不同遗传背景猪TNF-α基因启动子区多态性与其血清含量的关联分析
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摘要
为研究不同遗传背景猪TNF-α基因启动子区多态性及其与血清含量的相关性,本实验采用克隆测序方法对3个不同遗传背景猪群体(长白山野猪民猪杂交猪,简称野杂猪、民猪和大白猪) TNF-α基因启动子区进行基因多态性检测并测序,选取特定区域采用高分辨率熔解曲线(HRM)方法分析其基因型,并分析了其不同基因型与血清TNF-α含量的相关性。测序结果显示3个群体猪TNF-α基因启动子区存在7个单核苷酸多态位点(SNP)。选取其G-1256、G-1239位点区采用HRM分析显示3个猪群体TNF-α基因启动子区存在4种基因型,其中CC基因型作为优势基因型,在野杂猪群体中该基因型频率高达73.0%,民猪次之为52.0%,大白猪最低为31.0%。差异显著性分析显示,野杂猪、大白猪群体中CC基因型个体血清TNF-α含量显著高于其它基因型个体(p<0.05)。将所有猪看做统一群体,差异显著分析显示CC基因型个体血清中TNF-α含量同样显著高于其它基因型个体(p<0.05)。转录因子结合位点预测显示CC基因型G-1239位点A构成NF-κB亚基RelB潜在转录结合位点。上述结果表明NF-κB信号通路在猪TNF-α基因转录表达调节中具有重要作用,本实验为研究影响猪群抗逆表型性状的调控通路奠定基础。
In order to investigate the association of the polymorphisms on TNF-αgene promoter region and serum concentration in different genetic background pig populations,the crossbred from wild boars in Changbai Mountain with Min pig(shorted as crossbred wild boars),Min pig and Large White pig.In this study,we tested the TNF-αgene promoter polymorphisms by genomic PCR and sequencing,the genotype frequencies of specific region were measured by High Resolution Melting(HRM)and the significant difference analysis of genotype with serum TNF-αconcentration were tested by ANOVA in these populations.The genomic PCR sequencing showed there were seven SNPs in TNF-αpromotor region.Choosing the G-1256 and G-1239 region,The HRM results showed that there were four genotypes,and as the dominant genotype,the frequency of CC genotype was73%in crossbred wild boars population,52%in Min pig and 31%in Large White pig.The significant difference tests found that the CC genotype individual had higher serum concentration in crossbred wild boars,Large White pig and whole population when the three kinds of pigs were looked as one population(p<0.05).The transcription factor(TFs)binding site prediction hinted that the G-1236 A on CC genotype sequence could construct potential RelB binding site.The RELB is one of subunits of NF-κB.The founding of polymorphism in TNF-αpromotor region and the prediction of potential RelB binding site hinted that the NF-κB signal might affect the stress resistance phenomenon in pig by regulating the transcription of TNF-αgene.
In order to investigate the association of the polymorphisms on TNF-αgene promoter region and serum concentration in different genetic background pig populations,the crossbred from wild boars in Changbai Mountain with Min pig(shorted as crossbred wild boars),Min pig and Large White pig.In this study,we tested the TNF-αgene promoter polymorphisms by genomic PCR and sequencing,the genotype frequencies of specific region were measured by High Resolution Melting(HRM)and the significant difference analysis of genotype with serum TNF-αconcentration were tested by ANOVA in these populations.The genomic PCR sequencing showed there were seven SNPs in TNF-αpromotor region.Choosing the G-1256 and G-1239 region,The HRM results showed that there were four genotypes,and as the dominant genotype,the frequency of CC genotype was73%in crossbred wild boars population,52%in Min pig and 31%in Large White pig.The significant difference tests found that the CC genotype individual had higher serum concentration in crossbred wild boars,Large White pig and whole population when the three kinds of pigs were looked as one population(p<0.05).The transcription factor(TFs)binding site prediction hinted that the G-1236 A on CC genotype sequence could construct potential RelB binding site.The RELB is one of subunits of NF-κB.The founding of polymorphism in TNF-αpromotor region and the prediction of potential RelB binding site hinted that the NF-κB signal might affect the stress resistance phenomenon in pig by regulating the transcription of TNF-αgene.
引文
[1]Bradley J R.TNF mediated inflammatory disease[J].J Pathol,2008,214(2):149-160.
