TAP1 Gene Polymorphism And MRNA Expression Associated With Innate Immunity In Chicken

China is a big country with being rich in chicken resources and rapid development in the poultry industry. For a long time, disease control mainly rely on vaccines. Because the increasing change in variation and virulence of the virus such as MDV, the effect of using vaccine was declined.Therefore, researchers begin to focus on disease resistance breeding, hoping to improve the disease resistance of chicken. Research shows that, the polymorphism of MHC gene and resistance to a variety of infectious disease of poultry are highly relevant.Therefore MHC is the preferred marker gene for resistance breeding in poultry. TAP1 is to be located in the core position of the MHC genes, is a kind of transporters that playing an important role in disease resistance in the process of the endogenous antigens presented. TAP1 is also correlation between immune response, disease resistance and economic characters in chicken. We study 29 Chinese local breeds and foreign breeds. This research object is to study the TAP1 gene polymorphism in different breeds and determine haploid type. We also study TAP1 expression after guangxihuang chickens infected with salmonella enteritidis and TAP1 gene expression of different genotype individual, the main results were as follows:1. To study the polymorphism of TAP1 Gene,29 breeds were selected including Chinese local breeds and foreign breeds, each breeds has 10 individuals as DNA mix and sequenced TAP1 Gene. Comparing the sequencing results and the NCBI reference sequence (Gene ID: 427727), the result was that on TAP1 Gene exon, qingjiaoma exists the most number of SNPs. Wenchang chicken has only one SNP and rugaohuang chicken, xueshan chicken and taihu chicken do not exist SNP. For the foreign breeds such as Ross, recessive white chicken, anka chicken and AA chicken, they had less SNPs and INDEL than local breeds. On TAP1 Gene introns, wugu chicken, spf chicken, xueshan chicken and zangjizajiao chicken have more SNPs, and guangxihuang chicken, lvkedan chicken have the most INDEL. Foreign breeds have less mutation quantity than local varieties too. On TAP1 gene promoter, douji, gushi chicken, xiaoshan chicken, luyuan chicken and doujizajiao chicken have 10 bp deletion in position 67309, and lvkadan chicken have 20 bp. All the deletion breeds are the local varieties. The above results showed that the local breeds in our country, the TAP1 gene have abundant genetic diversity in Chinese local breeds, TAP1 gene can be used as candidate genes of innate immune research.2. Based on TAP1 gene promoter polymorphism on guangxihuang chicken for different haploid type, the results found TAP1 gene promoter is also a lack of ten bases. Guangxihuang chicken have AA, AB and BB genotype, wild (consistent with the reference sequence of red jungle fowl)and mutant allele frequency of alleles are 0.847,0.153, PIC is 0.239, the average heterozygosity is 0.278. Examined by the chi-square suitability, guangxihuang chicken populations in Hardy Weinberg equilibrium state. Predicted by the JASPAR database, there may be three transcription factor binding sites in the 10bp deletion of TAP1 gene promoter region.3. Microbial load of 1day age guangxihuang chickens infected with salmonella enteritidis 0-after 21 dpi (day-post-infection) in cecum show that group of salmonella attack reaches peak at 5 day. And microbial load increased within the period of 2 h to 5 d, then gradually reduce in 5 d to 21 d. The results cytokine levels in plasma show that the control group and infected group appeared peak at different time points. IL-1α and IFN-γ in the control group were peaked at 24 h, and infected group is a maximum concentration of 4 h. IgY of the control group concentration reaches the first peak in 24 h, in the infected group, concentration slowly continued to rise. IgM concentration change is not obvious in the control group, the concentration of basic remain unchanged. In the infected group, cytokine concentrations of IgM rise fast to peak at 2 h, and then falling fast in 2 h to 4 h. After 4 h concentration drops slowly. The results show that in guangxichicken early immune respond rapidly, time 2 h to 4 h belongs to the acute phase reaction and after 5 d belongs to a relatively stable mmune period.4. QRT-PCR was used to detect TAP1 gene mRNA expression of 1 day age guangxihuang chickens infected with salmonella enteritidis 0-21 dpi, the results showed that the TAP1 widely expressed in the four different groups in heart, liver, thymus and bursa of fabricius. Gene expression in thymus and bursa of fabricius is significantly higher than other groups (P<0.05). TAP1 gene infected group is significantly higher than the control group (P<0.05). In control group, expression of TAP1 gene increases with the increase of age and significantly increased in 3 dpi, shows that 3 d is the key to the immune state switching period. Compared different infection time and the TAP1 expression level, different organizations expressed were the highest in 3 dpi.The expression significantly increased in 1dpi and decreased after 3 dpi.This suggests that the chicken show immune reaction after salmonella infection.And TAP1 gene is involved in guangxihuang chicken immune response.5. QRT-PCR was used to study guangxihuang chicken TAP1 mRNA expression level of mutant and wild type individuals. In the control group in the same organization, mutant and wild type individuals TAP1 expression was not significantly different (P>0.05) at different time points.In infection group, TAP1 expression also has the same trend. This suggests 10 bp deletion in TAP1 promoter region in 67309 had no significant effect on the mRNA expression. And there was no significant difference in different genotype in innate immunity.