Detection of Escherichia coli O157 by Reverse Transcriptase Loop-Mediated Isothermal Amplification

doi:10.13560/j.cnki.biotech.bull.1985.2017-0932

Abstract

Abstract: A reverse transcriptase loop-mediated isothermal amplification（RT-LAMP）method was developed to specifically detect Escherichia coli O157. A number of primers were designed for specific conservative rfbE gene in E. coli O157. A real-time fluorescent RT-LAMP for detecting E. coli O157 was established by optimizing primer selection and reaction conditions. The specificity and sensitivity of the method were verified by using E. coli O157 and its artificial contaminated skim milk samples，and compared with the sensitivity of rRT-PCR（real-time fluorescent quantitative reverse transcription polymerase chain）. The RT-LAMP reaction was completed within 30 min under the condition of isothermal temperature of 65℃. RT-LAMP was highly specific since there was no specific amplification except two strains of E. coli O157. In the detection of artificial contaminated skim milk samples，the sensitivity reached 20 CFU/g，10 times of that by the rRT-PCR，meaning this method was highly sensitive. The established real-time fluorescent RT-LAMP will provide a powerful means for rapid detection of E. coli O157 on site.

Key words: reverse transcriptase loop-mediated isothermal amplification, Escherichia coli O157, detection, skim milk

LIANG Yu-lin, LIU Xiu ,ZHOU Peng-fei ,ZHOU Zhen-sen, YIN Jian-jun. Detection of Escherichia coli O157 by Reverse Transcriptase Loop-Mediated Isothermal Amplification[J]. Biotechnology Bulletin, 2018, 34(6): 59-65.

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