Rapid communication: isolation and regional localization of the porcine glial fibrillary acidic protein (GFAP) gene.

Genus and Species. Sus scrofa. Locus. Pig glial fibrillary acidic protein (GFAP) gene. Source and Description of Primers. The cDNA sequence of the human GFAP gene (GenBank accession no. J04569) was aligned to the mouse GFAP gene (GenBank accession no. X02801). Well-conserved sequence regions corresponding to exon 5 and exon 6 of the mouse GFAP gene were selected for PCR primers designed using the program PRIMER version 0.5 (Lincoln et al., 1991). The forward primer was 5′-TTG CAG TCC TTG ACC TGC G-3′ and the reverse primer was 5′-GTA GGT GGC GAT CTC GAT G-3′. The primers were used to amplify 412 bp of the porcine GFAP gene from genomic DNA. The 234-bp predicted exonic regions (including the primer sequences above) of the pig sequence showed 93.5% identity at the nucleotide level with the human GFAP cDNA (GenBank accession no. J04569). The 412-bp pig sequence reported in this paper has been submitted to GenBank (accession no. AF250778). Method of Detection. The PCR products were obtained in a 25L reaction volume of 1 × PCR buffer (Promega, Madison, WI), consisting of 100 ng of porcine genomic DNA, 10 pmol of each primer, 100 M of each dNTP, 1.5 mM MgCl2, and 2.0 Units Taq DNA Polymerase (Promega). The PCR profile included 3 min at 94°C followed by 35 cycles of 30 s at 94°C, 45 s at 60°C, 1 min at 72°C, and a final extension of 5 min at 72°C in a Mastercycler gradient (Eppendorf Limited, Germany). Porcine-specific PCR fragments were separated by electrophoresis on a 2.0% agarose gel in 1 × TAE buffer and stained with 0.5 g/mL ethidium bromide. Chromosomal Localization. The pig/rodent somatic cell hybrid panel was used for regional assignment of GFAP on the pig chromosome (Yerle et al., 1996). The panel consists of 19 pig/Chinese hamster somatic cell hybrids (numbers 1 to 19) and eight pig/mouse somatic cell hy-