| 摘要: |
| 【目的】探究哈萨克羊诱导型一氧化氮合酶基因（iNOS）多态性与布鲁菌易感性的关系。【方法】使用虎红平板凝集试验（RBPT）进行哈萨克羊布鲁菌病血清学检测。参考GenBank中绵羊iNOS基因序列，针对其2,4,5外显子及其邻近内含子片段（exon 2/intron,exon 4/intron and exon 5/intron）设计引物，采用PCR-SSCP和DNA测序技术对231只哈萨克羊的iNOS基因进行多态性检测，分析其单核苷酸多态性（SNPs）与布鲁菌易感性之间的关系。【结果】经检测发现67只哈萨克羊为布鲁菌感染阳性，阳性检出率为29.00%。在哈萨克羊iNOS基因的exon 2/intron片段上检测出F2-C1910T多态位点，检测到CC、CT 2种基因型，优势等位基因和基因型分别是C和CC，其频率分别是0.924和0.848。在exon 4/intron片段上检测出F4-T11519C和F4-A11546G两个多态位点，其中F4-T11519C位点有2个等位基因T、C，有TT、CC、TC 3种基因型，优势等位基因和基因型分别是T和TC，其频率分别是0.714和0.528；F4-A11546G位点有2个等位基因A、G，有AA、AG、GG 3种基因型，优势等位基因和基因型分别是A和AG，其频率分别是0.714和0.528。在exon 5/intron片段上检测出F5-C12707T位点，有2个等位基因C、T，有CC、TT、CT 3种基因型，优势等位基因和基因型分别是C和CT，其频率分别是0.600和0.506。F2-C1910T位点为低度多态性位点，F4-T11519C、F4-A11546G和F5-C12707T位点为中度多态性位点。χ2检验表明，哈萨克羊iNOS基因F2-C1910T和F5-C12707T位点与布鲁菌易感性的相关性不显著（P＞0.05），F4-T11519C和F4-A11546G位点与布鲁菌易感性的相关性极显著（P＜0.01）。哈萨克羊iNOS基因F2-C1910T位点的易感基因型为CC，F4-T11519C位点的易感基因型为TT、TC，F4-A11519G位点的易感基因型为AA、AG，F5-C12707T位点的易感基因型为CC、CT。【结论】哈萨克羊iNOS基因F2-C1910T和F5-C12707T位点与布鲁菌易感性无相关性，F4-T11519C和F4-A11546G位点与布鲁菌易感性存在相关性。 |
| 关键词: 哈萨克羊 iNOS基因 PCR-SSCP SNPs 布鲁菌 |
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| 基金项目:国家重点基础研究发展计划（“973”计划）项目“重要人兽共患胞内寄生菌病流行特征及病原致病机制研究”（2010CB530200） |
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| Relationship between iNOS gene polymorphism and Brucella susceptibility in Kazak sheep |
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XU Jie,LI Gang,ZHANG Fan,et al
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| Abstract: |
| 【Objective】This study aimed to explore the relationship between inducible nitric oxide synthetase (iNOS) gene polymorphism and Brucella susceptibility in Kazakh sheep.【Method】RBPT was used to detect brucellosis in Kazakh sheep.Referring to the sequence of sheep iNOS gene in GenBank,primers were designed for exon 2/intron,exon 4/intron and exon 5/intron.The polymorphism of iNOS gene in 231 Kazakh sheep was detected by PCR-SSCP and DNA sequencing,and the relationship between SNPs and susceptibility of Brucella Kazakh was analyzed.【Result】A total of 67 Kazak sheep were positive for Brucella infection with positive rate of 29.00%.F2-C1910T polymorphism was detected on exon 2/intron segment of iNOS gene in Kazak sheep.Two genotypes (CC,CT) were detected and the dominant allele and genotype were C and CC with frequencies of 0.924 and 0.848,respectively.F4-T11519C and F4-A11546G polymorphisms were detected on exon 4/intron fragment.F4-T11519C locus had two alleles T and C and three genotypes (TT,CC,TC),and the dominant allele and genotype were T and TC with frequencies of 0.714 and 0.528.F4-A11546G locus had two alleles A and G and three genotypes (AA,AG,GG),and the dominant allele and genotype were A and AG with frequencies of 0.714 and 0.528.F5-C12707T was detected on exon 5/intron fragment,with two alleles C and T and three genotypes (CC,TT,CT).The dominant allele and genotype were C and CT with frequencies of 0.600 and 0.506.F2-C1910T was a low-level polymorphism,while F4-T11519C,F4-A11546G and F5-C12707T were moderate polymorphisms.The χ2 test showed that there was no significant correlation between F2-C1910T and F5-C12707T polymorphisms of iNOS gene and Brucella susceptibility (P>0.05),while F4-T11519C and F4-A11546G polymorphisms were highly correlated with Brucella susceptibility (P<0.01).The susceptible genotype of iNOS gene in Kazakh sheep was CC at F2-C1910T,TT and TC at F4-T11519C,AA and AG at F4-A11519G,and CC and CT at F5-C12707T.【Conclusion】There was no correlation between F2-C1910T and F5-C12707T polymorphisms of iNOS gene and Brucella susceptibility in Kazakh sheep.F4-T11519C and F4-A11546G polymorphisms were correlated with Brucella susceptibility. |
| Key words: Kazakh sheep iNOS gene PCR-SSCP SNPs Brucella |
