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. 2012 Jan 20;18(6):289–297. doi: 10.1093/molehr/gas005

Figure 1.

Figure 1

T2R5-Cre/GFP construction. BAC DNA (RP23-342N5) contains more than 70 kb 3′ and 5′ sequence of T2R5 gene. The IRES-GFP/Cre-frt-KanR-frt fragment is obtained from plasmid pICGN21 by PCR. Two homologous arms of 48 bp from the mT2R5 gene are synthesized into both sides of the primers used to amplify the IRES-eGFPcre-frt-KanR-frt cassette in the pICGN21 plasmid. The IRES-eGFPcre-frt-KanR-frt cassette is introduced into the T2R5 gene downstream of the start codon (93 bp) and upstream of its stop codon (60 bp) by Red-mediated homologous recombination, followed by flp-mediated removal of the KanR selectable marker from the BAC-mT2R5-IRESCre/GFP construct. Final construct was verified by pulse field gel electrophoresis analysis and sequence. P1/P2 and P3/P4 primers are used for genotyping (A). In order to verify the function of Cre, we crossed the T2R5-Cre/GFP transgenic mouse with RosA-LoxP-LacZ mouse and got double transgenic mouse. After X-gal staining of sections of mouse tongue, we found positive cells in circumvallate papillae (B) and foliate papillae (C). Scale bar 12.5 μm.