RT Journal Article SR Electronic T1 Multiplex gene editing by CRISPR-Cpf1 through autonomous processing of a single crRNA array JF bioRxiv FD Cold Spring Harbor Laboratory SP 049122 DO 10.1101/049122 A1 Bernd Zetsche A1 Matthias Heidenreich A1 Prarthana Mohanraju A1 Iana Fedorova A1 Jeroen Kneppers A1 Ellen M. DeGennaro A1 Nerges Winblad A1 Sourav R. Choudhury A1 Omar O. Abudayyeh A1 Jonathan S. Gootenberg A1 Wen Y. Wu A1 David A. Scott A1 Konstantin Severinov A1 John van der Oost A1 Feng Zhang YR 2016 UL http://biorxiv.org/content/early/2016/10/01/049122.abstract AB Microbial CRISPR-Cas defense systems have been adapted as a platform for genome editing applications built around the RNA-guided effector nucleases, such as Cas9. We recently reported the characterization of Cpf1, the effector nuclease of a novel type V-A CRISPR system, and demonstrated that it can be adapted for genome editing in mammalian cells (Zetsche et al., 2015). Unlike Cas9, which utilizes a trans-activating crRNA (tracrRNA) as well as the endogenous RNaseIII for maturation of its dual crRNA:tracrRNA guides (Deltcheva et al., 2011), guide processing of the Cpf1 system proceeds in the absence of tracrRNA or other Cas (CRISPR associated) genes (Zetsche et al., 2015) (Figure 1a), suggesting that Cpf1 is sufficient for pre-crRNA maturation. This has important implications for genome editing, as it would provide a simple route to multiplex targeting. Here, we show for two Cpf1 orthologs that no other factors are required for array processing and demonstrate multiplex gene editing in mammalian cells as well as in the mouse brain by using a designed single CRISPR array.