[2]Arslan E,Colakoglu M,Celik C,et al.Serum TNF-alpha,IL-6,lupus anticoagulant and anticardiolipin antibody in women with and without a past history of recurrent miscarriage[J].Arch Gynecol Obstet,2004,270(4):227-229.
[3]Umapathy D,Ezhilarasi K,Mariappanadar V,et al.Increased levels of circulating(TNF-α)is associated with(-308G/A)promoter polymorphism of TNF-αgene in Diabetic Nephropathy.[J].Int J Biol Macromol,2017,107(Pt B):2113-2121.
[4]Jahid M,Rehan-Ul-Haq,Jha P K,et al.Tumor necrosis factor-α-308 polymorphism in North Indian rheumatoid arthritis patients and association with mRNA and serum TNF-α[J].Clin Rheumatol,2017,36(10):2209-2216.
[5]徐阿娟,刘小林,郭家中,等.TNF-α基因多态性及其与奶牛乳房炎的相关性分析[J].遗传,2010,32(9):929-934.
[6]Szydlowski M,Buszka A,Mackowski M,et al.Polymorphism of genes encoding cytokines IL6 and TNF is associated with pig fatness[J].Livest Sci,2011,136(2-3):150-156.
[7]张立春,李兆华,孙福亮,等.长白山野杂猪、民猪和大白猪血常规及免疫学指标比较分析[J].中国兽医学报,2018,38(1):171-176.
[8]许振英.中国一些地方猪种质特性的研究[J].中国农业科学,1985,18(2):1-13.
[9]孙丽,夏日炜,刘颖,等.大白猪肿瘤坏死因子α基因启动子区多态性及其对mRNA表达水平的影响[J].中国畜牧兽医,2015,42(6):1518-1523.
[10]牛步月,邢桂玲,查安东,等.猪肿瘤坏死因子α基因5'侧翼区域多态性分析[J].东北农业大学学报,2017,48(3):17-23.
[11]张立春,李兆华,李梦姝,等.不同遗传背景猪群体黏病毒耐药蛋白1基因多态性分析[J].中国畜牧兽医,2017,44(4):1046-1053.
[2]Arslan E,Colakoglu M,Celik C,et al.Serum TNF-alpha,IL-6,lupus anticoagulant and anticardiolipin antibody in women with and without a past history of recurrent miscarriage[J].Arch Gynecol Obstet,2004,270(4):227-229.
[3]Umapathy D,Ezhilarasi K,Mariappanadar V,et al.Increased levels of circulating(TNF-α)is associated with(-308G/A)promoter polymorphism of TNF-αgene in Diabetic Nephropathy.[J].Int J Biol Macromol,2017,107(Pt B):2113-2121.
[4]Jahid M,Rehan-Ul-Haq,Jha P K,et al.Tumor necrosis factor-α-308 polymorphism in North Indian rheumatoid arthritis patients and association with mRNA and serum TNF-α[J].Clin Rheumatol,2017,36(10):2209-2216.
[5]徐阿娟,刘小林,郭家中,等.TNF-α基因多态性及其与奶牛乳房炎的相关性分析[J].遗传,2010,32(9):929-934.
[6]Szydlowski M,Buszka A,Mackowski M,et al.Polymorphism of genes encoding cytokines IL6 and TNF is associated with pig fatness[J].Livest Sci,2011,136(2-3):150-156.
[7]张立春,李兆华,孙福亮,等.长白山野杂猪、民猪和大白猪血常规及免疫学指标比较分析[J].中国兽医学报,2018,38(1):171-176.
[8]许振英.中国一些地方猪种质特性的研究[J].中国农业科学,1985,18(2):1-13.
[9]孙丽,夏日炜,刘颖,等.大白猪肿瘤坏死因子α基因启动子区多态性及其对mRNA表达水平的影响[J].中国畜牧兽医,2015,42(6):1518-1523.
[10]牛步月,邢桂玲,查安东,等.猪肿瘤坏死因子α基因5'侧翼区域多态性分析[J].东北农业大学学报,2017,48(3):17-23.
[11]张立春,李兆华,李梦姝,等.不同遗传背景猪群体黏病毒耐药蛋白1基因多态性分析[J].中国畜牧兽医,2017,44(4):1046-1053.