Novel coronavirus SARS-CoV-2 detection proteome and application thereof

A coronavirus, sars-cov-2 technology, applied in the detection proteome and application field of the new coronavirus SARS-CoV-2, can solve the problems of reduced positive rate, high requirements, long detection time, etc., to increase specificity Effect

Pending Publication Date: 2021-04-16
SHANXI ACAD OF ADVANCED RES & INNOVATION
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] Therefore, the technical problem to be solved in the present invention is to solve the problem that the positive rate of the existing detection method is reduced due to the long detection time and the high requirements, thereby provid...

Method used

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  • Novel coronavirus SARS-CoV-2 detection proteome and application thereof
  • Novel coronavirus SARS-CoV-2 detection proteome and application thereof
  • Novel coronavirus SARS-CoV-2 detection proteome and application thereof

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0097] The present invention provides a kind of detection method of novel coronavirus SARS-CoV-2, and described detection method is enzyme-linked immunosorbent assay method, comprises the following steps:

[0098] (1) Cultivate the bacterial strain containing the plasmid pET-30a-nCoV-N in LB medium containing kanamycin at a culture temperature of 37° C. and shake at a speed of 180 r / min;

[0099] When the OD value of the optical density is 0.6, add the inducer isopropylthiogalactopyranoside to the LB medium, and induce overnight at 18°C;

[0100] Finally, the purified N protein was obtained after sonication, His-HP column elution using 1mol / L NaCl as the eluent and gel chromatography;

[0101] (2) Add 40 μg of pEZT-sp-nCoV RBD plasmid to DMEM medium and mix to make the final volume 1mL to obtain solution 1, then add 120ug polyethyleneimine to DMEM medium and mix to make the final volume to 1mL to obtain solution 2 ;

[0102] Then solution 2 was dropped into solution 1, and l...

Embodiment 2

[0111] The present invention provides a kind of detection method of novel coronavirus SARS-CoV-2, and described detection method is enzyme-linked immunosorbent assay method, comprises the following steps:

[0112](1) Cultivate the strain containing the plasmid pET-30a-nCoV-N in LB medium containing kanamycin at a culture temperature of 38° C. and shake at a speed of 200 r / min;

[0113] When the OD value of the optical density is 0.65, add the inducer isopropylthiogalactopyranoside to the LB medium, and induce overnight at 20°C;

[0114] Finally, the purified N protein was obtained after sonication, His-HP column elution using 1mol / L NaCl as the eluent and gel chromatography;

[0115] (2) Add 40 μg of pEZT-sp-nCoV RBD plasmid to DMEM medium and mix to make the final volume 1mL to obtain solution 1, then add 120ug polyethyleneimine to DMEM medium and mix to make the final volume to 1mL to obtain solution 2 ;

[0116] Then drop solution 2 into solution 1, and place it at room t...

Embodiment 3

[0125] The present invention provides a kind of detection method of novel coronavirus SARS-CoV-2, and described detection method is enzyme-linked immunosorbent assay method, comprises the following steps:

[0126] (1) Cultivate the bacterial strain containing the plasmid pET-30a-nCoV-N in LB medium containing kanamycin, the cultivation temperature is 36°C, and the culture is shaken at a speed of 150r / min;

[0127] When the OD value of the optical density is 0.55, add the inducer isopropylthiogalactopyranoside to the LB medium, and induce overnight at 15°C;

[0128] Finally, the purified N protein was obtained after sonication, His-HP column elution using 1mol / L NaCl as the eluent and gel chromatography;

[0129] (2) Add 40 μg of pEZT-sp-nCoV RBD plasmid to DMEM medium and mix to make the final volume 1mL to obtain solution 1, then add 120ug polyethyleneimine to DMEM medium and mix to make the final volume to 1mL to obtain solution 2 ;

[0130] Then drop solution 2 into solut...

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Abstract

The invention discloses a novel coronavirus SARS-CoV-2 detection proteome and an application thereof. A detection method for detecting novel coronavirus SARS-CoV-2 is an enzyme-linked immunosorbent assay method. The N protein is optimized, and the receptor binding region (RBD 331aa-532aa) of the S protein is selected for expression through structural analysis aiming at the S gene of the novel coronavirus, so that the detection specificity can be further improved; an enzyme-linked immunosorbent assay method is used, and whether serum to be detected is positive or not can be efficiently and accurately determined by optimizing conditions such as coating concentration and time, primary antibody and secondary antibody incubation time and developing time.

Description

technical field [0001] The invention relates to the field of viral nucleic acid detection, in particular to the detection proteome and application of the novel coronavirus SARS-CoV-2. Background technique [0002] The recent outbreak of pneumonia caused by a novel coronavirus (SARS-CoV-2) poses a huge threat to global public health. Therefore, rapid and accurate identification of pathogenic viruses plays a crucial role in selecting appropriate treatments, saving lives and preventing epidemics. It is important to establish rapid standard diagnostic tests to detect infectious disease (COVID-19) to prevent subsequent secondary transmission. [0003] ELISA is the abbreviation of Enzyme-Linked Immunosorbnent Assay. It is an immunoenzyme technique developed after immunofluorescence and radioimmunoassay techniques. Since the early 1970s, this technology has been reported by VAN MEEMEM, SCHUARS.ENGVALL and PERLMARN, etc., and has now attracted widespread attention. ELISA method ...

Claims

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Application Information

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IPC IPC(8): G01N33/569G01N33/543
Inventor 仝舟高山杨勃张德梅张智超闫红
Owner SHANXI ACAD OF ADVANCED RES & INNOVATION
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