Development of genetic diagnostic method of Shigella based on loop-mediated isothermal amplification

Objective To develop a method of loop-mediated isothermal amplification (LAMP) for detection of Shigella.Methods According to conserved nucleotide of Shigella ipaH gene and principle of LAMP, we designed a set of LAM Preaction system containing four primers.Twenty-two strains of Shigella and non-Shigella were detected with LAMP to evaluate the specificity of the method.The dilution of the ipaH gene recom binant plasmid was detected with PCR and LAMP to evaluate the sensitivity of the method.Inaddition, we evaluated the linearity between initial template copys Log value and reaction time to explore the feasibility of quantitative determination.Results The LAMP assay could be finished with in 1 hour and was no cross-reaction with other closely related members of food pathogens.The detectability was 200 copies/reaction and was 10 times more sensitive than PCR.The coefficient of variance between tests was less than 5%.The kinetics curves showed a good linearity between initial template copys Log value and reaction time with a correlation coefficient of 0.985 5.Conclusion The genetic diagnostic method of Shigella based on LAMP is rapid, sensitiv e and specific for detection of Shigella.