A novel “chassis” PSC line for precise safe harbor site integration via selection-aided RMCE
Merve Ay, Desi Veleva, Mohammad M. Chowdhury, Andrew B.J. Prowse, Dmitry A. Ovchinnikov · Stem Cell Research · 2025
We describe here a human pluripotent stem cell line (hPSC) designed to serve as an acceptor (a "chassis" cell line) for the recombination-mediated cassette exchange (RMCE)-mediated precise insertion of DNA fragments into a safe harbor locus. To enable the use of Flp recombinase for directional insertion, we targeted one of the AAVS1 safe harbor alleles with an excisable negative selection cassette flanked with FRT and FRT3 sites. The generated hPSC line displayed typical colony morphology, pluripotency signatures and normal karyotype. We expect it to provide a useful AAVS1 safe harbor "landing site" for FRT/FRT3-flanked transgenes.