Figure 2
From: Enhanced cell-permeant Cre protein for site-specific recombination in cultured cells
Cre-mediated recombination in cultured cells. (A). Structure of the recombination substrate in Tex.loxP.EG cells. The pBABE.lox.stp.EGFP retrovirus contains multiple polyadenylation sequences (stop cassette) flanked by loxP sites such that Cre-mediated recombination activates the expression of an enhanced green fluorescent protein (EGFP) reporter gene. (B). Dose-response of Cre-mediated recombination. Tex.loxp.EG cells were treated for two hours with increasing concentrations of each soluble Cre recombinase; were washed twice with PBS; and after 24 hours in normal growth medium, the percentage of GFP expressing cells was determined by FACS analysis. For clarity, the results obtained using different fusion proteins are plotted in two panels. The Upper Panel shows results for HNC (solid squares), HT7N'C (solid circles), HNCM (open squares), HC (open circles), and Cre (solid triangles). The Lower Panel plots the results for NCH6 (solid squares), H6C (solid circles), and CH6 (solid triangles). (C). Southern blot analysis of Cre-mediated recombination. Tex.loxp.EG cells were treated with 0.5, 1.0, 2.0, and 4.0 μM of HNC, HNCM and Cre proteins as described above, except after 24 hours DNA was extracted, digested with EcoR1 and analyzed by Southern blot analysis. Cre-mediated recombination results in the conversion of the loxP-containing fragment (upper band, U) to the recombination product (lower band, R).