Prior to the development of CRISPR/Cas9 genome-editing technologies, RNA interference (RNAi) was used broadly to suppress the expression of genes of interest and to screen large gene sets for their role in biological processes and phenotypes. RNAi is a biological process involved in immunity and gene regulation that is present in many eukaryotes, including humans. It involves RNA guided inhibition of gene expression by the RNA-induced silencing complex (RISC) through translational or transcriptional repression. The gene targeting in RNAi is achieved by a double-stranded RNA (siRNA) or a small micro RNA (miRNA). Similar to the gRNAs required for CRISPR/Cas9 technologies, siRNAs and miRNAs can be introduced into cells by transduction using viruses that encode shRNAs or miRNA, respectively. The shRNAs consist of sense and antisense strands separated by a hairpin loop that are converted into siRNAs by cleavage of the hairpin loop.

While RNAi technologies have been largely displaced by CRISPR technologies with regard to model generation and large-scale functional genomic screens, they can provide a complementary approach for validation of genes identified by CRISPR screens. At the NKI, whole-genome collections of individual lentiviral constructs encoding shRNAs as well as synthetic siRNAs are made available to researchers for this purpose.