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Recombinant human papilloma virus 16L1 protein and its use

A technology of HPV16L1 and protein, which is applied in the field of preventing cervical cancer caused by HPV16 virus infection, can solve the problems of large protein loss, inability to apply large-scale production, and difficulty of target protein

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 16L1 protein and its use
  • Recombinant human papilloma virus 16L1 protein and its use
  • Recombinant human papilloma virus 16L1 protein and its use

Examples

Experimental program
Comparison scheme
Effect test

Embodiment l

[0073] Embodiment 1: have the construction of the engineering bacterium of HPV16 L1 sequence 2

[0074] 1、 The full-length HPV16 L1 gene was synthesized by GENEWIZ Biotechnology Co., Ltd. (GENEWIZ). Nucleotide sequence SEQ ID NO: 1.

[0075] 2. A template for PCR reaction containing the gene fragment of SEQ ID NO:1. Forward primer sequence: 5'-CGCGGA TCCGGA AA AGAAGATCCCCTTAAAAAA -3'; the restriction endonuclease AccIII site is introduced at the 5' end of the H4 structure, and the sequence of the AccIII site is TCCGGA . Downstream contains XhoI endonuclease site, reverse primer sequence: 5'-GCTCTC CTCGAG TTA TAATGTAAAT TTTGGTTTGG C -3', its 5' end introduces a restriction endonuclease XhoI site, and the sequence of the XhoI site is CTCGAG . HPV16B was amplified by PCR reaction.

[0076] 3. A template containing the gene fragment of SEQ ID NO: 1 for PCR reaction. To contain the introduced restriction endonuclease BamH I site, the BamH I site sequence is GGATCC...

Embodiment 2

[0081] Embodiment 2: Construction of engineering bacteria with HPV16 L1 sequence 4

[0082] 1. The full-length target gene fragment of HPV16 L1 gene was purchased from the waste of clinical cell samples containing wild-type HPV16 virus from the Gynecology Clinic of Beijing Anzhen Hospital. Nucleotide The sequence is SEQ ID NO:3.

[0083] 2. A template containing the gene fragment of SEQ ID NO: 3 for PCR reaction. Forward primer sequence: 5'-CGCGGA TCCGGA AA AGAAGATCCCCTTAAAAAA -3'; the restriction endonuclease AccIII site is introduced at the 5' end of the H4 structure, and the sequence of the AccIII site is TCCGGA . Downstream contains XhoI endonuclease site, reverse primer sequence: 5'-GCTCTC CTCGAG TTA AGAGGTAGAT GAGGTGGTGG G -3', its 5' end introduces a restriction endonuclease XhoI site, the XhoI site sequence is CTCGAG . After PCR reaction, HPV16B was amplified.

[0084] 3. A template containing the gene fragment of SEQ ID NO: 3 for PCR reaction. To co...

Embodiment 3

[0089] Embodiment 3: Construction of engineering bacteria with HPV16 L1 sequence 6

[0090] 1、 The full-length HPV16 L1 gene was synthesized by GENEWIZ Biotechnology Co., Ltd. (GENEWIZ). Nucleotide sequence SEQ ID NO:5.

[0091] 2. A template containing the gene fragment of SEQ ID NO: 3 for PCR reaction. Forward primer sequence: 5'-CGCGGA TCCGGA AA AGAAGATCCCCTTAAAAAA-3'; a restriction endonuclease AccIII site is introduced at the 5' end of the H4 structure, and the sequence of the AccIII site is TCCGGA . Downstream contains XhoI endonuclease site, reverse primer sequence: 5'-GCTCTC CTCGAG TTA AGAGGTAGAT GAGGTGGTGG G-3', its 5' end introduces a restriction endonuclease XhoI site, and the sequence of the XhoI site is CTCGAG . PCR reaction, amplified to obtain HPV16B.

[0092] 3. A template containing the gene fragment of SEQ ID NO: 3 for PCR reaction. To contain the introduced restriction endonuclease BamH I site forward primer sequence: 5'-CGCGGA GGATCC GGA ...

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Abstract

The invention relates to a recombinant human papilloma virus 16L1 protein and its use, and provides a new polynucleotides gene fragment coding the recombinant HPV16L1 protein, a vector containing the gene fragment, a host cell containing the vector, an HPV16L1 protein pentamer expressed by the gene fragment in translation, and an anti-HPV16 infection cervical carcinoma 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 16 L1 protein, and a pentamer composed of it, a vaccine containing the protein and its role in preventing HPV16 virus infection, especially in preventing HPV16 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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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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