白介素6基因序列单核苷酸多态与京海黄鸡柔嫩艾美尔球虫抗性的关联分析

发布时间：2018-04-22 04:05

本文选题：柔嫩艾美尔球虫 + IL-6基因　；参考：《扬州大学》2017年硕士论文

【摘要】：本研究以京海黄鸡为实验材料,采用实时荧光定量法PCR法检测IL-6、IL-8(CXCLi2)、CCLi2基因在鸡柔嫩艾美尔球虫攻毒和非攻毒群体间的组织表达谱差异,研究3个基因在球虫感染中所起作用。进一步通过DNA测序技术,检测炎性细胞因子IL-6(白介素6)基因上游-2200bp至下游500bp区域中的单核苷酸多态(single nucleotide polymorphism,SNP),并分析所测SNPs及其单倍型与京海黄鸡柔嫩艾美尔球虫抗性指标之间的关系,为京海黄鸡抗病育种提供分子遗传标记。主要研究结果如下:1、IL-6基因在各组织中均有表达,感染组IL-6基因在盲肠、胸腺、肾脏中表达量极显著高于对(P0.01),在法氏囊中表达量显著高于对照组(P0.05);IL-8基因在各组织中均有表达,在腺胃、盲肠和胸腺中表达量极显著的高于对照组(P0.01),在脾和小肠中表达量显著高于对照组(P0.05);CCLi2在各组织中均有表达,感染组在脾和胸腺中表达量极显著高于对照组(P0.01),在腺胃、盲肠、肾脏中显著高于对照组(P0.05)。2、以IL-6为候选基因测序共发现了 28个突变位点,与GenBank上进行比较,新发现了 4个突变位点。其中位于5'调控区的单核苷酸突变有19个,除去GenBank有记录的SNPs,新发现3处突变;位于内含子的SNPs有2个;位于外显子的SNPs有2个,均为同义突变;位于3'区的SNPs有5个。3、本试验对京海黄鸡IL-6基因28个SNPs位点与抗性指标进行关联分析,发现其中有13个位点与抗性指标有关联。其中与丙二醛(Malondialdehyde,MDA)有显著关联的SNPs有3个,分别为1、6、25号变异位点;与白介素2有显著关联的SNPs有6个,分别是7、12、16、17、20、21号变异位点;与谷胱甘肽过氧化物酶(Glutathione peroxidase,GSH-Px)有关联的SNPs有3个,分别是12、15、17位点;14号位点SNP与一氧化氮有显著关联;与白介素17有显著关联的SNPs位点是17和19;与过氧化氢酶(Catalase,CAT)有显著关联的SNPs位点为19和28;19号位点与白介素16有显著相关。4、对5'调控区与抗性指标有关的9个SNPs位点进行单倍型分析:H2H3单倍型组合IL-17的浓度显著高于其他单倍型组合的浓度(P0.05);H2H5单倍型组合GSH-Px浓度显著高于H1H1、H1H2和H1H5单倍型组合(P0.05);H1H2、H1H4、H2H5单倍型组合MDA浓度显著的高于单倍型组合H1H1、H1H5、H2H3(P0.05);单倍型组合H2H5IL-2浓度显著的高于单倍型组合H1H1、H1H2和H1H5。对内含子和3'端与抗性指标有关的4个SNPs位点进行单倍型分析:H2'H3'单倍型组合的NO浓度显著的低于H1'H3'单倍型组合(P0.05),CAT浓度显著的低于H2'H2'单倍型组合(P0.05);H2'H2'单倍型组合超氧化物歧化酶(Superoxide Dismutase,SOD)浓度显著高于H2'H3'、H3'H3'单倍型组合(P0.05),H1'H5'单倍型组合的SOD浓度显著高于H3'H3'单倍型组合(P0.05);单倍型组合H1'H2'、H2'H3'IL-2的浓度显著的高于H1'H1,单倍型组合(P0.05),IL-16的浓度显著高于单倍型组合H1'H3'、H1'H5'(P0.05)。
[Abstract]:In this study, the tissue expression profiles of IL-6, IL-8, CXCLi2 and CCLi2 genes were detected by real-time fluorescence quantitative PCR method in Jinghai Yellow Chicken, and the effects of three genes on the infection of coccidia were studied. Further through DNA sequencing, The single nucleotide polymorphisms (SNPs) in the upstream to downstream 500bp region of the inflammatory cytokine IL-6 (IL-6) gene were detected. The relationship between the SNPs and its haplotypes and the resistance index of Eimeria tenella was analyzed. To provide molecular genetic markers for resistance breeding of Jinghai Yellow Chicken. The main results were as follows: the expression of IL-6 gene was significantly higher in the cecum, thymus and kidney than that in the control group, and the expression of IL-8 in the bursa of Fabricius was significantly higher than that in the control group. The expression of CCLi2 in the glandular stomach, caecum and thymus was significantly higher than that in the control group (P 0.01), and in the spleen and small intestine was significantly higher than that in the control group. The expression of CCLi2 in the spleen and thymus of the infected group was significantly higher than that in the control group (P 0.01), and in the glandular stomach, the expression of CCLi2 in the spleen and thymus was significantly higher than that in the control group. In the cecum, kidney was significantly higher than that in the control group (P0.05. 