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Truncated body based on novel coronavirus RBD-SD1 protein and application of truncated body

A coronavirus and protein technology, applied in the field of biomedicine, can solve problems such as production scale limitations, and achieve the effect of good safety and good immunogenicity

Inactive Publication Date: 2020-12-18
ACAD OF MILITARY SCI PLA CHINA ACAD OF MILITARY MEDICAL SCI INST OF MILITARY VETERINARY MEDICINE +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The inactivated vaccine against new coronary pneumonia has entered phase I / II clinical trials, but the production scale of the inactivated vaccine for new coronary pneumonia needs to consider the biosafety level of the production plant, so the production scale is limited

Method used

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  • Truncated body based on novel coronavirus RBD-SD1 protein and application of truncated body
  • Truncated body based on novel coronavirus RBD-SD1 protein and application of truncated body
  • Truncated body based on novel coronavirus RBD-SD1 protein and application of truncated body

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0055] The acquisition of embodiment 1 recombinant plasmid

[0056] The recombinant vector is obtained by inserting the DNA molecule shown in SEQ ID No.1 into the pPIC9K SnaBI and NotI sites, entrusting Shanghai Jierui Bioengineering Co., Ltd. to obtain a recombinant plasmid.

[0057] The recombinant plasmid pPIC9K-T was sequenced and the result was correct.

Embodiment 2

[0058] Expression and purification of embodiment 2 recombinant protein T

[0059] Transformation and screening of yeast: After the expression vector pPIC9K-T was constructed, a large amount of plasmid DNA was extracted, and after linearization with BglII, 5 μg of the linearized plasmid was taken and transformed into competent Pichia pastoris GS115 by electroporation (P. pastoris GS115 strain was purchased from (from Invitrogen Company), the competent cells transformed by electroporation were spread on MD solid plates, and cultured upside down in a constant temperature incubator at 30°C for 2-3 days until the transformants grew out. Pick the transformant on the YPD+G418 (700μg / mL) medium, culture it at 30°C for 1-2 days, pick a larger white single colony, and inoculate it into a culture tube of 3mL BMGY medium, 30°C, 220rpm Cultivate on a shaking table for 48 hours, centrifuge the bacterial liquid cultured on a shaking table for 48 hours at 3000×g for 15 minutes, remove the sup...

Embodiment 3

[0062] Example 3 Determination of Antigen Immunological Activity

[0063] (1) Immunization method

[0064] grouping:

[0065] Animals 6-week-old Balb / C female mice were selected and divided into PBS group, RBD-SD1 protein+MF59 adjuvant group, T protein+MF59 adjuvant group, with 10 mice in each group.

[0066] Immunization method:

[0067] PBS group, RBD-SD1 protein + MF59 adjuvant group, T protein + MF59 adjuvant group, took leg muscle injection, immunized once every 3 weeks (immunized on day 0, day 21 and day 42 respectively), 10 μg protein / Rats (protein concentration 50 μg / mL), PBS group immunization volume of 200 μL / rat, a total of 3 times of immunization, the first immunization with MF59 adjuvant, mixed at 1:1.

[0068] Blood collection method and time:

[0069] Blood was collected from the tail vein, and the blood collection time was once before immunization, and once on the 7th day after each immunization.

[0070] (2) Determination of antibody titer in serum

[0...

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Abstract

The invention relates to the field of biological medicine, in particular to a truncated body based on novel coronavirus RBD-SD1 protein and application of the truncated body. The protein truncated body disclosed by the invention has a sequence shown as follows: (I) an amino acid sequence shown as SEQ ID No. 1; and / or (II) an amino acid sequence which is obtained by substituting, deleting or addingone or more amino acids to the amino acid sequence shown in (I) and has the same function as the amino acid sequence shown in (I); and / or (III) an amino acid sequence having 80% or more identity withthe amino acid sequence shown in (I) or (II). The protein truncated body can induce relatively strong humoral immune response, has good immunogenicity, and can be applied to development of novel coronavirus vaccines and preparation of neutralizing antibodies.

Description

technical field [0001] The invention relates to the field of biomedicine, in particular to a truncated body based on the novel coronavirus RBD-SD1 protein and its application. Background technique [0002] On January 12, 2020, the World Health Organization (WHO) officially named it "2019 novel coronavirus" (2019 novel coronavirus, 2019-nCoV), and was later named severe acute respiratory virus by the International Virus Taxonomy Committee. Syndrome 2 (severe acute respiratory syndrome coronavirus 2, SARS-CoV-2). On February 11, WHO named the novel coronavirus pneumonia (corona virusdisease 2019, COVID-19). COVID-19 has brought enormous challenges and pressures to all human beings, and has become a global epidemic following the H1N1 influenza virus epidemic, wild-type polio virus epidemic, Ebola epidemic in West Africa, Zika virus epidemic in Brazil, Congo (Kinshasa) It is the sixth time since the Ebola virus outbreak that it has been designated as a public health emergency ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K14/165C12N15/50C12N15/81C12N1/19C07K16/10G01N33/569A61K39/42A61P31/14C12R1/84
CPCC07K14/005C12N15/815C07K16/10G01N33/56983A61P31/14C12N2770/20022G01N2333/165C12N2770/20034A61K2039/505
Inventor 金宁一田明尧黄火清姚斌鲁会军李昌李霄李乐天汪伟
Owner ACAD OF MILITARY SCI PLA CHINA ACAD OF MILITARY MEDICAL SCI INST OF MILITARY VETERINARY MEDICINE
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