摘要
为了快速、准确检测鸡滑液囊支原体(MS),对MS标准株vlhA基因进行分析,设计并合成特异性引物和探针,优化反应条件,构建了基于vlhA基因的TaqMan实时荧光定量PCR快速检测方法。结果显示,所构建方法标准曲线所对应的相关系数R^(2)=1.000,斜率S=-3.452,截距I=43.950,表明标准质粒浓度与Ct值的线性关系良好;该方法除MS外,其他参考菌(毒)株均无目的基因扩增,其最小检测浓度为1×10^(1)copies/μL,组内重复及组间重复的变异系数均低于0.1;运用所建立方法与普通PCR方法对4份疑似MS临床样本进行检测,检测结果完全相符合;应用该方法对贵州省7个地区107份疑似MS样本进行检测,发现总体阳性率为26.17%。说明建立的方法具有较好的特异性、敏感性和重复性,可应用于临床检测。
In order to diagnose Mycoplasma synoviae(MS)rapidly and accurately,the vIhA gene of MS standard strain was analyzed,the specific primers and probes were designed and synthesized,and the reac-tion conditions were optimized.A TaqMan real-time quantitative PCR method based on vlhA gene was es-tablished.The results showed that correlation coefficient R^(2) was 1.000,slope S was-3.452,and intercept I was 43.950,indicating a good linear relationship between standard plasmid concentration and CT value.The minimum detection concentration was 1×10^(1)copies/μL.The coefficient of variation of intra-and inter-group replicates was lower than 0.1.The established method and common PCR were used to detect 4 sus-pected MS clinical samples,and the results were consistent.This method was applied to 107 suspected MS samples from 7 regions of Guizhou province,and the overall positive rate was 26.17%.The established method has good specificity,sensitivity and repeatability,and can be applied to clinical detection.
作者
李梅
王柏林
岳筠
杨美
宋春
朱二鹏
单春兰
文明
程振涛
LI Mei;WANG Bo-ling;YUN Jun;YANG Mei;SONG Chun;ZHU Er-peng;SHAN Chun-lan;WENG Ming;CHENG Zhen-tao(College of Animal Science,Guizhou University,Guiyang,Guizhou,550025,China;Guizhou Provincial Key Laboratory of Animal Diseases and Veterinary Public Health,Gui yang,Guizhou,550025,China;Guizhou Animal Disease Prevention and Control Center,Guiyang,Guizhou,50008,China)
出处
《动物医学进展》
北大核心
2023年第3期15-20,共6页
Progress In Veterinary Medicine
基金
国家自然科学基金项目(32060786,31660723)
贵州省科学技术项目(黔科合基础[2019]1181号)
贵州省科技支撑计划项目(黔科合支撑[2021])
贵州省优秀青年科技人才项目(黔科合平台人才[2021]5646)。
