A novel screening system based on gene targeting to enrich the modified mammalian cells: without leaving selection marker and additional sequence

摘要

Schematic structure of targeting plasmids. a Targeting plasmid 1 contains a PNS-expression cassette in which genes of PuroR and TK, linked by the P2A sequence, are under the control of the PGK promoter and the bGH p(A) sequence. This cassette is surrounded by sequences of two homology arms. A CD expression cassette is located outside the homology arms, which is controlled by the EF-1α promoter and the EF-1α p(A) sequence. b Targeting plasmid 2 contains a GFP-expression cassette under the control of the CMV promoter and bGH p(A) sequence surrounded by two homology arms. P2A, porcine teschovirus-1 2A; PGK, phosphoglycerate kinase; bGH, bovine growth hormone; p(A), polyadenylation; EF-1α, elongation factor-1 alpha; CMV, cytomegalovirus; Ori, origin of replication; AmpR, ampicillin resistance