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Figure 1 | BMC Developmental Biology

Figure 1

From: Tetracycline-controlled transgene activation using the ROSA26-iM2-GFP knock-in mouse strain permits GFP monitoring of DOX-regulated transgene-expression

Figure 1

The gene targeted R26t1Δ allele controls the expression of DOX-inducible transcription from the endogenous promoter of the Gt(ROSA)26Sor locus. A) Gt(ROSA)26Sor and Gt(ROSA)26t1 (R26Rt1) loci. Exons are indicated as grey boxes; light grey show alternative spliced exons. Genetic elements inserted at the first XbaI site in intron1 of Gt(ROSA)26Sor are given in coloured boxes: first (yellow), the adenovirus major late splice acceptor sequence followed by the small intron of the adenoviral tripartite leader and the iM2 coding region terminated by the human growth hormone polyadenylation signal (hgh_pA); second (red), a loxP site (black triangles) flanked neomycin/uracil selection cassette [36] terminated by hgh_pA; third (green), the PtetO5-GFP gene module terminated by SV40_pA. Diagnostic restriction sites are indicated (E = EcoRV, Xh = XhoI, X = XbaI, P = PacI, A = AscI). The transparent green bar represents the chromosomal region covered by the targeting vector. Red-framed black boxes below R26Rt1 mark positions of Southern blot probes that detect restriction fragments given as red lines in kilo bases (kb). B) Left: Southern blots of genomic DNA isolated from wild type (WT) and PCR-preselected ES cell clones 27, 48, 60, 64, 65 and 67. Right: Images of R26t1 ES cells show DOX-dependent GFP expression. C) Schematic representation of the R26Rt1Δ allele after Cre mediated removal of neo/ura at the R26t1 allele (symbols are as in 1A). Transcriptional starts of the ROSA26 and Ptet promoter are indicated by arrows. Below the 5'-RACE identified exons (solid lines) and introns (dashed lines) of the iM2 encoding R26t1Δ mRNA are given. Below the 5' ends of mRNA from embryos, kidney liver and spleen are aligned to Gt(ROSA)26Sor transcript variants of the database (NCBI Accession numbers are indicated).

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