基于多聚腺嘌呤的二嵌段寡核苷酸探针用于卡那霉素可视化检测  

Visual Detection of Kanamycin by Poly-Adenine-based Diblock Oligonucleotide Probe

作　　者：席玥 王欣雨 王汇元 颜娟[1,2,3] XI Yue;WANG Xin-Yu;WANG Hui-Yuan;YAN Juan(College of Food Science and Technology,Shanghai Ocean University,Shanghai 201306,China;Shanghai Engineering Research Center of Aquatic-Product Processing&Preservation,Shanghai 201306,China;Laboratory of Quality and Safety Risk Assessment for Aquatic Products on Storage and Preservation(Shanghai),Ministry of Agriculture,Shanghai 201306,China)

机构地区：[1]上海海洋大学食品学院,上海201306 [2]上海水产品加工及贮藏工程技术研究中心,上海201306 [3]农业部水产品贮藏保鲜质量安全风险评估实验室(上海),上海201306

出　　处：《分析化学》2024年第8期1114-1122,共9页Chinese Journal of Analytical Chemistry

基　　金：上海市自然科学基金(No.20ZR1424100);国家市场监管重点实验室(生物分析计量溯源领域)开放课题(No.KLSMR2022-01)资助。

摘　　要：单链DNA(Single-stranded DNA,ssDNA)在未修饰的裸金纳米颗粒(AuNPs)上的吸附已成为基于核酸分子的比色分析的常用机制。本研究设计了一段包含多聚腺嘌呤(polyA20)尾部序列的二嵌段寡核苷酸探针(dP),构建了一种免标记的AuNPs比色生物传感器用于检测卡那霉素(Kanamycin,Kana)。在卡那霉素存在的情况下,磁珠@双链DNA(MBs@dsDNA)复合物上的卡那霉素适配体序列(Kana-Apt)与其特异性结合,导致dP从磁珠表面释放。由于polyA20与AuNPs之间有强吸附亲和力,dP被吸附在AuNPs表面,从而在高盐环境中保护AuNPs不发生团聚。与具有相同碱基数量的随机序列(Random sequence,RS)相比,利用含有polyA20尾部序列的AuNPs比色策略对卡那霉素的检测灵敏度提高了2个数量级。此比色传感器对卡那霉素的线性检测范围为0.01~1 nmol/L,检出限(LOD,S/N=3)为0.01 nmol/L。本方法无需大型仪器及其它信号放大策略,具有较高的灵敏度和特异性,适用于现场大批量样品的快速、可视化和实时检测,为其它抗生素以及影响食品安全的化学污染物的可视化检测提供了新思路。Adsorption of single-stranded DNA(ssDNA)onto unmodified gold nanoparticles(AuNPs)has been utilized as a common strategy for colorimetric assays,enabling facile detection of target analytes.In this study,a label-free,sensitive AuNP-based colorimetric biosensor was developed for visual detection of kanamycin(Kana)using a diblock oligonucleotide probe(dP)that included a poly-adenine(polyA20)tail.In the presence of Kana,the aptamer on the MBs@double-stranded DNA(dsDNA)complex specifically bound to Kana,leading to dissociation of dPs from complementary aptamer strands.The dP sequences were anchored to the surface of AuNPs due to the strong affinity between polyA20 tail and AuNPs,thus stabilizing the colloidal AuNPs against aggregation when high concentration level salt(NaCl)was introduced.Compared with random sequence(RS)with the same number of bases,AuNPs colorimetric strategy with polyA20 tail sequence significantly improved the detection sensitivity of Kana by two orders of magnitude.The results showed that the colorimetric sensor had good colorimetric performance when the concentration of Kana ranged from 0.01 to 1 nmol/L,and the detection limit(S/N=3)was 0.01 nmol/L.This sensing system did not require large instruments and other signal amplification technique,and was suitable for visualization and real-time detection of large quantities of samples with high sensitivity and specificity.This study provided a good idea for visual detection of other antibiotic drugs and chemical pollutants affecting food safety.

关 键 词：卡那霉素 金纳米颗粒 核酸适配体 生物传感器 食品安全 

分 类 号：O657.3[理学—分析化学] TS207.5[理学—化学]