Development of a Simple and Cost-Effective Method Based on T7 Endonuclease Cleavage for Detection of Single Nucleotide Polymorphisms
Development of a Simple and Cost-Effective Method Based on T7 Endonuclease Cleavage for Detection of Single Nucleotide Polymorphisms
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开发一种基于 T7 核酸内切酶切割的简单且经济有效的方法来检测单核苷酸多态性
DOI：
10.1089/gtmb.2018.0181
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发表时间：
2018
影响因子：
1.4
通讯作者：
Wu Xingan
中科院分区：
文献类型：
--
作者：
Zhang Xiaoxiao;Yang Lina;Wang Fang;Liu Ziyu;Liu Rongrong;Ying Qikang;Fan Chao;Wu Xingan
目的：单核苷酸多态(SNP)可作为遗传标记，用于等位基因连锁疾病的风险评估，为临床诊断提供信息。大规模微阵列和下一代测序方法使全基因组SNP基因分型成为可能。然而，除了成本高昂外，这些技术还依赖于拥有专门的设备。因此，需要一种可以在资源有限的环境中实施的简单的基因分型方法。方法：我们建立了一种基于T7内切酶I酶切和酶联微粒免疫分析的SNP基因分型方法。利用该方法，我们对两个常见的SNP位点(rs11526468和rs12979860)进行了基因分型。验证了基因分型过程的质量。结果：虽然有70%的假阴性率，但没有发现假阳性反应。因此，多个平行重复反应可以抵消关闭时间失败检测的可能性。讨论：由于该方法使用了重复的试剂依赖程序，因此有可能集成到野外操作的便携式试剂盒中。
Aims: Single nucleotide polymorphisms (SNP) can be used as genetic markers and for risk assessment of allele-linked diseases, which may provide information for clinical diagnosis. Large-scale microarray and next-generation sequencing methods have made genome-wide SNP genotyping possible. However, in addition to their high cost, these techniques are dependent on having specialized equipment. Thus, there is a need for a simple genotyping method that can be implemented in a resource-limited environment. Methods: We developed a strategy for SNP genotyping based on T7 Endonuclease I cleavage and an enzyme-linked microparticle immune assay. Using this method, we genotyped two common SNP sites (rs11526468 and rs12979860). The quality of the genotyping process was validated. Results: Although a 70% false-negative rate was observed, no false-positive reactions were found. Therefore, multiple parallel repeat reactions can offset the possibility off time failed detection. Discussion: As the method employs a duplicate reagent-dependent procedure, it has the potential for integration into a portable kit for field operation.