Abstract:Enterotoxigenic Escherichia coli （ETEC）can cause serious foodborne diseases.It is important to establish a rapid and accurate detection method for food safety and human health.Cross-priming amplification（CPA）primers were designed based on the conservative sequence of ETEC heat-labile enterotoxin（LT）gene，and the reaction temperature，deoxy -ribonucleoside triphosphate （dNTPs）concentration，Bst -DNA polymerase-large fragment concentration，Mg2+ concentration，betaine concentration and reaction time were optimized.Six strains of Escherichia coli and eight other near-source bacteria were selected to be specific for CPA method.In order to evaluate the application technology of CPA，46 samples of raw pork without LT enterotoxin were randomly contaminated by diluted enterotoxin-producing Escherichia coli.The results showed that the optimum reaction temperature of CPA was 62 ℃，the concentration of dNTPs was 0.3 mol/L，the concentration of Bst-DNA polymerase-large fragment concentration was 8 U/tube，the concentration of Mg2+was 2.0 mmol/L，the concentration of betaine was 0.8 mol/L，and the reaction time was 45 mins.The minimum detection rate of enterotoxigenic Escherichia coli by CPA was 40 cfu/mL，which was two times higher than loopmediated isothermal amplification（LAMP）.The results of CPA assay in 7 positive samples（15.2%）were consistent with those of LAMP and real-time fluorescence quantitative PCR.