Rapid Detection of Salmonella spp. by Loop-Mediated Isothermal Amplification Method
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Abstract:
Loop-mediated isothermal amplification (LAMP) is a novel DNA amplification technology which amplifies DNA with high specificity, efficiency, and rapidity under isothermal conditions. According to the specific invA gene of Salmonella spp., a set of specific primers were designed to amplify the special DNA sequences by LAMP. Moreover, the reaction conditions of LAMP were optimized. The results showed that the optimal concentrations of inner primers, exogenous primers, Mg2＋, dNTP, betaine, Bst DNA polymerase, reaction temperature and reaction time were 40 pmol/L, 5 pmol/L, 6 mmol/L, 0.8 mmol/L, 0.8 mmol/L, 8 U, 63 ℃ and 1h, respectively. Under those conditions, the detection limit for Salmonella spp. DNA template was 10 fg/tube and no cross reaction with other usual bacteria was found. For Salmonella spp. in milk products, the detection limit of the method was 102 cfu/mL. So this method was suitable for rapid detection of Salmonella spp. in contaminated foods.
