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. 2003 Aug 11;140(2):315–322. doi: 10.1038/sj.bjp.0705447

Figure 1.

Figure 1

Strategy for deletion of the GABAB(1) gene. (a) Map of the murine GABAB(1) locus, showing the exon–intron structure of the gene. Exon nomenclature is according to Lamp et al. (2001). The targeting construct comprised the SAβGeo(LoxP)2, flanked by a 4126 bp EcoR1–Not1 5′ fragment and a 5304 bp Hpa1–Sal1 3′ fragment. Recombination between the GABAB(1) locus and the targeting construct resulted in the replacement of the genomic sequence from exon E1a1 to the 5′ part of exon E1a/b. Homologous recombination was selected by PCR, using the primer CQ113 and CQ114 (arrows), and later confirmed by Southern blotting. (b) Southern blot analysis of Sca1-digested genomic DNA confirming the correct junction. 5′ probe label indicates the 253 bp probe generated by PCR amplification using the primers 5′-ACTGAGCCTGGTCAAGGTCAG-3′ and 5′-CAACGCCACCGTGAAACCCT-3′. (c) Detection of GABAB(1) mRNA by RT–PCR gave rise to the expected 427 bp fragment, confirming the absence of GABAB(1) transcript in GABAB(1) mutant mice. GABAB(2) (970 bp RT–PCR product) transcript levels are normal in GABAB(1) mutant mice, despite the finding that the protein product could not be detected by Western blot. Amplification of the S16 sRNA was used as a control for efficient cDNA synthesis, and generated a 102 bp PCR product.