Characterization, Polymorphism And Aplication In High Quality Pig Line Of Porcine Disease Resistant Genes-IRGC, MX1 Gene

With the development and application of DNA marker technology,the molecular breeding techniques such as marker-assisted selection(MAS) and marker-assisted introgression(MAI) combining with traditional breeding methods have greatly promoted the process of genetic improvement in pigs.As more and more bioinformatics and database are common,they afford the new method to separating and localization.In this paper,we used the method of the EST information and the Comparative Genomics to isolate two disease resistance candidate genes.We carried out the characterization prediction and association analysis.The main results are as follows:1.Based on the method of candidate gene,we found the porcine homologous EST from the ESTs.The specific primers were designed through the eontigs by the ESTs. immunity-related GTPase family,cinema(IRGC) and myxovirus(influenza virus) resistance 1(MX1)genes were isolated from Large White cDNA,which size is 1518bp and 2387bp respectivly.The sequence of IRGC was submitted to the NCBI GenBank and assigned the accession number of EU 703776.2.According to cloned cDNA sequence overlapping primers were designed to amplify the genome sequence of IRGC and MX1.The overlapping PCR products from Large White,Landrace,Meishan,Toughing,Qingping and Exi were cloned,sequenced, and assembled into respective contig.The IRGC contig with a length of 3345bp is covering the whole DNA sequence of IRGC gene.This sequence was submitted to the NCBI GenBank and assigned the accession numbers of EU 696780.A G-A mutation which makes the region be recognized by Hpaâ…¡in intron 12 was found in MX1 gene.3.Expression profiles of 1RGC and MX1 genes were analysis by semi-quantitative RT-PCR assay.IRGC expressed very lowly in tissues of heart,liver, spleen,lung,kidney,small intestine,stomach,backfat,muscle,uterus.MXI expressed higly in tissues of heart,spleen,stomach,uterus,expressed lowly in liver,small intestine,backfat.4.The profiles of functional domain in IRGC and MX1 protein were estimated and displayed primarily.The multiple sequence alignments were carried out by the CLUSTAL W program.IRGC amino acid sequence shared high similarity with dog (95%),cattle(93%),human(90%) and orangutang(90%).The deduced MX1 protein sequence shares 80%similarity with cattle,78%with human,77%with orangutang, 76%with dog.Polygenetictree of IRGC and MX1 protein in different species were analyzed by the MegAlign program in DNAStar software. 5.Studied on polymorphic sites of MX1 gene in intron 12 by PCR-RFLP,genotype and allele frequencies among different pig populations were determined.Analysis of PCR- Hapâ…¡-RFLP in MX1 gene showed that allele B was predominant in six pig breeds,with a frequencies of 0.83,0.89,1.00,0.94,0.87,0.89 in Large White,Landrace, Meehan,Toughing,Qingping,Exi pig,repectively.Association with PCR-Hpaâ…¡-RFLP in MX1 gene genotypes and phenotypes of economic traits in 295 F₂(L×M) pigs revealed that MX1 gene affected dressing percentage,lean meat percentage,loin eye height significantly(p＜0.05,the same below).Additive effects of dressing percentage,loin eye height were significantly.Dominance effect of height was significantly.Dominance effect of lean meat percentage was highly significantly(p＜0.01,the same below).Genotype AB has highly dressing percentage,lean meat percentage,loin eye height.6.The molecular breeding techniques and traditional bredding techniques were applied in high quality pig line breeding.G2 was the same as GO in reproductive traits, but G2 was better than GO in desease resistance in Large White and Duroc.Selective effection of average day gain,Lean meat percentage,fat thickness,Loin Eye area,was well.MX1 gene affected average day gain significantly,Additive effects of average day gain was significantly in Large White(p＜0.05).Selective effection of average day gain, Lean meat percentage,fat thickness,ratio of food and gain was well.MX1 gene affected average day gain and loin eye area significantly.Dominance effect of average day gain was significantly.AdditiVe effect of loin eye area was significantly in Duroc.