Looking for breakthrough ideas for innovation challenges? Try Patsnap Eureka!

Method for constructing recombinant virus for expressing ALV-K envelope protein

A technology of ALV-K and envelope protein, applied in the direction of retroRNA virus, virus, viral peptide, etc., can solve the problem of low expression level of viral protein, no commercial specific ALV-K antigen and antibody detection kit, The detection effect is not very good and other problems, to achieve the effect of speeding up the purification process

Active Publication Date: 2019-02-01
YANGZHOU UNIV
View PDF12 Cites 0 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Due to the weak replication ability of ALV-K virus and the low level of viral protein expression, the current general detection kit for ALV is not very effective in detecting it, and there is no commercial specific ALV-K antigen and antibody detection kit

Method used

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
View more

Image

Smart Image Click on the blue labels to locate them in the text.
Viewing Examples
Smart Image
  • Method for constructing recombinant virus for expressing ALV-K envelope protein
  • Method for constructing recombinant virus for expressing ALV-K envelope protein
  • Method for constructing recombinant virus for expressing ALV-K envelope protein

Examples

Experimental program
Comparison scheme
Effect test

Embodiment

[0021] (1) Preparation of ALV-K proviral DNA:

[0022] Take 400uL of the supernatant after ALV-K infection of the cells in a 1.5mL finger tube, add 400uL of cell lysate (50mmol / L Tris-HCl pH=8.0, 20mmol / L EDTA pH=8.0, 2% SDS and proteinase K), fully After mixing, place it in a 56°C water bath for 4 hours; add 400uL Tris to balance the phenol, mix well and centrifuge at 10000rpm for 10min, take the supernatant into another 1.5mL finger tube; add 400uL phenol: chloroform: isoamyl alcohol, and mix well Then centrifuge at 10,000rpm for 5min, take the supernatant into another 1.5mL finger tube; add 800uL of absolute ethanol, invert and mix well, incubate at -20°C for 30 minutes, centrifuge at 12,000rpm for 15 minutes, discard the supernatant; after natural drying at room temperature Add 30uL sterilized ultrapure water and 2uL RNase to the precipitate, fully dissolve to obtain ALV-K proviral DNA, and store at -20°C for later use.

[0023] (2) PCR amplification of ALV-K env gene fra...

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to View More

PUM

No PUM Login to View More

Abstract

The invention discloses a method for constructing a recombinant virus for expressing an ALV-K envelope protein, specifically comprising the following steps: designing a PCR amplification ALV-J infectious clone linear expression vector primer and a PCR amplification ALV-K-env gene primer sequence; carrying out PCR amplification on an ALV-K envelope protein gene fragment and the ALV-J infectious clone linear vector, constructing a recombinant infectious cloning plasmid, and transfecting the recombinant plasmid into DF-1 cells to rescue the recombinant virus for expression of the ALV-K envelope protein. By ALV-J infectious cloning, the recombinant virus for efficient replication and expression of the ALV-K envelope protein is constructed. The acquisition of the recombinant virus not only laysa foundation for studying functional receptors of the ALV-K envelope protein and its pathogenicity, but also provides a target virus for the clinical detection of neutralizing antibodies of the ALV-K.

Description

technical field [0001] The invention relates to a method for constructing a recombinant virus expressing ALV-K envelope protein. This invention not only lays the foundation for the study of the ALV-K envelope protein functional receptor and its pathogenicity, but also provides a target virus for clinical detection of ALV-K neutralizing antibodies. Therefore, the present invention has certain application value. Background technique [0002] Avian leukemia virus (ALV) is a retrovirus that can cause a variety of tumors and immunosuppressive diseases in poultry. According to the characteristics of its envelope protein, the virus can be divided into 11 subgroups, A-K. Among them, A, B, and J subgroups are more common, and J subgroup is currently the most common in China. ALV-J infection mainly causes malignant proliferation of hematopoietic cells in chickens, causing myeloid cell tumors, hemangiomas and severe immunosuppression, causing huge economic losses to poultry industry...

Claims

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to View More

Application Information

Patent Timeline
no application Login to View More
IPC IPC(8): C12N7/01C12N15/48C12N15/63
CPCC07K14/005C12N7/00C12N15/63C12N2740/11021C12N2740/11022
Inventor 叶建强吕璐李拓凡谢菁邵红霞秦爱建
Owner YANGZHOU UNIV
Who we serve
  • R&D Engineer
  • R&D Manager
  • IP Professional
Why Patsnap Eureka
  • Industry Leading Data Capabilities
  • Powerful AI technology
  • Patent DNA Extraction
Social media
Patsnap Eureka Blog
Learn More
PatSnap group products