溶藻弧菌适配子亲和力测定条件的优化及其特异性研究

Optimization of Affinity Determination Condition of Aptamer for Vibrio alginolyticus and Specificity Analysis

[目的]优化溶藻弧菌适配子亲和力的测定条件,并验证适配子的亲和特异性。[方法]采用地高辛-过氧化物酶显色系统,通过正交试验确定亲和力测定的适宜条件,并在该条件下进行特异性验证。[结果]优化后亲和力的测定条件为:菌浓度3.0×108个/ml,结合时间40 min,结合温度28℃;经验证,该适配子对溶藻弧菌具有较高的亲和特异性。[结论]获得了溶藻弧菌适配子亲和力测定的优化条件,并证明了该适配子与溶藻弧菌具有较好的亲和特异性,为溶藻弧菌的SELEX筛选及其快速检测提供了参考。

[Objective] The aim was to optimize the condition for the affinity detection of an aptamer against Vibrio alginolyticus and to validate the aptamer′s specific affinity to the microorganism.[Method] Based on the digoxigenin-peroxidase color system,the optimal condition was obtained by the orthogonal experiment and the specific affinity of the aptamer was validated under the optimized conditions.[Result] The optimum condition for the affinity detection of the aptamer was obtained as follows：bacteria concentration 3.0×10^8 cell/ml,binding time 40 min,and binding temperature 28 ℃;the aptamer was validated to have the specific affinity to the microorganism Vibrio alginolyticus under the optimum condition.[Conclusion] The optimum condition for the affinity detection of the aptamer against Vibrio alginolyticus was obtained,and the aptamer was proved to have good specific affinity to the microorganism,which would be a valuable reference for the SELEX of aptamers against Vibrio alginolyticus and the quick detection of the microorganism.