Fig. 1.

GABA_A receptor α₆subunit gene disruption by homologous recombination. A, Wild-type α₆ gene, targeting (replacement) vector, and the disrupted α₆ gene structures. Numbersindicate exons. On the Replacement vector, thebroken line indicates pBluescript (pBS) sequences. On the wild-type allele, theasterisk marks exon 8 where the IRES lacZ/neo cassette was inserted. Only relevant restriction sites are shown. A, AflII;B, BamHI; N,NcoI; S, SphI;X, XhoI. Arrows mark theneo and tk gene promoter sites and direction of transcription. The lacZ coding sequence orientation is the same as the α₆ gene, thus permitting its translation from the IRES sequence (striped box) to be initiated on the mRNA derived from the α₆ promoter. Expected restriction fragment lengths diagnostic for homologous recombination and the probes used to detect these are marked by double-headed arrows andhorizontal bars, respectively. B, Confirmation of α₆ mutant allele germline transmission. Biopsy tail DNA samples were digested with SphI, electrophoresed, and Southern-blotted. The membrane was probed withPROBE A (3′ flanking). Wild-type (+/+) individuals give a 15 kb band, the homozygous null (−/−) animals give a 9 kb band, and heterozygotes (+/−) give both bands.