Systematic truncating of aptamers to create high-performance graphene oxide (GO)-based aptasensors for the multiplex detection of mycotoxins.
Systematic truncating of aptamers to create high-performance graphene oxide (GO)-based aptasensors for the multiplex detection of mycotoxins.
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DOI：
10.1039/c9an00624a
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发表时间：
2019-06
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通讯作者：
Xing-Xia Wang;Xiaoyi Gao;Jiale He;Xiaochen Hu;Yunchao Li;Xiaohong Li;L. Fan;Hua-Zhong Yu
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作者：
Xing-Xia Wang;Xiaoyi Gao;Jiale He;Xiaochen Hu;Yunchao Li;Xiaohong Li;L. Fan;Hua-Zhong Yu
基于氧化石墨烯（GO）的适体传感器是目前最流行的传感平台之一，用于简单快速地检测各种目标。不幸的是，具有长适体链的基于GO的适体传感器通常显示出不令人满意的性能，这是由于在靶结合时不显著的结构转化。我们在此报告利用适体截短策略来应对这样的挑战。以预先筛选的抗黄曲霉毒素B1（AFB 1）适体（P-AFB 1 -50）为试验体系，通过圆二色谱（CD）和结合亲和力分析，依次去除适体中的外源核苷酸。特别地，对于每个截短的适体，确定在不存在和存在靶标的情况下GO片层和截短的适体（用荧光团标记）之间的猝灭常数的比率，以评估最佳序列。结果，证实包含40个核苷酸的截短适体在与GO片缀合后显示出最高的FL输出和最佳的检测限。更重要的是，我们证明了这种截断策略是通用的，即，它可以很容易地扩展到其它适体系统（例如抗赭曲霉毒素A（OTA）适体，P-OTA-61），用于外来核苷酸鉴定。令人印象深刻的是，两种最佳截短适体可以在GO片上一起工作，以实现两种不同真菌毒素的同时检测（即，AFB 1和OTA）。从本质上讲，这项研究为设计和测试基于适体/GO的传感平台开辟了一条新的途径，用于快速，低成本和多重定量分析目标。
Graphene oxide (GO)-based aptasensors are currently one of the most popular sensing platforms for the simple and rapid detection of various targets. Unfortunately, the GO-based aptasensors with long aptamer strands typically show unsatisfactory performance resulting from insignificant structural transformations upon target binding. We report herein the utilization of an aptamer-truncating strategy to combat such a challenge. Taking a pre-selected anti-aflatoxin B1 (AFB1) aptamer (P-AFB1-50) as a trial system, we sequentially remove the extraneous nucleotides within the aptamer by means of circular dichroism (CD) spectroscopy and binding affinity analysis. Particularly, the ratio of the quenching constants between the GO sheets and the truncated aptamers (labelled with fluorophores) in the absence and presence of the target was determined for each of the truncated aptamers to evaluate the optimal sequence. As a result, the truncated aptamer comprising 40 nucleotides was confirmed to show the highest FL output and the best detection limit upon conjugation with GO sheets. More importantly, we demonstrated that this truncating strategy is versatile, i.e., it can be easily extended to other aptamer systems (anti-ochratoxin A (OTA) aptamer, P-OTA-61, as an example) for extraneous nucleotide identification. Impressively, the two optimal truncated aptamers can work together on GO sheets to achieve a simultaneous detection of two different mycotoxins (i.e., AFB1 and OTA) in one single test. Essentially, this research opens a new avenue for the design and testing of aptamer-/GO-based-sensing platforms for rapid, low-cost and multiplex quantification of analytical targets of interest.