PiggyBac-mediated genomic integration of linear dsDNA-based library for deep mutational scanning in mammalian cells.

ABSTRACT: Deep mutational scanning (DMS) makes it possible to perform massively parallel quantification of the relationship between genetic variants and phenotypes of interest. However, the difficulties in introducing large variant libraries into mammalian cells greatly hinder DMS under physiological states. Here, we developed two novel strategies for DMS library construction in mammalian cells, namely 'piggyBac-in vitro ligation' and 'piggyBac-in vitro ligation-PCR'. For the first strategy, we took the 'in vitro ligation' approach to prepare high-diversity linear dsDNAs, and integrate them into the mammalian genome with a piggyBac transposon system. For the second strategy, we further added a PCR step using the in vitro ligation dsDNAs as templates, for the construction of high-content genome-integ

SUBMITTER: Wang Y 

PROVIDER: S-EPMC11071730 | biostudies-literature | 2023 Oct

REPOSITORIES: biostudies-literature