Pan Jiafeng, Deng Fang, Zeng Lingwen, Liu Zhi, Chen Junhua
College of Bioscience and Biotechnology, Hunan Agricultural University, Changsha, 410128, China.
National-Regional Joint Engineering Research Center for Soil Pollution Control and Remediation in South China, Guangdong Key Laboratory of Integrated Agro-Environmental Pollution Control and Management, Institute of Eco-Environmental and Soil Sciences, Guangdong Academy of Sciences, Guangzhou, 510650, China.
Anal Bioanal Chem. 2022 Dec;414(29-30):8255-8261. doi: 10.1007/s00216-022-04354-3. Epub 2022 Sep 30.
Based on aptamer recognition and target-mediated competitive hybridization of hairpin probes, we developed a fluorescence sensor for kanamycin (KAN) detection. The aptamer and KAN binding will open hairpin H1 to release the trigger DNA fragment, which can initiate the competitive hybridization between hairpins H2 and H3. Then, exonuclease III (Exo III) can cleave H2 and H3 to produce numerous DNA3 and DNA4. Through the synergetic hybridization among DNA1, DNA2, DNA3, and DNA4, an active Mg-DNAzyme can be formed. The cleavage reaction toward FAM-BHQ-modified DNA2 will produce a high fluorescence signal for KAN assay. Through Exo III-guided cleavage and Mg-DNAzyme-based catalysis, the sensor exhibits high sensitivity, with a detection limit of 3.1 fM. This method is robust and has been applied to the detection of KAN in milk and water samples with good accuracy and reliability. Our developed fluorescence sensor exhibits the advantages of simple operation, high sensitivity, and good robustness, which are beneficial for KAN detection in food samples.
基于适体识别和发夹探针的靶标介导竞争性杂交，我们开发了一种用于检测卡那霉素（KAN）的荧光传感器。适体与KAN结合会打开发夹H1以释放触发DNA片段，该片段可引发发夹H2和H3之间的竞争性杂交。然后，核酸外切酶III（Exo III）可切割H2和H3以产生大量的DNA3和DNA4。通过DNA1、DNA2、DNA3和DNA4之间的协同杂交，可形成活性Mg-DNAzyme。对FAM-BHQ修饰的DNA2的切割反应将产生用于KAN检测的高荧光信号。通过Exo III引导的切割和基于Mg-DNAzyme的催化，该传感器具有高灵敏度，检测限为3.1 fM。该方法稳健，已应用于牛奶和水样中KAN的检测，具有良好的准确性和可靠性。我们开发的荧光传感器具有操作简单、灵敏度高和稳健性好的优点，有利于食品样品中KAN的检测。
