Fig 1.
Targeted disruption of the α-synuclein locus by homologous recombination. (A) Schematic drawing of the wild-type (WT) locus, the targeting construct, and the recombined allele. A null allele was generated by blocking transcription from the α-synuclein allele with a “STOP” cassette (12) inserted upstream of the start ATG. The location of hybridization probes for Southern blot analysis (5′ probe, 3′ probe) are shown. (B) Southern blot analysis of embryonic stem cell clones. DNA was digested with BglII and was hybridized with the 5′ probe. The WT allele produced a 9.5-kb fragment, and the targeted allele produced a 6.5-kb fragment. BglII-digested DNA was also hybridized with the 3′ probe (data not shown). (C) Southern blot analysis of genomic DNA from littermates of a α-synuclein heterozygote mating. Tail DNA was isolated and analyzed as in B. (D) Northern blot analysis of whole brain mRNA. Total RNA was hybridized with α-synuclein cDNA probe containing the entire coding region. The lower panel shows ethidium bromide staining of the 28s ribosomal bands. (E) Western blot analysis of whole brain protein blotted with an α-synuclein antibody (Transduction Laboratories).