基于核酸适配体检测饲料中黄曲霉毒素B_(1)的方法

Study on Method for Detecting Aflatoxin B_(1) in Feed Based on Nucleic Acid Aptamer

【目的】建立一种基于核酸适配体检测饲料中黄曲霉毒素B_(1)的方法,为饲料中黄曲霉毒素B_(1)的检测提供技术支撑。【方法】利用FAM荧光标记的核酸适配体与黄曲霉毒素B_(1)的特异性结合,而与偶联BHQ1的互补序列无法配对,导致荧光值变化而实现检测。【结果】在优化条件下,链亲和素浓度为100μg/mL,核酸适配体浓度为250 nmol/L,互补序列浓度为500 nmol/L,黄曲霉毒素B_(1)在0.01~10.0 ng/mL范围具有较好的线性关系,最低检测限为0.01 ng/mL;与黄曲霉毒素B 2、黄曲霉毒素G 1、黄曲霉毒素G 2、桔霉素和赭曲霉素A的交叉反应率均较低,饲料样品中添加黄曲霉毒素B_(1)的平均回收率为91.3%~105.0%。【结论】建立的方法快速、准确、灵敏,选择性好,可用于饲料中黄曲霉毒素B_(1)的分析检测。

【Objective】The study aims to establish a method for detecting aflatoxin B_(1) in feed based on nucleic acid aptamer and to provide the technological support for detecting aflatoxin B_(1) in feed.【Method】The combination of nucleic acid aptamer labeled with FAM fluorescence and the specificity of aflatoxin B_(1) cannot match with the complementary sequence of coupling BHQ1,resulting in the variation of fluorescence value,thereby realizing the detection.【Result】Under the optimized condition,that is,100μg/mL streptavidin concentration,250 nmol/L nucleic acid aptamer concentration and 500 nmol/L complementary sequence concentration,the aflatoxin B_(1) has a good linear relation in the range of 0.01─10.0 ng/mL,the minimal detection limit is 0.01 ng/mL.The cross reaction rate with aflatoxin B 2,aflatoxin G 1,aflatoxin G 2,citrinin and ochratoxin A are all low,the average recovery of adding alfatoxin B_(1) in feed is 91.3%─105.0%.【Conclusion】The established method is rapid,accurate,sensitive and of good selectivity,which can be used in the analysis and detection of aflatoxin B_(1) in feed.