Maruyama Fumito, Kenzaka Takehiko, Yamaguchi Nobuyasu, Tani Katsuji, Nasu Masao
Graduate School of Pharmaceutical Sciences, Osaka University, Osaka 565-0871, Japan.
Appl Environ Microbiol. 2003 Aug;69(8):5023-8. doi: 10.1128/AEM.69.8.5023-5028.2003.
A new in situ DNA amplification technique for microscopic detection of bacteria carrying a specific gene is described. Loop-mediated isothermal amplification (LAMP) was used to detect stxA(2) in Escherichia coli O157:H7 cells. The mild permeabilization conditions and low isothermal temperature used in the in situ LAMP method caused less cell damage than in situ PCR. It allowed use of fluorescent antibody labeling in the bacterial mixture after the DNA amplification for identification of E. coli O157:H7 cells with an stxA(2) gene. Higher-contrast images were obtained with this method than with in situ PCR.
本文描述了一种用于显微镜检测携带特定基因细菌的新型原位DNA扩增技术。环介导等温扩增（LAMP）用于检测大肠杆菌O157:H7细胞中的stx A(2)。原位LAMP方法中使用的温和通透条件和较低等温温度比原位PCR造成的细胞损伤更少。它允许在DNA扩增后对细菌混合物进行荧光抗体标记，以鉴定带有stx A(2)基因的大肠杆菌O157:H7细胞。与原位PCR相比，该方法获得的图像对比度更高。
