三重荧光定量PCR检测水产动物源细菌磺胺类耐药基因

发布时间：2018-04-29 01:21

本文选题：水产动物源细菌 + 磺胺类耐药基因　；参考：《湖南农业大学学报(自然科学版)》2017年01期

【摘要】：根据Gen Bank中细菌磺胺类耐药基因Sul1、Sul2和Sul3的保守序列设计3对特异性引物及相应Taq Man探针,以重组质粒标准品为模板对反应条件进行优化,建立一种可同时检测上述3种耐药基因的三重荧光定量PCR方法,并对该方法进行敏感性、特异性、重复性及临床菌株检测试验。结果表明:该方法定量范围为1×10~1~1×10~8拷贝/反应时,其标准曲线呈良好的线性关系;该方法灵敏度高(3个基因的最低质粒检测限均可达到10拷贝/反应)、特异性强(与其他病原菌及病毒无交叉反应)、重复性好(变异系数均小于2%);对临床菌株的检测结果与药敏试验结果的符合率达94.2%,且整个检测过程可在2 h内完成。所建立的可同时检测Sul1、Sul2和Sul3基因的三重荧光定量PCR方法可用于临床检测细菌的磺胺类耐药基因。
[Abstract]:Three pairs of specific primers and corresponding Taq Man probes were designed according to the conserved sequences of sulfamethylamine Sulfur2 and Sul3 in Gen Bank, and the reaction conditions were optimized by using the standard plasmid as template. A triple fluorescence quantitative PCR method was developed for the simultaneous detection of the three drug resistant genes, and the sensitivity, specificity, reproducibility and clinical strains were tested. The results showed that the standard curve showed a good linear relationship when the quantitative range of the method was 1 脳 10 ~ (-1) ~ 1 脳 10 ~ (8) copy / reaction. This method has high sensitivity (the lowest plasmid detection limit of three genes can be up to 10 copies / reaction), strong specificity (no cross reaction with other pathogens and viruses, good reproducibility (coefficient of variation less than 2%), and detection of clinical strains The coincidence rate between the test results and the drug sensitivity test results was 94.2%, and the whole detection process could be completed within 2 hours. The triplex fluorescence quantitative PCR method for simultaneous detection of Sul1-Sul2 and Sul3 genes can be used in clinical detection of sulfanilamide-resistant genes in bacteria.
【作者单位】：广西水产科学研究院;
【基金】：广西水产畜牧科技项目(桂渔牧科201528044) 广西科技攻关项目(桂科攻1355010 3)
【分类号】：S941.4

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