期刊
JOURNAL OF VIROLOGICAL METHODS
卷 214, 期 -, 页码 60-74出版社
ELSEVIER SCIENCE BV
DOI: 10.1016/j.jviromet.2015.01.003
关键词
Reverse-transcription PCR; Luminex direct hybridization assay; Self-avoiding molecular recognition system (SAMRS); Artificially expanded genetic information system (AEGIS); Dengue virus
资金
- DARPA under its ADEPT program [HR0011-11-2-0018]
- NIAID [R01AI098616]
- Nucleic Acids Licensing LLC
- Firebird Biomolecular Sciences LLC from the Office of the Secretary of Defense [W911NF-12-C-0059]
- Defense Threat Reduction Agency [HDTRA1-13-1-0004]
- NATIONAL INSTITUTE OF ALLERGY AND INFECTIOUS DISEASES [R01AI098616] Funding Source: NIH RePORTER
Mosquito-borne arboviruses are emerging world-wide as important human and animal pathogens. This makes assays for their accurate and rapid identification essential for public health, epidemiological, ecological studies. Over the past decade, many mono- and multiplexed assays targeting arboviruses nucleic acids have been reported. None has become established for the routine identification of multiple viruses in a single tube setting. With increasing multiplexing, the detection of viral RNAs is complicated by noise, false positives and negatives. In this study, an assay was developed that avoids these problems by combining two new kinds of nucleic acids emerging from the field of synthetic biology. The first is a self-avoiding molecular recognition system (SAMRS), which enables high levels of multiplexing. The second is an artificially expanded genetic information system (AEGIS), which enables clean PCR amplification in nested PCR formats. A conversion technology was used to place AEGIS component into amplicon, improving their efficiency of hybridization on Luminex beads. When Luminex liquid microarrays are exploited for downstream detection, this combination supports single-tube PCR amplification assays that can identify 22 mosquito-borne RNA viruses from the genera Flavivirus, Alphavirus, Orthobunyavirus. The assay differentiates between closely-related viruses, as dengue, West Nile, Japanese encephalitis, and the California serological group. The performance and the sensitivity of the assay were evaluated with dengue viruses and infected mosquitoes; as few as 6-10 dengue virions can be detected in a single mosquito. (C) 2015 Elsevier B.V. All rights reserved.
作者
我是这篇论文的作者
点击您的名字以认领此论文并将其添加到您的个人资料中。
推荐
暂无数据