基于环介导恒温扩增和微流控芯片的生殖道支原体检测方法构建  

Construction of A Microfluidic Chip for Four Genital Mycoplasmas Based on Loop-mediated Isothermal Amplification

作　　者：张薇 李丽娟 荣长仙 方虹舒[1] ZHANG Wei;LI Lijuan;RONG Changxian;FANG Hongshu(Clinical Laboratory,Chongqing Emergency Medical Center,Chongqing 400014,China;Department of Obstetrics and Gynecology,Chongqing Emergency Medical Center,Chongqing 400014,China)

机构地区：[1]重庆市急救医疗中心检验科,重庆400014 [2]重庆市急救医疗中心妇产科,重庆400014

出　　处：《中国皮肤性病学杂志》2021年第3期343-348,共6页The Chinese Journal of Dermatovenereology

基　　金：重庆市科学技术委员会资助项目(cstc2018jscx-msybX0107)。

摘　　要：目的建立一种结合环介导恒温扩增和微流控芯片技术检测生殖道支原体的方法。方法针对4种生殖道支原体包括解脲脲原体(UU)、微小脲原体(UP)、生殖支原体(MG)、人型支原体(MH)的DNA序列进行特异性引物设计和合成,制备微流控芯片,配制恒温扩增反应体系后进行芯片加样,于核酸分析仪中进行核酸扩增和结果判读,并对该芯片进行准确性、灵敏度、特异性和重复性验证。结果成功制备的4种生殖道支原体的微流控芯片具有较好的准确性、灵敏度、特异性和重复性,整个检测过程在1 h内即可完成,UU、MG、MH的最低检测限为1000拷贝/μL,而UP的最低检测限为5000拷贝/μL。结论基于环介导恒温扩增技术的4种生殖道支原体微流控芯片具有快速、准确、高通量等优点,在临床上具有广泛的应用前景。Objective To establish a microfluidic chip method for four genital mycoplasmas based on loop-mediated isothermal amplification(LAMP).Methods Specific primers were designed and synthesized of four genital mycoplasmas including Ureaplasma urealyticum(UU),Ureaplasma parvum(UP),Mycoplasma genitalium(MG)and Mycoplasma hominis(MH).The microfluidic chip were constructed,the samples were amplificated by PCR.The accuracy,sensitivity,specificity and repeatability of the results were also verified.Results The microfluidic chip established successfully,and it had good accuracy,sensitivity,specificity and repeatability,and the detection process was finished within 1 hour.The detection limit of Ureaplasma urealyticum,Mycoplasma genitalium,Mycoplasma hominis were 1000 copies/μL,while that of Ureaplasma parvum was 5000 copies/μL.Conclusion The fast,accurate,high-throughput microfluidic chip method may has an extensive application for clinical diagnosis.

关 键 词：微流控芯片 环介导恒温扩增 解脲脲原体 微小脲原体 生殖支原体 人型支原体 

分 类 号：R446.5[医药卫生—诊断学]