Application of a Loop-mediated Isothermal Amplification Method for Rapid Diagnosis of Francisella tularensis

doi:10.13560/j.cnki.biotech.bull.1985.2018-0573

Abstract

Abstract: This study aims to apply the loop-mediated isothermal amplification（LAMP）to detect Francisella tularensis. LAMP primers were designed for amplifying FopA gene from F. tularensis,reaction conditions were optimized,and the specificity and sensitivity of the method were evaluated. Results showed that：（1）12 bacteria and tick’s DNA（Containing a large number of Pseudomonas bacteria）were amplified with LAMP reaction;however,only F. tularensis demonstrated positive reaction;（2）the detection limit of pure F. tularensis culture was 4.9×10²pg/mL,the minimum threshold for cloned plasmid was 10 copies/μL,which were below those of conventional PCR,1×10³copies/μL;（3）the minimum threshold for simulated soil samples was 44 CFU/g;（4）24 soil samples（8 dilutions）contaminated by F. tularensis were compared with the imported fluorescence quantitative PCR reagent,and all of them were accurately detected in the sensitivity range. In conclusion,applying LAMP technology in the detection of F. tularensis shows superiority of efficiency,high specificity and sensitivity,thus this has practical value for etiological diagnosis of tularemia.

Key words: Francisella tularensis, gene FopA, LAMP, rapid detection technology

LI Zi-wei, DENG Zhong-liang. Application of a Loop-mediated Isothermal Amplification Method for Rapid Diagnosis of Francisella tularensis[J]. Biotechnology Bulletin, 2019, 35(2): 212-217.

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