Identification and characterization of six new alternatively spliced variants of the human μ opioid receptor gene, Oprm
Section snippets
Primer design
Using Vector NTI (Informax, Bethesda, MD, USA), we identified potential mouse homologs of exon 5 in the human Oprm gene using a sequence alignment with an approximately 7.7 kb mouse genomic sequence containing exons 5a, 5b, 5c, 5d and 5e, an approximately 7.3 kb rat and a corresponding approximately 8 kb human corresponding genomic sequence obtained from the genome databases (NBCI and Ensembl). Three sets of antisense primers were designed based upon the alignment (see below). One of the
Cloning new splice variants from the human Oprm gene
To isolate potential human MOR-1 variants homologous to those identified in mice, we designed three sets of human antisense primers based upon the human genomic sequence alignment with the mouse and rat exon 5′s and then used these antisense primers together with two sense primers from 5′UTR of exon 1 in an RT-PCR strategy. We identified six new variants, hMOR-1B1, hMOR-1B2, hMOR-1B3, hMOR-1B4, hMOR-1B5 and hMOR-1Y, from Be(2)C, a human neuroblastoma cell line that expresses high level of μ
Discussion
Our previous studies revealed that the mouse Oprm gene undergoes extensive splicing to generate multiple MOR-1 variants (Bolan et al 2004, Pan et al 1999, Pan et al 2000, Pan et al 2001), with similar splicing patterns in the rat (Pasternak et al., 2004). A question arose as to whether the same complex splicing existed in the human Oprm gene. Recently, we isolated two splice variants, hMOR-1O and hMOR-1X, from the human Oprm gene (Pan et al., 2003), in addition to hMOR-1A which had been
Acknowledgments
This work was supported, in part, by a research grant to Y.-X.P. (DA13997), and a research grant and a Senior Scientist Award to G.W.P. (DA02615 and DA00220) from the National Institute on Drug Abuse and by a Core Grant to MSKCC from the National Cancer Institute (CA8748).
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