环介导等温扩增法快速检测乳中阪崎肠杆菌

Rapid detection of Enterobacter sakazakii in milk by loop-mediated isothermal amplification

利用环介导等温扩增方法的快速、简便等优点,建立一种检测乳中阪崎肠杆菌的方法。以阪崎肠杆菌的OmpA序列为靶基因,设计特异性引物。优化并建立LAMP检测乳中阪崎肠杆菌的方法。结果表明,LAMP检测阪崎肠杆菌纯培养物的灵敏度为3.7×101cfu/mL,其灵敏度是PCR方法的10倍。人工污染阪崎肠杆菌灭菌乳的检测限为4.3×101cfu/mL。对23株致病菌进行特异性实验,特异性良好。该方法具有特异性强、灵敏度高、设备简便、耗时短等优点,在食品检测中具有良好的应用前景。

In order to rapidly and conveniently detect Enterobacter sakazakii in milk,a new method was developed by loop-mediated isothermal amplification （LAMP）.The LAMP primers were designed on the basis of outer membrane protein A gene （OmpA）in Enterobacter sakazakii. The LAMP reaction mixture was optimized. The determination limit of LAMP when testing E sakazakii templates in pure culture was 3.7 ×10^1cfucfu/mL, and its sensitive was better than PCR.For dairy samples, LAMP could specifically detect E.sakazakii cells as low as 4.3 ×10^1cfucfu/mL.Only four Enterobacter sakazakii strains were amplified using LAMP,and no amplicon was observed in other 19 bacteria strains.Therefore LAMP method was an effective technology with high specificity and sensitivity,to conveniently detect E.sakazakii in milk.