Figure 2.

Identification of ES cell clones with targeted disruption of the MKK4 gene. (A) Genomic DNA prepared from ES cell clones indicates the presence of all three expected genotypes. EcoRI restricted DNA was probed with a radiolabeled fragment of the MKK4 gene (Fig. 1). The upper band (6 kb) corresponds to the wild-type allele, and the lower band (2.5 kb) corresponds to the mutant allele. (B) Northern blot analysis of total RNA (10 μg) isolated from wild-type and homozygous knockout ES cells. Blots were probed with a random-primed ³²P-labeled mouse MKK4 cDNA probe. The blots also were probed for β-actin mRNA as an internal control. (C) Reverse transcription-PCR analysis of MKK4 mRNA expressed by ES cells. RNA isolated from ES cells was amplifed with primers specific for MKK4 and hypoxanthine–guanine phosphoribosyltransferase mRNA.