In vitro selection and characterization of DNA aptamers recognizing chloramphenicol
In vitro selection and characterization of DNA aptamers recognizing chloramphenicol
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DOI：
10.1016/j.jbiotec.2011.06.043
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发表时间：
2011-10-10
影响因子：
4.1
通讯作者：
Robbens, Johan
中科院分区：
文献类型：
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作者：
Mehta, Jaytry;Van Dorst, Bieke;Robbens, Johan
氯霉素（Cam）虽然是一种有效的抗生素，但由于一些致命的副作用而失去了人们的青睐。因此，迫切需要快速、灵敏的方法来检测食品、饲料和环境中的残留物。我们通过进行体外选择，设计了能够将 Cam 识别为目标的 DNA 适体。适体是核酸识别元件，对其靶点具有高度特异性和敏感性，并且可以以对动物友好的方式合成生产，使其成为抗体的道德创新替代品。本研究中分离的适配体彼此之间没有序列同源性或结构相似性，这表明在所使用的选择条件下，不同的DNA序列可以实现特定的Cam识别。分析序列的结合亲和力，证明解离常数 (K-d) 在极低的微摩尔范围内，低于之前报道的 Cam 特异性 RNA 适体的解离常数 (K-d)。两个最好的适体具有富含 G (> 35%) 的核苷酸区域，这是它们区别于其他适体的属性，并且显然是它们高选择性和亲和力的原因（K-d 分别类似于 0.8 和 1 μM）。这些适体开辟了通过基于适体的生物传感器轻松检测 Cam 的可能性。 (C) 2011 Elsevier B.V. 保留所有权利。
Chloramphenicol (Cam), although an effective antibiotic, has lost favour due to some fatal side effects. Thus there is an urgent need for rapid and sensitive methods to detect residues in food, feed and environment. We engineered DNA aptamers that recognize Cam as their target, by conducting in vitro selections. Aptamers are nucleic acid recognition elements that are highly specific and sensitive towards their targets and can be synthetically produced in an animal-friendly manner, making them ethical innovative alternatives to antibodies. None of the isolated aptamers in this study shared sequence homology or structural similarities with each other, indicating that specific Cam recognition could be achieved by various DNA sequences under the selection conditions used. Analyzing the binding affinities of the sequences, demonstrated that dissociation constants (K-d) in the extremely low micromolar range, which were lower than those previously reported for Cam-specific RNA aptamers, were achieved. The two best aptamers had G rich (> 35%) nucleotide regions, an attribute distinguishing them from the rest and apparently responsible for their high selectivity and affinity (K-d similar to 0.8 and 1 mu M respectively). These aptamers open up possibilities to allow easy detection of Cam via aptamer-based biosensors. (C) 2011 Elsevier B.V. All rights reserved.