Toehold链置换辅助智能手机比色法特异性检测废水中新冠病毒靶标RNA  

Toehold-mediated strand displacement reaction assisted smart phone colorimetry for specific detection of SARS-CoV-2 RNA in wastewater

作　　者：张琳 张颖[1] 宋九华[1] 韩耀霞[1] 杨孝容[1] 雷欣梦 黄莉翔 徐君茹 何思仪 邱小凤 史铠 ZHANG Lin;ZHANG Ying;SONG Jiuhua;HAN Yaoxia;YANG Xiaorong;LEI Xinmeng;HUANG Lixiang;XU Junru;HE Siyi;QIU Xiaofeng;SHI Kai(College of New Energy Materials and Chemistry,Leshan Normal University,Leshan 614000,China)

机构地区：[1]乐山师范学院新能源材料与化学学院,乐山614000

出　　处：《分析试验室》2024年第1期36-41,共6页Chinese Journal of Analysis Laboratory

基　　金：国家自然科学基金(22204066);四川省科技计划(2022JDRC0126);大学生创新创业训练计划项目(202210649023)资助。

摘　　要：基于Toehold介导的链置换反应(TSDR)与DNA酶,构建了一个强特异性及低成本检测废水中SARS-CoV-2靶标RNA单碱基突变的比色生物传感器。SARS-CoV-2靶标RNA与双链DNA(dsDNA)引发TSDR,释放出完整的G-四链体序列并与氯化血红素(Hemin)结合,形成G-四链体/Hemin DNA酶,进而催化双氧水氧化3,3’,5,5’-四甲基联苯胺,使溶液呈蓝色。当SARS-CoV-2靶标RNA存在单碱基突变时,TSDR被抑制,导致裸眼观察到的溶液颜色与未突变异的靶标相比更浅。同时采用智能手机摄像头捕捉溶液的RGB值变化(ΔRGB),对裸眼观察结果的准确性进行验证。在最优条件下,构建的比色生物传感器能够特异性识别单个碱基突变的SARS-CoV-2靶标RNA,具有用于废水中SARS-CoV-2变异毒株检测的潜力。A colorimetric biosensor based on toehold-mediated strand displacement reaction(TSDR) and DNAzyme with high specificity and low cost was constructed to detect single base mutation of SARS-CoV-2 target RNA in wastewater.The SARS-CoV-2 target RNA could bind to the toehold region of the DNA1/DNA2 duplex,and displace the DNA1 through TSDR,followed by liberating the pre-blocked G-quadruplex sequence,which could associate with Hemin to form G-quadruplex/Hemin DNAzyme.Subsequently,the G-quadruplex/Hemin DNAzyme could catalyze H_2O_(2) to oxide the TMB efficiently,and change the colorless solution to be blue.However,the TSDR procedure could be inhibited when a single base was mutational in the target RNA.This resulted in a lighter color of the solution.Meanwhile,a smartphone was employed to capture the change of RGB value of the solution to verify the accuracy of the measurement results.Under the optimal conditions,the developed biosensor could specifically identify SARS-CoV-2 target RNA with a single base mutation,and has an application prospect for the detection of SARS-CoV-2 variants in wastewater.

关 键 词：新冠病毒靶标RNA Toehold链置换 智能手机 废水 

分 类 号：O654.2[理学—分析化学]