I was wondering if anybody has information on how to choose guides for optimal gene activation/repression using multiplex CRISPR/Cas9s with only two guides?
I have tried different combinations e.g. two guides with average activity v.s. one poor guide plus an average guide and got mixed results. Sometimes the latter outperformed the former, which is counter-intuitive.
So my question is, how should one choose the two guides for best gene activation/repression? More specifically, how far apart should these two guides be, how far away should them be from the TSS, and what should their individual activity level be for best synergistic results etc?
Any suggestions will be highly appreciated. Many thanks!