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Method for efficiently expressing fowl adenovirus Fiber-2 protein in escherichia coli

A technology of fiber-2 and fiber-2-f, which is applied in the field of high-efficiency expression of fiber-2 protein of poultry adenovirus, can solve the problems of high lethality, loss of poultry industry, strong pathogenicity, etc., and achieve reduction of operation steps, Effects of enhanced expression and improved replication efficiency

Pending Publication Date: 2021-04-06
ZHAOQING INST OF BIOTECHNOLOGY CO LTD
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

The serotype 4 epidemic strain HN strain that broke out in 2015 has strong pathogenicity and high lethality, and is quite different from the classic strain, which has brought huge losses to the poultry industry. necessary
[0004] Although the above-mentioned literature and patent methods have achieved the expression of Fiber2 protein, there is still room for further improvement in the preparation of expression vectors and optimization of soluble expression

Method used

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  • Method for efficiently expressing fowl adenovirus Fiber-2 protein in escherichia coli
  • Method for efficiently expressing fowl adenovirus Fiber-2 protein in escherichia coli
  • Method for efficiently expressing fowl adenovirus Fiber-2 protein in escherichia coli

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Embodiment

[0049] (1) Amplification of Fiber-2 target fragment:

[0050] With PET-21a-GX-1-fiber2 (nucleotide sequence is SEQ ID NO: 6, its plasmid map is as follows figure 1 Shown) is the template, Fiber-2-F, Fiber-2-R are the primers to amplify the Fiber-2 target fragment, and the amplification PCR system and program are shown in Table 2 and Table 3 respectively.

[0051] Table 2. Fiber-2 amplification PCR system

[0052]

[0053]

[0054] Table 3. Fiber-2 amplification PCR program

[0055]

[0056] After PCR amplification, 1% nucleic acid gel electrophoresis was carried out for identification, and the results were as follows: image 3 shown. The obvious target band appears at around 1505bp, cut off the gel containing the target band under ultraviolet light, try to make the cut gel size just contain all the target bands, not too large, cut the gel quickly, cut Turn off the ultraviolet light immediately after completion to prevent the target fragment from being degraded by ...

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Abstract

The invention belongs to the technical field of bioengineering, and discloses a method for efficiently expressing avian adenovirus Fiber-2 protein in escherichia coli. A PET-21a-GX-1-fiber2 plasmid and a PXMJ19-C1786T-hexon (T7) plasmid are respectively used as templates, and amplification is carried out by virtue of designed primers, so that a Fiber-2 target fragment and a PXMJ19-C1786-T7 carrier fragment are obtained; then homologous recombination is carried out, and a recombinant plasmid PXMJ19-T7-his-fiber2 is obtained; finally, the recombinant plasmid is transformed into a Shuffle T7-B competent cell to carry out Fiber-2 protein expression. According to the invention, a plasmid capable of increasing the copy number of an exogenous gene is used as a carrier to construct a PXMJ19-T7-his-fiber2 recombinant plasmid so as to enhance the soluble expression of the Fiber-2 protein.

Description

technical field [0001] The invention relates to the technical field of bioengineering, in particular to a method for highly expressing avian adenovirus Fiber-2 protein in Escherichia coli. Background technique [0002] Avian adenoviruses belong to the Adenoviridae family and belong to the genus Avian Adenoviruses. They exist in poultry, and chickens, ducks, geese, etc. are all susceptible hosts. Avian adenovirus is highly contagious and spreads stably, and generally does not cause disease in the host body, and is an opportunistic pathogen. In recent years, the main epidemic strains in China are serotype 4 and 8b avian adenovirus, among which serotype 4 virus strains dominate. FADV4 was first reported in August 1987 and found in Angara, Pakistan. The serotype 4 epidemic strain HN strain that broke out in 2015 has strong pathogenicity and high lethality, and is quite different from the classic strain, which has brought huge losses to the poultry industry. necessary. The fi...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/70C12N15/67C12N15/64C12R1/19
CPCC12N15/70C12N15/67C12N15/64C07K14/005C12N2710/10222
Inventor 方倪冉郑航辉叶俊贤董楠杨小云
Owner ZHAOQING INST OF BIOTECHNOLOGY CO LTD
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