摘要
DNA环介导等温扩增(LAMP)方法是一种新型的核酸扩增技术,该方法是在等温的条件下进行的,具有反应时间短、灵敏度高、特异性强的特点。本文以产麻痹性贝毒(PSP)的亚历山大藻为研究对象,采用简易法提取DNA模板,设计特异性LAMP引物,利用LAMP技术进行产毒藻种的快速检测,同时,着重对LAMP技术与PCR技术在检测微小亚历山大藻细胞的灵敏度方面做了比较。向LAMP终产物中加入SYBRGreenI染料后可直接用肉眼观察结果而不需要通过凝胶电泳来观察。结果表明,LAMP技术在恒温65℃,1h内就可以检测到产毒藻种;LAMP技术检测微小亚历山大藻的最低检测限为200个/ml,而PCR技术的最低检测限为1000个/ml,LAMP扩增方法比PCR扩增方法的程序简单、反应时间短、灵敏度高;LAMP技术不需要精密的温度循环装置,有恒温加热设备就可以满足检测条件,可用于野外检测。
Loop-mediated isothermal amplification method （LAMP） is a novel nucleic acid amplification technology. The LAMP method amplifies DNA with rapidity, high specificity and sensitivity under isothermal conditions. This thesis took toxic Alexandrium as the major research object. DNA template was extracted by boiling method and specific primer was designed for LAMP assays. Toxic Alexandrium were quickly detected by LAMP technology. LAMP and PCR were successfully compared in the sensitivity of detecting Alexandrium minutum. Without gel electrophoresis, the LAMP amplification was directly visualized in the reaction tube by addition of SYBR Green I for a naked-eye inspection. The result indicated that toxic Alexandrium was detected by LAMP technology within 1 h under isothermal condition at 65 ℃. The lower detection limit of the LAMP reaction was 200 cell/ml, while that of the PCR was 1000 cell/ml. The LAMP assay is more simple, rapid and sensitive than the PCR. Furthermore, as LAMP technology does not require sophisticated instrumentation, it is very suitable for diagnosis in the fields.
出处
《现代食品科技》
EI
CAS
2007年第11期71-74,共4页
Modern Food Science and Technology
基金
广东省科技厅农业攻关计划(2005A11601101)