2). A total of 28 mutation sites were identified using IL-6 as a candidate gene. Compared with GenBank, 4 new mutation sites were found. There were 19 single nucleotide mutations in the 5 'regulatory region, 3 new mutations except for those recorded in GenBank, 2 in intron SNPs, 2 in exon SNPs, and 2 in exon, all of which were synonymous mutations. There are 5 SNPs in 3 'region. In this experiment, 28 SNPs loci of IL-6 gene and resistance index of Jinghai Yellow chicken were analyzed, and 13 of them were found to be related to resistance index. Among them, there were three SNPs with significant association with malondialdehyde malondialdehyde (malondialdehyde), six SNPs with significant association with interleukin 2, and three SNPs with glutathione peroxidase glutathione peroxidase glutathione peroxidase glutathione peroxidase glutathione peroxidase glutathione peroxidase glutathione peroxidase glutathione peroxidase glutathione peroxidase glutathione peroxidase glutathione peroxidase glutathione peroxidase glutathione peroxidase glutathione peroxidase glutathione peroxidase glutathione peroxidase GSH-Px. There was a significant correlation between SNP and nitric oxide at site 14. The number of SNPs sites significantly associated with interleukin 17 was 17 and 19; the number of SNPs sites with significant correlation with catalase catalase was 19 and 281.There was a significant correlation between IL-16 and IL-16 at locus 19, and 9 SNPs loci associated with resistance index for 5 'regulatory region. Haplotype analysis: the concentration of IL-17 in haplotype combinations was significantly higher than that in other haplotype combinations P0.05H2H5 haplotype GSH-Px was significantly higher than that of H1H1H1H2 and H1H5 haplotype combinations P0.05H2H2H2H5 haplotype combinations were significantly higher than that of haplotype combinations. The concentration of H2H5IL-2 in haplotype combination was significantly higher than that in haplotype combination H _ 1 H _ 1 and H _ 1 H _ 2 and H _ 1 H _ 5. The H2H5IL-2 concentration of haplotype combination was significantly higher than that of haplotype combination H _ 1 H _ 1 and H _ 1 H _ 5. Haplotype analysis of four SNPs loci related to resistance index in intron and 3 '-terminal haplotype: the concentration of no was significantly lower than that of H1 / H3' haplotype combination (P0.05) and the concentration of cat was significantly lower than that of H2H2'haplotype combination P0.05H2H2'haplotype combination P0.05H2H2'haplotype combination (P0.05H2H2'haplotype combination), which was significantly lower than that of H2H3'haplotype combination (P0.05H2' haplotype combination). The SOD concentration of H2H2H2H3H3H3'haplotype combination was significantly higher than that of H3H3H3'haplotype combination P0.05'H3H3'haplotype (P0.05N), and the concentration of H2H2H2H3H3H3H3H _ (3) H2H3H3H3H _ (3) (H2H2H3H3H _ (3)) was significantly higher than that of the haplotype combination (H0.05H _ (0. 05) IL-16), and the concentration of IL-16 was significantly higher than that of the haplotype combination (H0. 05).
【学位授予单位】：扬州大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：S858.31

【参考文献】