Fig. 1
From: Mutagenic consequences of cytosine alterations site-specifically embedded in the human genome
Overview of the TATAM system. Structures of cytosine alteration (a) and the principle of the TATAM system (b). X on the targeting vector indicates the position of cytosine, 5-mC, 5-BrC, U, 5-BrU, or thymine at the BssSI site. The targeting vectors pvINTC:G, pvINT5mC:G, pvINT5BrC:G, pvINTU:G, pvINT5BrU:G, or pvINTT:G and the I-SceI expression plasmid pCBASce were co-transfected into TSCER122 cells. Double-strand break at the I-SceI site enabled gene targeting by inducing site-specific homologous recombination. The targeting vector contained an MseIR site that was resistant to MseI digestion and thereby distinguished targeted and non-targeted revertants of TK. TK revertants were selected by using HAT. Genomic DNA of the revertant colonies was prepared, and part of the TK gene containing the modified DNA integrated site was amplified by PCR. The amplified fragment was sequenced as described in the Materials and Methods section