Figure 2

Schematic overview of the miRNA-based CRISPR vector construction system for multiplex genome engineering. In this construction miRNAs (or shRNAs) and sgRNAs alternately emerge in a polycistron. 5′ and 3′ arm of miRNAs (or shRNAs) were required for Drosha to recognize and cut the pri-miRNAs (or pri-shRNAs) from the original transcription. Pre-miRNA (or pri-shRNAs) can be transported into cytoplasm then cut by Dicer to form a double stranded RNA. Further, these double stranded-RNAs can be loaded onto RISC complex and function as a mature miRNA (or siRNA) to target on its corresponding mRNA. Simultaneously, sgRNAs bind to Cas9 (including dCas9-VP64, dCas9-KRAB, etc.). Redundant nucleotides can be degraded by ubiquitous RNase.