Endpoint visual detection of three genetically modified rice events by loop-mediated isothermal amplification.
Endpoint visual detection of three genetically modified rice events by loop-mediated isothermal amplification.
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DOI：
10.3390/ijms131114421
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发表时间：
2012-11-07
影响因子：
5.6
通讯作者：
Xu J
中科院分区：
文献类型：
--
作者：
Chen X;Wang X;Jin N;Zhou Y;Huang S;Miao Q;Zhu Q;Xu J
转基因水稻KMD 1、TT 51 -1和KF 6是我国最著名的三个转Bt基因水稻品系。因此，需要一种快速、灵敏的转基因水稻风险评估方法。聚合酶链反应（PCR）是目前检测转基因生物最常用的方法，需要温度循环和相对复杂的程序。本文建立了一种直观、快速的环介导等温扩增（LAMP）方法，扩增了三个转基因水稻事件特异性连接序列。在63 °C的等温温度下，在60 min内通过两种指示剂（SYBR绿色或羟基萘酚蓝[HNB]）扩增并可视化靶DNA。选择不同种类的植物以保证检测的特异性，非目标样品的检测结果均为阴性，表明针对三个转基因水稻品种的引物组具有良好的特异性。LAMP的灵敏度为常规PCR的10 ~ 100倍，检测限为0.01%~ 0.005%GM。另外，与指示剂（SYBR绿色或HNB）偶联的LAMP测定促进分析。这些研究结果显示，快速检测方法适合作为一种简单的实地测试，以确定转基因作物的状况。
Genetically modified (GM) rice KMD1, TT51-1, and KF6 are three of the most well known transgenic Bt rice lines in China. A rapid and sensitive molecular assay for risk assessment of GM rice is needed. Polymerase chain reaction (PCR), currently the most common method for detecting genetically modified organisms, requires temperature cycling and relatively complex procedures. Here we developed a visual and rapid loop-mediated isothermal amplification (LAMP) method to amplify three GM rice event-specific junction sequences. Target DNA was amplified and visualized by two indicators (SYBR green or hydroxy naphthol blue [HNB]) within 60 min at an isothermal temperature of 63 °C. Different kinds of plants were selected to ensure the specificity of detection and the results of the non-target samples were negative, indicating that the primer sets for the three GM rice varieties had good levels of specificity. The sensitivity of LAMP, with detection limits at low concentration levels (0.01%–0.005% GM), was 10- to 100-fold greater than that of conventional PCR. Additionally, the LAMP assay coupled with an indicator (SYBR green or HNB) facilitated analysis. These findings revealed that the rapid detection method was suitable as a simple field-based test to determine the status of GM crops.
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影响因子：
56.9
作者：
SONG, WY;WANG, GL;RONALD, P
通讯作者：
RONALD, P
影响因子：
5.1
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作者：
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作者：
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