Novel coronavirus N-S dominant epitope fusion protein, preparation method, application, expression protein, microorganism, application and kit

A dominant epitope, fusion protein technology, applied in the field of genetic engineering, can solve the problems of difficult nucleic acid sampling, long detection time, missed detection of infected persons, etc., to achieve excellent sensitivity and specificity, improved detection sensitivity, and reduced missed detection rate. Effect

Active Publication Date: 2020-06-02
HEBEI GENSURE BIOTECH LIC
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, due to the limitation of nucleic acid detection by unfavorable factors such as difficult sampling, complicated operation, and long detection time, some infected persons will be missed (for example, many cases of nucleosides have appeared

Method used

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  • Novel coronavirus N-S dominant epitope fusion protein, preparation method, application, expression protein, microorganism, application and kit
  • Novel coronavirus N-S dominant epitope fusion protein, preparation method, application, expression protein, microorganism, application and kit

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0026] Example 1 N protein dominant epitope segment antigen screening and determination

[0027] According to the full-length amino acid sequence of the new coronavirus N protein published in Genebank, the amino acid sequence of the N protein was analyzed with the B-cell epitope analysis and signal peptide analysis functions of BIOSUN software, and the antigenicity was selected according to the strength of hydrophobicity and signal peptide. Good 20-220aa segment (denoted as N1), 230-300aa segment (denoted as N2) and 335-419aa segment (denoted as N3) are used as segment antigens for screening the dominant epitope of N protein in this example .

[0028] The gene sequences of segments N1 to N3 are:

[0029] The N1 gene sequence is:

[0030] PSDSTGSNQNGERSGARSKQRRPQGLPNNTASWFTALTQHGKEDLKFPRGQGVPINTNSSPDDQIGYYRRATRRIRGGDGKMKDLSPRWYFYYLGTGPEAGLPYGANKDGIIWVATEGALNTPKDHIGTRNPANNAAIVLQLPQGTTLPKGFYAEGSRGGSQASSRSSSRNSSRNSSPRMNSTPNGGSSRGTALP

[0031] The N2 gene sequence is:

[0032] ...

Embodiment 2

[0049] Example 2 S protein dominant epitope segment antigen screening and determination

[0050] According to the full-length amino acid sequence of the new coronavirus S protein published in Genebank, the amino acid sequence of the S protein was analyzed by using the BIOSUN software B-cell epitope analysis and signal peptide analysis functions, and combined with the distribution of functional domains, selected 65- The 290aa segment (denoted as S1), the 340-510aa segment (denoted as S2) and the 520-681aa segment (denoted as S3) three dominant epitope segment antigens are used to screen the S protein dominant epitope in this example segment antigens.

[0051] The S1 gene sequence is:

[0052]fhaihvsgtngtkrfdnpvlpfndgvyfasteksniirgwifgttldsktqsllivnnatnvvikvcefqfcndpflgvyyhknnkswmesefrvyssannctfeyvsqpflmdlegkqgnfknlrefvfknidgyfkiyskhtpinlvrdlpqgfsaleplvdlpiginitrfqtllalhrsyltpgdsssgwtagaaayyvgylqprtfllkynengtitdavd

[0053] The S2 gene sequence is:

[0054] evfnatrfasvyawnrkr...

Embodiment 3

[0065] Example 3 Preparation of N-S dominant epitope fusion protein of novel coronavirus

[0066] In this example, the coding genes of the N1 dominant epitope segment antigen and the S3 dominant epitope segment antigen were linked together for fusion expression using the homologous enzyme method with complementary sticky ends to obtain the N-S dominant epitope fusion protein of the new coronavirus, Its amino acid sequence is as follows:

[0067] PSDSTGSNQNGERSGARSKQRRPQGLPNNTASWFTALTQHGKEDLKFPRGQGVPINTNSSPDDQIGYYRRATRRIRGGDGKMKDLSPRWYFYYLGTGPEAGLPYGANKDGIIWVATEGALNTPKDHIGTRNPANNAAIVLQLPQGTTLPKGFYAEGSRGGSQASSRSSSRSRNSSRNSTPGSSAPATVCGPKKSTNLVKNKCVNFNFNGLTGTGVLTESNKKFLPFQQFGRDIADTTDAVRDPQTLEILDITPCSFGGVSVITPGTNTSNQVAVLYQDVNCTEVPVAIHADQLTPTWRVYSTGSNVFQTRAGCLIGAEHVNNSYECDIPIGAGICASYQTQTNSP

[0068] The nucleic acid sequence of the above-mentioned novel coronavirus N-S dominant epitope fusion protein is as follows:

[0069]CCGAGCGATTCTACCGGCTCCAACCAGAATGGTGAACGTTCGGGCGCGCGCAGCAAACA...

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Abstract

The invention discloses a novel coronavirus N-S dominant epitope fusion protein, a preparation method, application, an expression protein, a microorganism, application and a kit, and belongs to the technical field of genetic engineering. The novel coronavirus N-S dominant epitope fusion protein is a novel coronavirus N protein and S protein dominant epitope fusion protein. The preparation method comprises the following steps: firstly, constructing a fusion expression antigen by an S protein dominant epitope segment antigen and an N protein dominant epitope segment antigen of a novel coronavirus; then removing rare codons through gene optimization to obtain an optimized gene sequence; and finally, obtaining a soluble high-activity antigen with efficient expression. The novel coronavirus N-Sdominant epitope fusion protein disclosed by the invention has soluble expression, high yield, high purity and good activity, and is suitable for accurate diagnosis and detection of patients with thenovel coronavirus.

Description

technical field [0001] The invention belongs to the technical field of genetic engineering, and relates to the preparation and application of a fusion protein, in particular to a novel coronavirus N-S dominant epitope fusion protein, preparation method, application, expression protein, microorganism, application, and kit. Background technique [0002] Coronaviruses are a large family of viruses known to cause colds as well as more serious illnesses such as Middle East Respiratory Syndrome (MERS) and Severe Acute Respiratory Syndrome (SARS). On January 31, 2020, the World Health Organization suggested that the new type of coronavirus pneumonia that appeared in 2019 be temporarily named "2019-nCoV acute respiratory disease", and the virus was temporarily named "2019-nCoV" (2019 represents the year when it first appeared, n means novel (new), CoV means coronavirus (coronavirus)). On February 7, China's National Health and Medical Commission issued a notice, deciding to tempora...

Claims

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Application Information

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IPC IPC(8): C07K19/00C07K14/165C12N15/70C12N1/21G01N33/569C12R1/19
CPCC07K14/005C07K2319/00C12N15/70C12N2770/20022G01N33/56983G01N2333/165
Inventor 陈庆全李丽满
Owner HEBEI GENSURE BIOTECH LIC
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