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为学术实验室生产 SARS-CoV-2 定量 RT-PCR 检测试剂盒制定蓝图。

A blueprint for academic laboratories to produce SARS-CoV-2 quantitative RT-PCR test kits.

机构信息

School of Biological Sciences, Georgia Institute of Technology, Atlanta, Georgia, USA.

School of Chemistry and Biochemistry, Georgia Institute of Technology, Atlanta, Georgia, USA.

出版信息

J Biol Chem. 2020 Nov 13;295(46):15438-15453. doi: 10.1074/jbc.RA120.015434. Epub 2020 Sep 3.

Abstract

Widespread testing for the presence of the novel coronavirus severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in individuals remains vital for controlling the COVID-19 pandemic prior to the advent of an effective treatment. Challenges in testing can be traced to an initial shortage of supplies, expertise, and/or instrumentation necessary to detect the virus by quantitative RT-PCR (RT-qPCR), the most robust, sensitive, and specific assay currently available. Here we show that academic biochemistry and molecular biology laboratories equipped with appropriate expertise and infrastructure can replicate commercially available SARS-CoV-2 RT-qPCR test kits and backfill pipeline shortages. The Georgia Tech COVID-19 Test Kit Support Group, composed of faculty, staff, and trainees across the biotechnology quad at Georgia Institute of Technology, synthesized multiplexed primers and probes and formulated a master mix composed of enzymes and proteins produced in-house. Our in-house kit compares favorably with a commercial product used for diagnostic testing. We also developed an environmental testing protocol to readily monitor surfaces for the presence of SARS-CoV-2. Our blueprint should be readily reproducible by research teams at other institutions, and our protocols may be modified and adapted to enable SARS-CoV-2 detection in more resource-limited settings.

摘要

在出现有效的治疗方法之前,广泛检测个体中新出现的严重急性呼吸综合征冠状病毒 2(SARS-CoV-2)的存在对于控制 COVID-19 大流行仍然至关重要。检测方面的挑战可以追溯到最初缺乏定量 RT-PCR(RT-qPCR)检测病毒所需的用品、专业知识和/或仪器,RT-qPCR 是目前最强大、最敏感和最特异的检测方法。在这里,我们展示了配备适当专业知识和基础设施的学术生物化学和分子生物学实验室可以复制市售的 SARS-CoV-2 RT-qPCR 检测试剂盒并填补管道短缺。由佐治亚理工学院生物技术区的教职员工和学员组成的佐治亚理工学院 COVID-19 检测试剂盒支持小组合成了多重引物和探针,并配制了由内部生产的酶和蛋白质组成的主混合物。我们的内部试剂盒与用于诊断测试的商业产品相当。我们还开发了一种环境测试方案,可轻松监测表面是否存在 SARS-CoV-2。我们的蓝图应该可以由其他机构的研究团队轻松复制,我们的方案可以进行修改和调整,以在资源有限的环境中检测 SARS-CoV-2。

https://cdn.ncbi.nlm.nih.gov/pmc/blobs/4e57/7667971/d835cb2eb718/SB-JBCJ200618F001.jpg

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