Signal amplified strategy based on target-induced strand release coupling cleavage of nicking endonuclease for the ultrasensitive detection of ochratoxin A
Signal amplified strategy based on target-induced strand release coupling cleavage of nicking endonuclease for the ultrasensitive detection of ochratoxin A
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基于切口核酸内切酶靶标诱导链释放耦合裂解的信号放大策略用于赭曲霉毒素 A 的超灵敏检测
DOI：
10.1016/j.bios.2012.07.005
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发表时间：
2013-01-15
影响因子：
12.6
通讯作者：
Luo, Xiliang
中科院分区：
文献类型：
--
作者：
Hun, Xu;Liu, Fang;Luo, Xiliang
在这项工作中，一个新的信号放大策略的基础上的目标诱导链释放耦合切割的切口核酸内切酶的超灵敏检测赭曲霉毒素A（OTA）。将OTA适体（DNA 1）和OTA适体互补物（DNA 2）固定在磁珠（MB）上。在OTA存在下，DNA 2从MB解离并释放。然后，释放的DNA 2与DNA 3杂交，DNA 3连接在扩增模板的5'末端，并且可以在Phi 29 DNA聚合酶存在下沿着模板延伸。形成的双链DNA被切口核酸内切酶Nb.BbvCI切割，并产生短的单链DNA。在Phi 29 DNA聚合酶的作用下，DNA链断裂后产生一个新的位点，延伸和断裂的过程是循环的。因此，产生了一定量的短单链DNA。利用DNA和ABEI标记的羧基二氧化硅纳米颗粒化学发光探针，可以灵敏地检测短单链DNA。CL强度（Δ I）与OTA浓度在1.0 x 10（-12）至5.0 x 10（-8）g mL（-1）范围内呈线性关系。方法的检出限为3.0 × 10（-13）g mL（-1），在1.0 × 10（-10）g mL（-1）浓度下，RSD为3.4%（n=7）。该方法已应用于天然污染小麦样品中OTA的检测。由于其简单，灵敏度和不需要特异性识别的适配体切割，这种CL生物测定提供了一个有前途的方法检测OTA和其他生物分子。(C)2012爱思唯尔有限公司版权所有。
In this work, a new signal amplified strategy based on target-induced strand release coupling cleavage of nicking endonuclease for the ultrasensitive detection of ochratoxin A (OTA) is reported. OTA aptamer (DNA1) and OTA aptamer complementary (DNA2) were immobilized onto a magnetic bead (MB). In the presence of OTA, DNA2 was dissociated and released from the MB. The released DNA2 then hybridized with DNA3, which was linked at the 5' terminus of the amplification template and can extend along the template in the presence of Phi 29 DNA polymerase. The formed double-stranded DNA was cleaved by nicking endonuclease Nb.BbvCI and produced a short single-stranded DNA. The cleaved DNA strand generated a new site by Phi 29 DNA polymerase and the process of extension and cleavage was cyclical. Thus, a amount of the short single-stranded DNA were produced. Using DNA and ABEI labeled carboxylic silica nanoparticles chemiluminescence (CL) probe, the short single-stranded DNA could be sensitively detected. The CL intensity (Delta I) versus the concentration of OTA was linear in the range from 1.0 x 10(-12) to 5.0 x 10(-8) g mL(-1). The detection limit was 3.0 x 10(-13) g mL(-1), and the RSD was 3.4% at 1.0 x 10(-10) g mL(-1) (n=7). The developed method has been applied to detect OTA in naturally contaminated wheat samples. Due to its simplicity, sensitivity and no need of specific recognition of aptamer for cleavage, this CL bioassay offers a promising approach for the detection of OTA and other biomolecules. (C) 2012 Elsevier B.V. All rights reserved.