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

A kind of porcine igfn1 protein polyclonal antibody and its preparation method and application

A polyclonal antibody and protein technology, applied in the field of porcine IGFN1 protein polyclonal antibody, can solve problems such as unclear, affecting meat quality, and limiting IGFN1 function research, etc., to achieve strong detection specificity, long antigen sequence, and high detection sensitivity Effect

Active Publication Date: 2017-08-25
INST OF ANIMAL SCI OF CHINESE ACAD OF AGRI SCI
View PDF0 Cites 0 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

For example, using a long-chain oligonucleotide chip to analyze the genes differentially expressed in the skeletal muscle of normal turkey and PSE meat turkey, it was found that the expression of IGFN1 gene in PSE meat was down-regulated (Malila et al. PoultryScience. 2013, 92: 1621-1633), It shows that the expression of this gene may affect meat quality, but the expression rule of IGFN1 and its relationship with meat quality have not been reported
In terms of pig research, except for the complete sequence information (XM_005656791) of the IGFN1 gene published by the NCBI database on September 26, 2013, there is no report on the pig IGFN1 gene at home and abroad. Its exact function and whether it will also affect meat quality is not clear
There are only two commercially available IGFN1 antibodies (mouse monoclonal antibody and goat polyclonal antibody), and they are only directed against humans or mice, and these two antibodies are not ideal for detecting IGFN1 expression in pig samples
There is currently no IGFN1 antibody specific to pigs, which greatly limits the functional study of IGFN1 in pigs

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
  • A kind of porcine igfn1 protein polyclonal antibody and its preparation method and application
  • A kind of porcine igfn1 protein polyclonal antibody and its preparation method and application
  • A kind of porcine igfn1 protein polyclonal antibody and its preparation method and application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0042] Example 1 The optimization, cloning and construction of prokaryotic expression vector of porcine IGFN1 gene

[0043] Proceed as follows:

[0044] (1) Sequence optimization of porcine IGFN1 gene and preparation of freeze-dried plasmid. First, optimize the codons of the porcine IGFN1 gene sequence (GenBank, XM_005656791) (see SEQ ID No.5 for the specific nucleotide sequence) to avoid restriction sites EcoRI and BglII, so as to facilitate the construction of subsequent expression vectors, optimize The post porcine IGFN1 gene is composed of the nucleotide sequence shown in SEQ ID No:6. Entrusted Nanjing GenScript Biotechnology Co., Ltd. to artificially synthesize the optimized porcine IGFN1 gene (SEQ ID No: 6) using the GenScript standard vector pUC57 using the EcoRV blunt-end cloning strategy, and obtain the optimized porcine IGFN1 after freeze-drying Gene lyophilized plasmid.

[0045] (2) The amino acid sequence of porcine IGFN1 was analyzed using the CFSSP protein sec...

Embodiment 2

[0053] Example 2 Induced expression and purification test of porcine IGFN1 protein of the present invention

[0054] Proceed as follows:

[0055] (1) The prokaryotic expression vector pET28a-IGFN1-I plasmid prepared in Example 1 was extracted with a plasmid mini-extraction kit from OMEGA, USA.

[0056] (2) Transform the obtained pET28a-IGFN1-I plasmid into BL21 competent cells (Beijing Tiangen Biotechnology Co., Ltd.) to obtain a pET28a-IGFN1-I / BL21 expression strain.

[0057] (3) Inoculate the constructed expression strain pET28a-IGFN1-I / BL21 in LB medium (containing 100 μg / mL ampicillin) for shaking culture at 37°C, activate overnight, and transfer to fresh In LB medium (containing 100 μg / mL ampicillin), shake culture at 37°C and 250 rpm until OD 600 At about 0.6, the final concentrations of 10 μM, 0.1 mM, and 1 mM IPTG (Shanghai Sangon Bioengineering Co., Ltd.) were added, and induced for 4 hours at 37° C. and 250 rpm. Take 1 mL of the bacterial solution, centrifuge at 8...

Embodiment 3

[0059] Example 3 Preparation of porcine IGFN1 protein polyclonal antibody of the present invention

[0060] Proceed as follows:

[0061] Two New Zealand white rabbits were immunized with the IGFN1-I polypeptide obtained in Example 2 (see Table 1 for the immunization methods). Seven days after the fourth immunization, heart puncture blood was collected from each immunized rabbit, antiserum was separated, and the titer and band specificity of the antiserum were detected by Western Blot technology.

[0062] Table 1 Antigen polypeptide IGFN1-I immunization method of the present invention

[0063]

[0064] The His-MBP recombinant protein antigen was coupled to an NHS-activated agarose column, and the antiserum was affinity-purified to obtain a high-purity polyclonal antibody to porcine IGFN1 protein. After the porcine IGFN1 protein was separated by 12% SDS gel electrophoresis, it was transferred to PVDF membrane, and the serum diluted 1:2000 was used as the primary antibody fo...

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 pig IGFN1 protein polyclonal antibody, further discloses antigenic polypeptide for inducing the pig IGFN1 protein polyclonal antibody and belongs to the field of animal gene engineering. The antigenic polypeptide is formed by the amino acid sequence as shown by SEQ ID No: 1. The nucleotide sequence shown by SEQ ID No: 2 forms the antigenic polypeptide encoding genes. The pig IGFN1 protein polyclonal antibody is high in pig IGFN1 protein expression level detection specificity, and an important tool is provided for expression analysis and correlational studies of IGFN1 in pig tissues. The antigen sequence is long, and the detection sensitivity is high.

Description

technical field [0001] The invention belongs to the field of animal genetic engineering, and in particular relates to a porcine IGFN1 protein polyclonal antibody, a preparation method and application of the polyclonal antibody. Background technique [0002] Immunoglobulin-like and fibronectin type III domain containing 1 (IGFN1 for short), is a eukaryotic translation elongation factor 1A (Eukaryotic translation elongation) discovered by Mansilla et al. in 2008 factor 1A, eEF1A) binding protein. The IGFN1 gene is specifically expressed in skeletal muscle and exhibits the domain characteristics of a sarcomere protein combining immunoglobulin I and fibronectin III. At the same time, IGFN1 protein has high homology with fast type myosin binding protein C (myosin binding protein C fast-type) and slow type myosin binding protein C (myosin binding protein C slow-type) in sequence and structure Sex (Mansilla et al. Journal of Cell Biology. 2008, 105:847-858). [0003] The study o...

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
Patent Type & Authority Patents(China)
IPC IPC(8): C07K14/47C12N15/12C12N15/11C07K16/18C07K16/06G01N33/68
CPCC07K14/47C07K16/06C07K16/18G01N33/6803G01N2333/46
Inventor 郝月顾宪红
Owner INST OF ANIMAL SCI OF CHINESE ACAD OF AGRI SCI
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

Browse by: Latest US Patents, China's latest patents, Technical Efficacy Thesaurus, Application Domain, Technology Topic, Popular Technical Reports.

© 2024 PatSnap. All rights reserved.Legal|Privacy policy|Modern Slavery Act Transparency Statement|Sitemap|About US| Contact US: help@patsnap.com