鸡鸭源性成分荧光LAMP检测方法的建立及应用

Establishment and Application of Fluorescent Loop-Mediated Isothermal Amplification for Detecting Chicken or Duck Derived Ingredients

为快速鉴别牛羊肉中是否掺杂了鸡肉、鸭肉等其他肉类,基于Genie II等温扩增荧光检测系统,建立了检测鸡鸭源性成分的荧光LAMP检测方法。同时,优化了反应体系和条件,进行了特异性和灵敏度试验,以及市场检测应用。结果显示:鸡鸭源性成分荧光LAMP检测方法具有很强的特异性,除了相对应的动物源性成分外,其他13种动物源性成分均呈阴性;该方法检测鸡、鸭源性成分的灵敏度分别为0.88、3.32 pg/μL,均高于普通PCR 10倍;加入双加速引物,该方法的反应时间仅为15~20 min,10 min左右时就可进行实时初判;市场应用检测结果与普通PCR的符合率为100%。结果表明,本方法特异、灵敏,所需时间短,可用于鸡鸭源性成分的市场快速检测。

In order to rapidly detect whether the chicken and duck meat were mingled in beef and mutton,a fiuorescent LAMP technology for detecting chicken or duck derived ingredients were developed based on the Genie II real-time fiuorescence isothermal amplification detection system. Meanwhile,the reaction system and conditions were optimized,and the specificity and sensitivity were tested,then the LAMP technology was applied to market detection. The results showed that the fiuorescent LAMP technology was very specific,except ingredients derived from chicken or duck,the detection results of other 13 animal derived materials were all negative. According to the sensitivity test,the lowest limits of chicken or duck meat that could be detected were 0.88、3.32 pg/μL respectively,which were 10 folds higher than that of conventional PCR. By adding the loop primes,the reaction time was only about 15-20 min,and the preliminary assessment could be conducted after 10 min of reaction. In addition,the coincidence rate was 100% between detection results of market application and conventional PCR. In conclusion,the LAMP technology was sensitive,specific and short time-consuming,and could be used in rapid detection of ingredients derived from chicken or duck.