霍乱弧菌核酸标准物质定值方法:ICP-MS法和数字PCR法

Quantification of Plasmid DNA Reference Material for Vibrio Cholerae by Using ICP-MS and Digital PCR

霍乱弧菌是人类霍乱的病原体,霍乱是一种古老且流行广泛的烈性传染病之一。曾在世界上引起多次大流行,主要表现为剧烈的呕吐、腹泻、失水,死亡率甚高,属于国际检疫传染病。霍乱弧菌定量检测质粒标准物质,霍乱弧菌的外膜蛋白编码基因omp W为靶基因,可作为霍乱弧菌核酸扩增(PCR)荧光检测试剂盒中的阳性物质替代物。构建霍乱弧菌质粒DNA标准样品,采用电感耦合等离子体质谱(ICP-MS)方法和数字PCR两种方法定值,最终定值结果为(79. 2±5. 2) ng/μL,可为霍乱弧菌核酸检测的量值准确性、可比性和有效性提供保障。

Vibrio cholerae is the pathogen of human cholera,and cholera is an ancient and widespread infectious disease.It as an international quarantine infectious disease has caused many pandemics in the world,and mainly manifested as severe vomiting,diarrhea,dehydration,and high mortality rate.Vibrio cholerae plasmid reference material (RM),containing a target gene coding the outer-membrane protein ompW of Vibrio cholerae.It can be used as a positive substance substitute in polymerase chain reaction (PCR)fluorescence detection reagent kit for Vibrio cholerae.Plasmid DNA reference material for Vibrio cholerae analysis and performed the quantification by using two different methods:inductively coupled plasma mass spectrometry (ICP-MS)and digital PCR.The result was (79.2±5.2)ng/μL.Reference material will play important role for the accuracy,comparability,and effectiveness of the Vibrio cholerae quantification.