Targeted disruption of the PCAF-B/GCN5
locus. (A) Targeting strategy for
PCAF-B/GCN5 gene. The configuration of
the wild-type (WT) allele is shown (Top). The genomic
region containing exons encoding the 3′ terminus of PCAF-B/GCN5
(shown by boxes) was replaced by a neomycin resistance gene cassette in
the targeting vector (Middle), yielding the targeted
allele (Bottom) by homologous recombination. Restriction
sites: Bg, BglII; E, EcoRI; X,
XhoI. (B) Southern blot of
EcoRI-digested genomic DNAs isolated from offspring of
heterozygous mice. The probe hybridizes with 13- and 10-kb fragments in
the wild-type and targeted allele, respectively. (C)
Immunoblot of protein extracts from
PCAF-B/GCN5+/+,
PCAF-B+/−, and
PCAF-B−/− embryos. PCAF-B/GCN5 could
not be detected in PCAF-B−/− embryos
(Upper). Note that an immunoreactive band around 55 kDa
could not be detected. Further characterization would be required to
resolve the issue of the N-terminally truncated form of mouse
PCAF-B/GCN5. As an internal control, TFIIB was also detected
(Lower).