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利用有机磷农药与分子信标竞争结合核酸适体的原理,建立了核酸适体活性的荧光鉴定方法,对前期筛选的15条核酸适体的活性进行了鉴定。结果表明,分子信标设计合理,甲醇和4种有机磷农药对分子信标发夹结构的稳定性影响较小,室温孵育时间为60 min时体系荧光强度趋于稳定,当体系中适体与分子信标的添加比例为1∶5时,荧光强度的净增量达到最大值。适体鉴定结果表明适体ss4,ss6,ss12,ss16,ss24和ss27对甲基对硫磷表现出了一定的活性,但亲和活性均较弱;适体ss5,ss12,ss13和ss24对马拉硫磷表现出了一定的活性,其中适体ss5活性最好,荧光抑制率达到了64.97%;适体ss1,ss4,ss6,ss7,ss10,ss16,ss17,ss23,ss27和ss29对辛硫磷表现出了一定的活性,其中适体ss27活性最好,荧光抑制率达到了49.42%;适体ss13,ss26和ss27对乐果表现出了一定的活性,其中ss26活性最好,荧光抑制率达到了44.35%。
Based on the principle that organophosphorus pesticides and molecular beacons competed for the binding of aptamers, a fluorescent assay was established for the activity of aptamers. The activity of 15 aptamers selected in the previous study was identified. The results showed that the design of molecular beacons was reasonable and methanol and four organophosphorus pesticides had little effect on the stability of the molecular beacon hairpin. The fluorescence intensity tended to be stable when the incubation time was 60 min at room temperature. When the molecular beacon is added in a ratio of 1: 5, the net increase in fluorescence intensity reaches a maximum value. The results of aptamer identification showed that aptamers ss4, ss6, ss12, ss16, ss24 and ss27 showed some activities on methyl parathion, but their affinity activities were weaker. Aptamers ss5, ss12, ss13 and ss24 The activity of aptamer ss5 was the best, and the fluorescence inhibition rate reached 64.97%. The aptamers ss1, ss4, ss6, ss7, ss10, ss16, ss17, ss23, ss27, Phosphorus showed a certain activity, of which aptamer ss27 activity was the best, the fluorescence inhibition rate reached 49.42%; aptamers ss13, ss26 and ss27 on dimethoate showed some activity, of which ss26 activity is best, fluorescence inhibition rate Reached 44.35%.