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Preparation method of Norovirus nucleic acid standard sample

A standard sample and viral nucleic acid technology, applied in the field of viral nucleic acid standard sample preparation, can solve the problems of difficulty in maintaining stability and uniformity, no animal model, time-consuming and labor-intensive, saving time and effort in the preparation process and ensuring quality control. , the effect of high stability

Inactive Publication Date: 2012-08-15
DALIAN NATIONALITIES UNIVERSITY
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  • Description
  • Claims
  • Application Information

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Problems solved by technology

The virus is highly contagious and can easily cause outbreaks. If the treatment is not timely or the treatment method is incorrect, the mild cases will affect the growth and development of the human body, and the severe cases may lead to dehydration and death.
In recent years, the number of cases caused by this virus has been on the rise, and the diagnosis and control of this disease is particularly important. However, norovirus cannot be reproduced in vitro, there is no animal model, and it cannot be isolated and tested by tissue culture or animal experiments. Research on the etiology, pathogenic mechanism, and diagnostic methods of the disease and formulate prevention and control measures
[0004] Because the virus not only has the above-mentioned particularity, but also the virus is an RNA virus, and its RNA is extremely easy to be degraded. It is not only time-consuming and labor-intensive to extract the RNA virus in each laboratory, but it is also difficult to maintain its stability and uniformity, and it is difficult to ensure that the laboratory quality control of

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  • Preparation method of Norovirus nucleic acid standard sample
  • Preparation method of Norovirus nucleic acid standard sample
  • Preparation method of Norovirus nucleic acid standard sample

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Embodiment Construction

[0034] a. To extract norovirus RNA, add 1000 μl of Trizol lysate and 200 μl of human norovirus-infected feces (purchased from the Chinese Center for Disease Control and Prevention) to a centrifuge tube, and set a positive control and a negative control at the same time. Add 200 μl of chloroform, shake and mix well, and centrifuge at 10 000 g for 15 minutes; absorb 500 μl of the supernatant and add it to 500 μl isopropanol, invert and mix repeatedly, and centrifuge at 10 000 g for 15 minutes; discard the supernatant, place it upside down on absorbent paper, and To dry the liquid, add 1,000 μl of 75% alcohol, wash it upside down, and centrifuge at 10,000 g for 10 minutes; discard the supernatant gently, put it upside down on absorbent paper, and dry the liquid; centrifuge at 4,000 g for 10 sec, and blot dry with a micro-sampler Dry the liquid at the bottom of the tube at room temperature for 3 minutes; add 11.5 μl of DEPC water, mix gently to dissolve the RNA in the tube, centrif...

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Abstract

The invention discloses a preparation method of a Norovirus nucleic acid standard sample, which is characterized by including the steps of extracting RNA (ribonucleic acid) of Norovirus; performing RT-PCR (reverse transcription-polymerase chain reaction); purifying a target gene; connecting and transforming target segments; recycling plasmids; and performing in-vitro transcription. The preparation process is timesaving and laborsaving, and the prepared Norovirus nucleic acid standard sample is highly stable and uniform and can be used for Norovirus detection and research, pharmaceutical research, application research and the like so as to achieve comparison of different lab results and guarantee quality control in labs.

Description

technical field [0001] The invention relates to a preparation method of a viral nucleic acid standard sample, in particular to a preparation method of a norovirus nucleic acid standard sample which saves time and effort, has high stability and good uniformity. Background technique [0002] Norovirus (Norovirus, NV), a small round-shaped virus belonging to the Caliciviridae family and the genus Walkervirus, has a 2O-hedron symmetry, no envelope, and lacks significant morphological features under the electron microscope. It is different from animal caliciviruses. There are no obvious cup-embedded depressions or surface holes, and the floating density in cesium chloride (CsCl) is 1.38~1.41 g / cm. Strong resistance to various physical and chemical factors, resistant to ether, acid and heat, and cannot be completely inactivated at 100°C for 30 minutes. [0003] Norovirus is a common cause of intestinal infectious diseases caused by viruses, and it has become the most important ca...

Claims

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Application Information

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IPC IPC(8): C12N15/63C12N15/40
Inventor 杜雄伟李叶
Owner DALIAN NATIONALITIES UNIVERSITY