Cloning, Chromosomal Localization, And SNPs Detection Of The Porcine IRS-â…  Gene

Insulin receptor substrate (IRS) molecules are key mediator in insulin signaling and play a central role in maintaining basic cellular functions. In this study, insulin receptor substrate-1 (IRS-1), which play a important role in mediating the growth, the metabolism of skeletal muscle, was selected as a candidate gene for studying the molecular mechanisms of muscle' growth and type 2 diabetes in the pig. The porcine IRS-1 gene was cloned, located in porcine chromosome, and scanned for SNPs. The results are as follows:1. By designing primers from the homologous region between the human and mouse, the coding region of IRS-1 gene was cloned by using the RT-PCR approach. Eight fragments was amplified and sequenced for makeing a contig of porcine IRS-1 gene. The length of this contig is 3756 base pairs and covers ninety five percent of the coding region of porcine IRS-1 gene.2. The radiation hybrid panel (IMpRH) was used for doing the regional assignment of IRS-1 gene in pig. Using the two point analysis, statistical result revealed that the most significantly linked marker is SJF315 on chromosome 15q25, with a LOD score 7.65. Thus the porcine IRS-1 gene was located in SrSrC15q25.3. Two fragments of the IRS-1 gene which had been found some important SNPs in human was scanned for searching SNPs in two pig populations, Duroc(42 individuals) and Tibetan pig (28 individuals), by using the SSCP approach combined with the sequencing method. Two SNPs which caused amino acid variation were detected in those fragments. One is G/A in position 2650, another is C/A in position 3257. Comparing to the ORF of human's, both of the SNPs can cause the amino acid change, and maybe affect the functions of IRS-1.