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. 2015 Sep 30;24(24):7097–7110. doi: 10.1093/hmg/ddv409

Figure 2.

Figure 2.

I-SceI-induced correction (A and B) or mutagenesis (C) of an integrated mHPRT minigene. (A) Gene correction templates are shown schematically, aligned with the target mHP-I*-RT minigene. Templates were sense or antisense single-stranded oligonucleotides (SssO, AssO), dsOs or dsL (for oligonucleotide sequences, see Supplementary Material, Table S3). Each of these four template classes was made with the I-SceI site and stop codon (-I*), the I-SceI site only (-I) or neither (-wt; wild-type). (B) HPRT − cells (clone 2.1) carrying the mHP-I*-RT minigene and a tetracycline-regulated I-SceI expression cassette were grown with or without tetracycline and transfected with the indicated templates detailed in (A). The frequencies of resulting HAT-resistant colonies are shown for oligonucleotides templates (left panel) and dsL templates (right panel). Error bars show SDs for three experiments. (C) Three HPRT+ clones (14.2, 7.1 and 5.1) carrying the mHP-I-RT minigene were transfected with I-SceI expression plasmid (+), or a vector control (−), and selected in 6TG. Colonies resulting from selection of 5 × 105 cells are shown.