Synthetic gene circuits combining CRISPR interference and CRISPR activation inE. coli: importance of equal guide RNA binding affinities to avoid context-dependent effects

Abstract

Gene expression control based on CRISPR has emerged as a powerful approach for constructing synthetic gene circuits. While the use of CRISPR interference (CRISPRi) is already well-established in prokaryotic circuits, CRISPR activation (CRISPRa) is less mature and combination of the two in the same circuits is only just emerging. Here, we report that combining CRISPRi with SoxS-based CRISPRa inEscherichia colican lead to context-dependent effects due to different affinities in the formation of CRISPRa and CRISPRi complexes, resulting in loss of predictable behaviour. We show that this effect can be avoided by using the same scaffold guide RNA structure for both complexes.