.

A. Plasmids engineered to constitutively express the CRISPR array in sense or anti-sense orientations. A variant of the antisense plasmid was constructed with a single mutation in the PFS flanking spc2 (spc sequence in orange, repeat sequences in black, mismatch highlighted in red). B. Conjugation assay showing CRISPR-dependent toxicity of a single mismatch in antisense CRISPR array transcript C. An aTc-inducible target construct matching spc4 was integrated into the tRNA^Arg locus of the L. seeligeri genome. D. Conjugation assay demonstrating inhibition of CRISPR interference by expression of antisense CRISPR array. Plasmids bearing sense or antisense array were conjugated into WT or ΔCRISPR recipients carrying the inducible target, and plated in the presence of 100 ng/mL aTc.