Abstract
Single-cell CRISPR (Perturb-seq) screens have primarily relied on Cas9 whereas Cas12a, despite its unique effectiveness for multiplex guide expression, remains underexplored. This may be due to Cas12a’s guide RNA array (pre-crRNA) self-processing activity and the subsequent challenges associated with pre-crRNA sequence recovery. By developing modified pre-crRNA expression vectors and a degron-based Cas12a system, we overcome the self-processing constraint, allowing for accurate detection of pre-crRNAs at the single-cell level, thus greatly expanding possibilities for future Perturb-seq efforts.
Competing Interest Statement
Authors have submitted a provisional patent application that is based on the technology described in this manuscript. All authors are or were employed by Genentech Inc., a member of the Roche group, South San Francisco, CA, USA, at the time of their contribution to this work. V.S., C.G., K.D., S.W., and B.J.H. are equity holders in Roche.
