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鸡固有免疫MDA5信号通路相关分子的实时荧光定量PCR检测方法的建立 被引量:2

Development of Real Time Fluorescence Quantitative PCR Method for Detection of the Innate Immune MDA5 Signaling Pathway Related Molecules in Chicken
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摘要 固有免疫中的MDA5信号通路是机体识别入侵胞内病原体的重要途径之一,其中MDA5、LGP2和MAVS在识别入侵病原体、激活固有免疫下游的转录因子、诱导干扰素、各种细胞因子及炎症因子的产生中发挥重要作用。本研究根据NCBI已发表的鸡MDA5、LGP2和MAVS基因序列设计引物,构建重组质粒作为标准品,在荧光定量PCR仪上建立标准曲线,并进行灵敏度、重复性和特异性试验。结果显示,成功建立检测鸡MDA5、LGP2和MAVS mRNA表达的实时荧光定量PCR检测方法,此方法具有快速、特异性强、灵敏度高、线性范围广、稳定性高的特点,为快速检测和定量分析鸡源MDA5、LGP2和MAVS表达提供技术平台,也为进一步探究鸡相关疾病的固有免疫机制奠定了技术基础。 MDA5 signaling pathway is an important method for organisms to identify intracellular pathogens invading in innate immune. Meanwhile, MDA5, LGP2 and MAVS play a critical role in the recognition of cytoplasm pathogens and transmitting the signal to the transcription factors which then are activated. As a result, the interferon, cytokines and inflammatory cytokines are induced. In the present study, primers were designed according to the chicken MDA5, LGP2 and MAVS gene sequences published in NCBI GenBank. The recombinant plasmids were constructed as standard for establishing the standard curves. The sensitivity,reproducibility and specificity of established real-time quantitative PCR were assessed. The results showed that the real-time fluorescence quantitative PCR method for detecting the expression of MDA5,LGP2 and MAVS mRNA were efficient,specific,sensitive,wide linear range and stable. This method would provide technical foundation for the rapid detection and quantification of chicken MDA5, LGP2 and MAVS mRNA expression,for further exploring the innate immune mechanism in related avian diseases.
出处 《中国家禽》 北大核心 2014年第17期15-19,共5页 China Poultry
基金 国家自然科学基金项目(31360611) 人力资源和社会保障部留学人员科技活动项目择优资助经费(人社厅函[2012]258号) 广西自然科学基金项目(2014GXNSFDA118011 2013GXNSFCA019010)
关键词 MDA5 LGP2 MAVS 实时荧光定量PCR chicken MDA5 LGP2 MAVS real-time fluorescence quantitative PCR
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