An In Vitro Assay of β-Galactosidase from Yeast

mouse genotyping by the three-primer multiple PCR. However, both of the smaller PCR products of 1068 (specific for the wild-type allele of mGluR4) and 1170 bp (specific for the mGluR4 transgene) were visible. Thus, all three allele combinations were clearly distinguishable on agarose gels stained with ethidium bromide. DNA from wildtype mice give a single smaller band (1068 bp; see Figure 2B, lanes 4, 5, 6 and 10). All transgenic homozygous mice showed the 1170-bp product (Figure 2B, lanes 2 and 9), and the heterozygous mice showed both PCR products (Figure 2B, lanes 1, 3 and 8). The results of the three-primer multiple PCR genotyping of mGluR4 mutant mice were confirmed by Southern blot and immunoblot analysis (data not shown). The results presented here confirm that multiple primer PCR can be applied to the genotyping of transgenic mice. The three-primer protocol outlined above increases the reliability of a genotyping screen because, unlike conventional two-primer PCR, threeprimer PCR will give a PCR product regardless of the genotype. Our results indicate that three-primer multiple PCR provides a very useful alternative for genotyping transgenic mouse and ES cells lines.