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Method for constructing two novel prawn expression vectors

The technology of an expression vector and a construction method, which is applied in the construction of two new shrimp expression vectors, can solve the problems affecting the sustainable development of the shrimp aquaculture industry and the lack of a good solution for virus prevention and control.

Inactive Publication Date: 2013-03-20
GUANGXI INST OF FISHERIES
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, due to the widespread spread of shrimp viruses, the economic loss of tens of billions of yuan is caused to the shrimp farming industry every year, which seriously affects the sustainable development of the shrimp farming industry.
Although great progress has been made in the diagnosis technology of shrimp pathogens, there is still no good solution for virus control.
There is no report on the transformation of pcDNA3.1 using the IE1 promoter

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  • Method for constructing two novel prawn expression vectors
  • Method for constructing two novel prawn expression vectors
  • Method for constructing two novel prawn expression vectors

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Embodiment Construction

[0038] The present invention will be further described in detail below in conjunction with the accompanying drawings and specific experiments.

[0039] figure 1 It is the electrophoresis result diagram of the PCR amplification WSSV promoter IE (-94 / +52) of the present invention.

[0040] In the picture: figure 1 The PCR method is as follows: refer to the WSSV genome sequence in GenBanK to synthesize upstream and downstream primers for the early gene promoter IE (-94 / +52) of the WSSV virus, see Table 1. The PCR method amplified WSSV promoter IE (-94 / +52). Take 10 μL of PCR product for agarose gel electrophoresis. Lane M is DNA molecular marker, and lane 1 is IE (-94 / +52). Results: The size of the IE (-94 / +52) band was 146bp, which was consistent with the expected result, indicating that IE (-94 / +52) of WSSV was amplified.

[0041] figure 2 It is the electrophoresis result diagram of the PCR amplification WSSV promoter IE (-945 / +52) of the present invention.

[0042] I...

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Abstract

The invention discloses a method for constructing two novel prawn expression vectors. A gene-specific primer is designed, a prawn white spot syndrome virus (WSSV) virus is taken as a template, different length fragments of immediately-early gene (IE) (-94 / +52) and IE (-945 / +52) of an immediately-early gene promoter IE 1 are amplified; and a restriction enzyme digestion and connection method is utilized, the IE (-94 / +52) and the IE (-945 / +52) are taken to replace a cytomegalovirus (CMV) promoter of an eukaryotic expression vector pc deoxyribose nucleic acid 3.1(pc DNA 3.1)-green fluorescent protein (GFP), so that two novel expression vectors pcDNA 3.1-IE (-94 / +52)-GFP and pcDNA 3.1-IE (-945 / +52)-GFP which contain prawn cytomegalovirus promoters are constructed. The constructed expression vectors are mixed with a cationic lipofectin transfection reagent TransLipid TM for the intramuscular injection of prawns, results show that the two expression vectors can well express prawn cells, and moreover, host prawns do not have any obvious toxic action. Consequently, the two expression vectors can be used for researching prawn oral vaccines or prawn cytobiology.

Description

technical field [0001] The invention relates to the field of molecular biology research on prawn cells and the development of anti-disease drugs for prawns, and specifically relates to the construction method of two novel expression vectors for prawns. Background technique [0002] Litopenaeus vannamei occupies an extremely important position in my country's mariculture industry. However, due to the widespread spread of shrimp viruses, the economic loss of tens of billions of yuan is caused to the shrimp farming industry every year, which seriously affects the sustainable development of the shrimp farming industry. Although great progress has been made in the diagnosis technology of shrimp pathogens, there is still no good solution in virus prevention and control. Shrimp antiviral drugs are still in the research stage. The main research directions include subunit vaccines, nucleic acid vaccines, yolk antibodies, and Chinese herbal medicines. Among them, there are many resea...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/85C12N15/66
Inventor 黎铭陈晓汉马春霞谢达祥赵永贞
Owner GUANGXI INST OF FISHERIES