Rapid communication: Mapping of the myeloperoxidase (MPO) gene to pig chromosome 12.

Name of Gene Marker. MPO, myeloperoxidase. Source and Description of Primers. Primers (set A) were designed based on human (Genbank Accession no. D14466) and mouse (Genbank Accession no. X15313) MPO exon 9 and 10 sequences available in the Genbank database. Using porcine genomic DNA as template in the PCR, these primers amplified a 1-kb fragment. Set B primers were designed based on pig DNA sequence data generated using set A primers. This 893-bp pig sequence has been deposited into Genbank as a sequence tagged site (STS), Accession no. AF208527. Because the predicted exon 9 and 10 sequences total 172 bp, the predicted pig intron 9 is 721 bp, slightly longer than the human MPO intron 9 length of 495 bp. Primer Sequences. Set A primers were as follows: forward 5′ CCA ACC CCC GTG TCC CCC TCA GC 3′ and reverse 5′ CGT GGT CCC TGC TGC GCT GCA TG 3′. Set B primers were as follows: forward 5′ GAG GGT AGT GCT GGA AGG 3′ and reverse 5′ GCT CCC GAA TCT CAT CTA C 3′. Method of Detection. The PCR with set A primers was performed in 10L PCR reactions including 0.38 U Taq Gold DNA polymerase (Perkin-Elmer, Branchburg, NJ), 1× PCR buffer, 1.5 mM MgCl2, 0.2 mM of each dNTP, 3 pmol of each primer, and 12.5 ng genomic DNA. Thermocycling was carried out in a Robocycler (Stratagene, La Jolla, CA) or a MJ Research, PTC-100 instrument (Watertown, MA) and included initial denaturation at 95°C for 5 min, followed by 35 cycles at 95°C for 45 s, 63°C for 1 min, 72°C for 1.5 min, and a final extension step at 72°C for 5 min. Products from Yorkshire, Landrace, and Meishan breeds were directly sequenced using dye terminators and an ABI 377 sequencer (Perkin-Elmer, Foster City, CA). The set B primers were used to amplify an 828-bp fragment. This product was used for physical and linkage mapping. Set B-PCR was performed using the MJ Research, PTC100 instrument and conditions as for set A except that Taq DNA polymerase (Promega, Madison, WI) was used and that the first denaturation step in the PCR