基于核酸适配体检测玉米中的赭曲霉毒素A

Determination of ochratoxin A in corn based on aptamer

目的建立一种基于核酸适配体检测玉米中赭曲霉毒素A(ochratoxin A,OTA)的方法。方法以微孔板为载体,采用偶联生物素和Cy3荧光标记的核酸适配体与OTA的特异性结合,而与偶联黑洞淬灭探针(black hole quencher 2,BHQ2)的互补序列无法配对,导致荧光值变化从而实现对OTA的定量检测。结果优化的条件为链亲和素质量浓度为100μg/mL,核酸适配体浓度为200 nmol/L,互补序列浓度为400 nmol/L,OTA在0.05-10.00 ng/mL范围具有较好的线性关系。与赭曲霉毒素B、黄曲霉毒素B1、脱氧雪腐镰刀菌烯醇和玉米赤霉烯酮的交叉反应率均低于1%,玉米样品中添加OTA的平均回收率为89.0%-93.8%。结论该方法快速、准确、灵敏,交叉反应率低,可用于样品中OTA的分析检测。

Objective To establish a method for the determination of ochratoxin A(OTA) in corn based on aptamer.Methods In microplate,biotin and Cy3 labeled aptamer could bind specifically to OTA and not hybridize with the complementary strand which labeled with black hole quencher 2(BHQ2),and it leaded to the change of fluorescence value,so as to realize the quantitative detection of OTA.Results The optimization of parameters were that the mass concentration of streptavidin was 100 μg/mL,aptamer concentration was 200 nmol/L and complementary strand concentration was 400 nmol/L,and OTA had good linear relationship in the range of0.05-10.00 ng/mL.The cross-reactivity rate was lower than 1% with ochratoxin B,aflatoxin B1,zearalmone and deoxynivalenol,and the average recoveries of OTA in corn samples were 89.0%-93.8%.Conclusion This method is rapid,accurate and sensitive with low cross-reactivity rate.It will be used for the analysis and detection of OTA in samples.