Rapid and sensitive method for detection of Marburg virus RT-LAMP

A technology of RT-LAMP and Marburg virus, which is applied to the determination/inspection of microorganisms, biochemical equipment and methods, etc., can solve the problems of long time required, low sensitivity, and large equipment, so as to reduce time and complexity, Improve the detection ability, fast and accurate detection effect

CN105936943AActive Publication Date: 2016-09-14SHANGHAI ENTRY EXIT INSPECTION & QUARANTINE BUREAU OF P R C
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Patent Information

Authority / Receiving Office
CN · China
Patent Type
Applications(China)
Current Assignee / Owner
Publication Date
2016-09-14

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Abstract

The invention relates to a rapid and sensitive method for detection of Marburg virus RT-LAMP. The detection method of the invention employs specific primers design, loop mediated isothermal amplification (LAMP) technique, and fluorescent visual reagent for simple and fast detection of amplification products. The method includes: specimen RNA extraction, RT-LAMP amplification, and interpretation of results. The method can conduct fast and sensitive detection of Marburg virus, uses simple equipment and operations and has less testing time. The invention can greatly improve the detection efficiency of the inspection and quarantine officers in the front line of import and export ports, and can reduce the workload, and solve the positive leak detection in the traditional detection method by the maximum, so as to maximally prevent the introduction of Marburg virus.
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Description

technical field

[0001] The invention relates to the technical field of in vitro diagnostic reagents, in particular to a rapid and sensitive Marburg virus RT-LAMP detection method. Background technique

[0002] With global economic integration and trade liberalization, foreign medical vectors can quickly spread infectious diseases from one country or region to the world through advanced means of transportation and international trade, resulting in international spread. In recent years, new viral infectious diseases have frequently broken out in countries all over the world, posing a serious threat to human health and causing huge economic losses to society. For example, the Severe Acute Respiratory Syndrome (SARS) epidemic in some countries and regions in 2003, the avian influenza epidemic in my country and neighboring countries in 2004, the global H1N1 influenza epidemic in 2009, and the Ebola epidemic in Africa in 2014 have given society It has had a major impact on stabili...

Examples

Embodiment 1

[0036] Example 1 Specificity and sensitivity of Marburg virus RT-LAMP detection method

[0037] 1. Experimental steps

[0038] 1. In vitro synthesis of Marburg virus and Ebola virus RNA

[0039] Using bioinformatics methods to analyze the genome sequence (Ang1379c) of the published Marburg virus Angola strain, the artificially synthesized fragment (about 600 bp) containing the specific detection Marburg virus VP35 was cloned on the PCR cloning vector (TOPO TA cloning kit), and used Plasmid DNA extraction kit to purify the plasmid DNA, sequence to confirm that the insert is correct, and then perform in vitro transcription: First, digest the plasmid with Not I to linearize it to exclude downstream RNA products, then use MAXIscript T7Kit for in vitro transcription to generate target RNA, and then use TurboDNase Perform DNase treatment to remove untranscribed plasmid DNA templates, and finally use ethanol precipitation to purify the RNA product, which is the positive control RNA ...