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Recombinant human papilloma virus 18L1 protein and its use

A protein and combination technology, applied in the direction of viruses, viral peptides, antiviral agents, etc., can solve the problems of inability to produce protective antibodies, difficult target proteins, and low expression levels.

Active Publication Date: 2014-09-17
BEIJING HEALTH GUARD BIOTECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, since most of the HPV L1 protein expressed by E. coli loses its natural conformation, it cannot produce protective antibodies against HPV
Or although the above protein can be purified by inclusion bodies, renaturation and other steps can also obtain HPV VLP, but the loss of protein is large during the renaturation process, and the yield is low, so it is difficult to apply in large-scale production.
Although the full-length sequence protein of HPV L1 can also be solublely expressed in the correct conformation in Escherichia coli and dissolved in the lysed supernatant of the bacteria, the expression level is low, and there are many types and large amounts of foreign proteins in the supernatant. It is quite difficult to purify the target protein, and it is still not applicable to large-scale production

Method used

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  • Recombinant human papilloma virus 18L1 protein and its use
  • Recombinant human papilloma virus 18L1 protein and its use
  • Recombinant human papilloma virus 18L1 protein and its use

Examples

Experimental program
Comparison scheme
Effect test

Embodiment l

[0071] Embodiment 1: have the construction of the engineering bacterium of HPV18 L1 sequence 2

[0072] 1、 The full-length HPV18 L1 gene was synthesized by GENEWIZ Biotechnology Co., Ltd. (GENEWIZ). Nucleotide sequence SEQ ID NO: 1, which is derived from GeneBank, the sequence number is GenBank: M14119.1.

[0073] 2. A template for PCR reaction containing the gene fragment of SEQ ID NO:1. With the forward primer sequence: 5'-CGCGGA TCCGGA GAAAATAAGGATCCCTATGAT-3'; a restriction endonuclease AccIII site was introduced at the 5' end of the H4 structure, and the sequence of the AccIII site was TCCGGA. The downstream contains an XhoI endonuclease site, and the reverse primer sequence is: 5'-GCTCTCCTCGAG TTA ACGTTTACGA GGGCCTACGG T -3', and its 5' end introduces a restriction endonuclease XhoI site, and the sequence of the XhoI site is CTCGAG. HPV18B was amplified by PCR reaction.

[0074] 3. A template containing the gene fragment of SEQ ID NO: 1 for PCR reaction. To contain...

Embodiment 2

[0079] Embodiment 2: Construction of engineering bacteria with HPV18 L1 sequence 4

[0080] 1. The target gene fragment of the full-length HPV18 L1 gene was purchased from the waste of clinical cell samples containing wild-type HPV18 virus from the Gynecology Clinic of Beijing Anzhen Hospital. Nucleotide The sequence is SEQ ID NO: 3 (GenBank: FN870689.1).

[0081] 2. A template containing the gene fragment of SEQ ID NO: 3 for PCR reaction. With the forward primer sequence: 5'-CGCGGATCCGGA GAAAATAAGGATCCCTATGAT-3'; a restriction endonuclease AccIII site was introduced at the 5' end of the H4 structure, and the sequence of the AccIII site was TCCGGA. The downstream contains the XhoI endonuclease site, the reverse primer sequence: 5'-GCTCTCCTCGAG TTA AGGTTTAGAA GACGTAGTGG C -3', the 5' end introduces the restriction endonuclease XhoI site, and the XhoI site sequence is CTCGAG. After PCR reaction, HPV18B was amplified.

[0082] 3. A template containing the gene fragment of...

Embodiment 3

[0087] Embodiment 3: Construction of engineering bacteria with HPV18 L1 sequence 6

[0088] 1、 The full-length HPV18 L1 gene was synthesized by GENEWIZ Biotechnology Co., Ltd. (GENEWIZ). Nucleotide sequence SEQ ID NO: 5, which is derived from GeneBank, the sequence number is GenBank: FN870694.1.

[0089] 2. A template containing the gene fragment of SEQ ID NO: 3 for PCR reaction. With the forward primer sequence: 5'-CGCGGATCCGGA GAAAATAAGGATCCCTATGAT-3'; a restriction endonuclease AccIII site was introduced at the 5' end of the H4 structure, and the sequence of the AccIII site was TCCGGA. The downstream contains the XhoI endonuclease site, the reverse primer sequence: 5'-GCTCTCCTCGAG TTA AGGTTTAGAA GACGTAGTGG C -3', the 5' end introduces the restriction endonuclease XhoI site, and the XhoI site sequence is CTCGAG. PCR reaction, amplified to obtain HPV18B.

[0090] 3. A template containing the gene fragment of SEQ ID NO: 3 for PCR reaction. The forward primer sequence con...

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Abstract

The invention relates to a recombinant human papilloma virus 18L1 protein and its use, and provides a new polynucleotides gene fragment coding the recombinant HPV18L1 protein, a vector containing the gene fragment, a host cell containing the vector, an HPV18L1 protein pentamer expressed by the gene fragment in translation, and an anti-HPV18 infection condyloma acuminatum vaccine composed of the pentamer.

Description

technical field [0001] The present invention relates to the prevention and / or treatment of human papilloma virus infection. More specifically, the present invention relates to a recombinant human papillomavirus type 18 L1 protein, and a pentamer composed of it, a vaccine containing the protein and its role in preventing HPV18 virus infection, especially in preventing HPV18 type Use in cervical cancer disease caused by viral infection. Background technique [0002] Human Papillomavirus (HPV) is a DNA virus transmitted through close contact. In human tissues, HPV mainly infects skin and mucous membrane tissues. HPV can be divided into high-risk type and low-risk type. Long-term persistent infection of high-risk type can cause cancer, such as cervical cancer, and endanger life. Low-risk HPV can cause genital lesions, such as genital warts, affecting quality of life. Cervical cancer is the second most malignant tumor in women. The annual global incidence is about 540,000 (20...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K14/025C12N15/37C12N15/70C12N1/21A61K39/12A61P31/20A61P35/00C12R1/93
CPCA61K39/00C07K14/005C12N2710/20022C12N2710/20034
Inventor 刘永江陈小江陈林盖大海许铮曹科陈建平潘勇昭银飞阮芳勇
Owner BEIJING HEALTH GUARD BIOTECH
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