A simple approach for multi-targeted shRNA-mediated inducible knockdowns using Sleeping Beauty vectors.

ABSTRACT: shRNA expression is an established technique to transiently or permanently deplete cells of a particular mRNA/protein. In functional analyses of oncogenic pathways it can thus be used as an alternative to pharmacologic inhibitors, or as a means to address otherwise undruggable targets. Here we describe and functionally validate a simple reiterative cloning system to generate concatenated multi-shRNA expression plasmids. The multi-shRNA expression cassette can eventually be subcloned into any suitably designed vector for the stable transfection of cells, here tested with derivatives of the Sleeping Beauty transposon vector for stable transfection of multiple myeloma cell lines at the lowest biosafety level. We finally test inducible versions of such multi-cassette knockdown vectors and show

SUBMITTER: Fink S 

PROVIDER: S-EPMC6195277 | biostudies-literature | 2018

REPOSITORIES: biostudies-literature