Multiplex Visualized Closed-Tube PCR with Hamming Distance 2 Code for 15 HPV Subtype Typing

化学 放大器 打字 一致性 聚合酶链反应 多路复用 金标准(测试) 多重聚合酶链反应 分子生物学 计算生物学 生物 遗传学 基因 内科学 医学 生物化学
作者
Jixue Weng,Nan Sheng,Runyuan Wang,Shuo Liang,Chen Wang,Xue Bai,Guohua Zhou,Bingjie Zou,Qinxin Song
出处
期刊:Analytical Chemistry [American Chemical Society]
卷期号:93 (13): 5529-5536 被引量:13
标识
DOI:10.1021/acs.analchem.1c00035
摘要

Cervical cancer is the fourth leading cause of death in women, especially in developing countries. Specific and economic methodologies for HPV typing are crucial in cancer diagnosis and further disease control. However, routine assays based on real-time polymerase chain reaction (qPCR) or DNA-chip hybridization are either incapable of offering detailed subtype information or involve tedious open-tube operations with the risk of cross-contamination from PCR amplicons. Herein, we proposed a multiplex visualized closed-tube PCR (Multi-Vision) for HPV typing. Using gold nanoparticle probes (AuNPs) as a color change indicator combined with a Hamming distance 2 coding scheme, 13 high-risk HPVs and two subtypes associated with high-incidence benign lesions were successfully typed by performing six closed-tube PCRs. The assay demonstrates high specificity with no cross-reaction among different subtypes under several artificial sample concentrations (from 100 to 103 copies per reaction) and enables highly sensitive detection of as low as 0.5 copies/μL. Further, 105 clinical samples were successfully analyzed using our method with a high concordance rate of 99.05% (104/105) compared to a HPV typing kit. The inconsistent sample was confirmed by sequencing to be consistent with the typing results determined by our method, indicating that Multi-Vision could be a useful tool for HPV detection, especially in resource-limited regions.
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