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Fusion protein, use thereof, and anti-malarial vaccine and antibody thereof

A fusion protein and anti-malarial technology, applied in the field of biomedicine, can solve the problem of difficulty in obtaining artificial polyepitope antigens, and achieve the effects of good development prospects, stable physical and chemical properties, and inhibition of in vitro growth.

Active Publication Date: 2013-06-19
NAT VACCINE & SERUM INST
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, in actual research, there is a huge diversity in the combination sequence of multiple epitopes connected to each other, and it is difficult to obtain an artificial multi-epitope antigen with an optimal connection sequence by manual operation.

Method used

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  • Fusion protein, use thereof, and anti-malarial vaccine and antibody thereof
  • Fusion protein, use thereof, and anti-malarial vaccine and antibody thereof
  • Fusion protein, use thereof, and anti-malarial vaccine and antibody thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment approach

[0056] According to one embodiment of the present invention, the M.RCAg-1 gene is cloned into the prokaryotic expression vector pDS-ex containing different carrier tags or no carrier tag, expressed and purified to obtain M.RCAg-1 / Exp.V-1 , M.RCAg-1 / Exp.V-2, M.RCAg-1 / Exp.V-3 three kinds of multi-epitope proteins, the amino acid sequences of which are respectively as SEQ ID No.: 1, SEQ ID No.: 3 and Shown in SEQ ID No.: 5; its DNA sequences are shown in SEQ ID No.: 2, SEQ ID No.: 4 and SEQ ID No.: 6, respectively.

[0057] figure 1 Shows the amino acid sequence comparison schematic diagram of M.RCAg-1 / Exp.V-1, M.RCAg-1 / Exp.V-2 and M.RCAg-1 / Exp.V-3 three kinds of multi-epitope proteins, wherein M.RCAg-1 / Exp.V-1 is a multi-epitopic protein with 43 carrier amino acids fused at the N-terminus with an enterokinase digestion recognition site, and M.RCAg-1 / Exp.V-2 is in A multi-epitope protein with 43 carrier amino acids fused to the N-terminus with a PreScission Protease cleavage re...

Embodiment 1

[0063] Embodiment 1 constructs VR1012-312 plasmid and pDS-ex plasmid

[0064] According to the method disclosed in Chinese Patent Application No. 200410080982.6, VR1012-312 plasmid and pDS-ex plasmid were constructed. details as follows:

[0065] According to the method disclosed in Chinese Patent Application No.200410080982.6, the gene fragment m.rcag-1 (that is, the ES312 gene fragment disclosed in Chinese Patent Application No.200410080982.6) was obtained, and the resulting gene fragment m.rcag-1 was connected to pDS- On the ex plasmid (according to the method disclosed in Chinese Patent Application No.200410080982.6, the prokaryotic expression vector pET-30a (+) (obtained from Novagen) was replaced by Bgl II with the Not I and Eag I restriction sites by PCR technology, Kpn I is moved forward, and EcoRV, Bgl II, and Nsp V restriction sites are removed, thereby obtaining the pDS-ex plasmid), its DNA sequence is shown in SEQ ID No.: 10, and its physical map is Figure 7 sho...

Embodiment 2

[0066] Embodiment 2 obtains M.RCAg-1 / Exp.V-1 protein

[0067] 1. The VR1012-312 plasmid and pDS-ex plasmid obtained in Example 1 were subjected to EcoR I and BglII double enzyme digestion respectively, so that m.rcag-1 was connected into the pDS-ex vector, thereby constructing the recombinant plasmid M.RCAg- 1 / pDS-ex.

[0068] The specific steps are as follows:

[0069] 1) The restriction enzyme digestion system of VR1012-312 plasmid and pDS-ex plasmid (taking EcoR I and Bgl II as examples) is as follows:

[0070]

[0071] Digest at 37°C for 2 hours, identify by agarose gel electrophoresis, and recover with a DNA fragment recovery kit to obtain gene fragment m.rcag-1 and pDS-ex plasmid fragment.

[0072] 2) Dephosphorylation of the recovered gene fragment m.rcag-1 and pDS-ex plasmid fragment

[0073] Concrete reaction system is as follows:

[0074]

[0075] 3) In the ligation reaction of the recovered gene fragment m.rcag-1 and the pDS-ex plasmid fragment, the molar ...

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Abstract

The invention provides use of a fusion protein, and an anti-malarial vaccine and an antibody thereof. The amino acid sequence of the fusion protein of the present invention from N to C terminal in sequence comprises an amino acid sequence shown in SEQ ID No.: 7 and an amino acid sequence shown in SEQ ID No.: 5. The fusion protein of the invention and the vaccine have advantages and positive effects that: the recombinant protein has strong immunogenicity, physicochemical properties are stable, sustainable and high-titer specific antibodies can be stimulated, plasmodium falciparum natural antigens can be identified in mouse immune sera, significant T-cell response can be induced, and in vitro growth of plasmodium falciparum can be suppressed, and the vaccine is a high promising candidate and has good development prospects.

Description

technical field [0001] The invention belongs to the field of biomedicine, in particular, it relates to a fusion protein and its use in the preparation of medicines for preventing and / or treating malaria, an anti-malarial vaccine containing the fusion protein, and antibodies or hybridoma cells or antiserum. Background technique [0002] Malaria, transmitted by mosquitoes, is the oldest disease that threatens human life. It has a history of thousands of years, but it is still listed by the World Health Organization (WHO) as one of the three most serious infectious diseases in the world. According to the latest statistics, it is estimated that about 40% of the world's population is at risk of suffering from malaria, and 500 million people are infected with Plasmodium every year, and about 1 million people die from falciparum malaria, most of whom are under five years old children and pregnant women. Malaria not only poses a serious threat to the life and health of people in d...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K19/00C07K16/20C07K16/06C12N15/62C12N5/20A61K39/015A61K39/395A61P33/06
CPCY02A50/30
Inventor 王健张振龙
Owner NAT VACCINE & SERUM INST
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