Full length mouse glycophorin gene constructed using recombinant polymerase chain reaction

Abstract

Recently, an incomplete cDNA clone for a major mouse glycophorin gene, pGP315, and a genomic clone, pGX7 (which contains the first exon and nucleotide sequences around the transcription start sites) was isolated and sequenced by Matsui et al. (1). Since there were no available restriction sites for the construction of a full length mouse glycophorin A gene, the recombinant PCR technique was adapted to splice together the above two partial sequence clone inserts to obtain a full length recombinant DNA fragment (1053 bp) containing the proper sequence of mouse glycophorin A cDNA. The PCR reconstructed DNA fragments were verified by: gel electrophoresis to contain the expected sizes, hybridization to probes made from the DNA components before recombination, and confirmed by the restoration of a previously destroyed restriction enzyme site. The corrected gene sequence for pGP315 is also reported.