High-throughput identification of synthetic riboswitches by barcode-free amplicon-sequencing in human cells.

ABSTRACT: Synthetic riboswitches mediating ligand-dependent RNA cleavage or splicing-modulation represent elegant tools to control gene expression in various applications, including next-generation gene therapy. However, due to the limited understanding of context-dependent structure-function relationships, the identification of functional riboswitches requires large-scale-screening of aptamer-effector-domain designs, which is hampered by the lack of suitable cellular high-throughput methods. Here we describe a fast and broadly applicable method to functionally screen complex riboswitch libraries (~1.8 × 10⁴ constructs) by cDNA-amplicon-sequencing in transiently transfected and stimulated human cells. The self-barcoding nature of each construct enables quantification of differential mRNA

SUBMITTER: Strobel B 

PROVIDER: S-EPMC7002664 | biostudies-literature | 2020 Feb

REPOSITORIES: biostudies-literature