Polymorphism Of The FUT1 And SLA-DQB Gene And The Basic Research On Disease Resistance For Pig Breeding

The FUT1 (Escherichia coil F18 receptor-related gene, ETECF 18 gene) and SLA-DQB gene were identified as candidate genes for disease resistance in pig, which were very important in the research field of pig breeding for disease resistance. Based on the epidemiological investigation, the aim of this study is to analyze the relationship between polymorphism of FUT1 and SLA-DQB genes and productive performance, further to probe the genetic difference of FUT1 and SLA-DQB genes between native pig breeds and foreign breeds, to explore the correlation of the F18 resistance/sensitivity in different genotypes, in order to establishment a reasonable molecular breeding scheme for F18 E.coil disease resistance in pig.The results showed:1) By the PCR-RFLP and PCR-SSCP methods, the loci of 307bp and 857bp of FUT1 gene in 14 pig breeds for polymorphism were screened respectively. The results showed that, in 307 locus, the AA genotype was only detected in Duroc and Pietrain, however, the AA genotype was not detected in Yorkshire, Landrace, cross-breeding pigs and Chinese native pig breeds. The most of indigenous pig breeds showed GG genotype, and the gene frequency ranged from 0.789 to 1.000. Meanwhile, the AG genotype only could be detected in Lingao pigs of all tested native pig breeds (the genotype frequency was 0.290). In M857 locus, AA genotype was not detected in the all pig breeds; the native pig breeds only carried only genotype of M857^GG, and showed extremely abbreviation distribution.2) M857 locus and it flanked genes from 10 individuals of genotype M307^GG and 7 individuals of genotype M307^AG were amplified by PCR, cloned into the pMD-19 vector and sequenced. A single neucleotide change from G to A (G/A mutation) was detected in the 307 locus of FUT1 gene. The mutation resulted in an amino acid residues substitution (Thr→Ala). Unexpected, we first time detected a C/T mutation in the 857 locus with an amino acid residues substitution (Arg→Ser).3) By the PCR-RFLP method, the polymorphism of exon 2 of SLA-DQB gene was analyzed in 16 pig breeds. With the analysis of restrictive enzyme HaeⅢdigestion, there were 11 genotypes and 6 alleles, and the A, B and D genes were dominant alleles, which showed abundant polymorphism in all the foreign pigs, cross-breeding pigs and native pigs. There were 10 genotypes and 6 alleles with the analysis of restrictive enzyme RsaⅠdigestion, and A and B genes were dominant alleles in foreign pigs, cross-breeding pigs and native pigs.4) SLA-DQB gene was analyzed by the combination of restrictive enzyme RsaⅠ, HaeⅢand DNA sequencing, two new genotypes DD (27/116)and FF (40/167/171) were found in this study.5) According to the gene frequency of M307 of FUT1 gene in different pig breeds, based on Nei's genetic distance, two unrooted consensus trees were constructed using both Neighbor-joining method and UPGMA method. The results showed that, all native pig breeds were clustered in a large population, all of foreign pig breeds tested formed the other cluster. The UPGMA phylogenetic dendogram showed all native pig breeds were also clustered in a large population, all of foreign pig breeds tested formed the other cluster.6) According to the gene frequency of SLA-DQB gene analyzed by the restriction enzyme HaeⅢenzyme, we constructed NJ and UPGMA phylogenetic dendograms, and the results of two dendograms showed that Lingao pig, Leping spotted pig, Erhualian pig, Fengjing pig, Pietrain pig, Xiushui hang pig, Sutai pig and Tibetan pig were always in a cluster, the other native pigs and foreign pigs were not confined into the same cluster.7) Compared the productive performances of Sutai pig with different genotypes of FUT1 gene in 307bp locus, the results showed that, the average productive performance with AG individuals was higher than that of GG individuals. And the postweaning weight at the second, the third and the sixth litter, and number of litter at the third litter were significantly higher than individuals with GG genotype.8) Analyzed the correlation of different genotypes and productive performances in Sutai pigs with the BB,BD and DD genotypes of HaeⅢdigestion analysis and Sutai pigs with the AA,AB,BB genotypes of RsaⅠdigestion analysis, the results showed significant correlation with postweaning viability in the second litter, and the significant correlation with the total litter, number of viable pigs, litter weight at birth, number of ablactation pigs and litter weight after ablactation in the third litter. The significant correlation existed in the total litter, number of ablactation pigs and litter weight after ablactation.9) The adhesion test showed that small intestinal epithelium cells of 30-35-day-old postweaning piglets with both genotype M307^GG and M307^AG could adhere to the standard E.coli strain express F18ab fimbriae, the recombinant E.coli express F18ac rE.coli1534 and the recombinant pnirBMisL-fedF E.coli displaying FedF subunit on the surface of E.coli, but small intestinal epithelium cells with genotype M307^AA and 3-day-old piglet could not adhere to the three kinds of bacteria descried above, the latter with good adherence capability of 987P fimbrial E.coli.10) In the adhesion inhibition test with piglet small intestinal epithelium cell, the results showed that, the adhesive ability of the30-35-day-old postweaning piglets small intestinal epithelium cells with two genotypes of M307^GG and M307^AG was almost lost when standard E.coli strain express F18ab fimbriae, the recombinant rE.coli1534 expressing F18ac and the recombinant pnirBMisL-fedF E.coli displaying F18ab FedF subunit incubated with the corresponding anti-serum.11) Under the guide of experimental methods and results of this study, using Sutai pig as an example, we may set up a reasonable molecular breeding schedule for F18 E.coil disease resistance in pig.