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Avian leukosis P27 monoclonal antibody hybridoma cell strain and application thereof

A hybridoma cell line and monoclonal antibody technology, which is applied in the field of serological diagnosis of animal epidemics, can solve the problems of avian leukosis P27 monoclonal antibody with low sensitivity and achieve the effect of low cost, easy operation and improved sensitivity

Inactive Publication Date: 2013-08-14
LANZHOU INST OF ANIMAL SCI & VETERINARY PHARMA OF CAAS +2
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0007] But so far, there is no report of avian leukosis P27 monoclonal antibody with good sensitivity, strong specificity and high titer

Method used

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  • Avian leukosis P27 monoclonal antibody hybridoma cell strain and application thereof
  • Avian leukosis P27 monoclonal antibody hybridoma cell strain and application thereof
  • Avian leukosis P27 monoclonal antibody hybridoma cell strain and application thereof

Examples

Experimental program
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Effect test

preparation example Construction

[0084] Preparation of LB liquid medium: Add 5g of yeast extract, 10g of peptone and 10g of NaCl into the container; add distilled water to 1000ml, adjust the pH to 7.0-7.2 with NaOH and HCl, and pack and sterilize.

[0085] instrument

[0086] Table 1 Instrument source and model

[0087]

[0088]

Embodiment 1

[0089] Embodiment 1. Preparation of the recombinant plasmid of the avian leukosis virus P27 gene

[0090] According to the nucleotide sequence of RSV P27 gene included in GenBank (the sequence is 1381agactggctg atacggtcag gaccaagggc ttacgatccc cgatcactat ggcggaggtg

[0091] 1441 gaagcgctta tgtcctcccc gctgctgccg catgacgtta cgaatctaat gagagttatt

[0092] 1501 ttaggacctg ccccatatgc cttgtggatg gacgcttggg gtgtccaact acagacggtt

[0093] 1561 atagcggcag ccactcgcga cccccgacac ccagcgaatg gtcaagggcg gggggaacgg

[0094] 1621 actaatttgg atcgcttaaa gggtttagct gatgggatgg tgggcaaccc gcagggtcag

[0095] 1681 gccgcattat taagaccggg ggaattggtt gctattacgg cgtcggctct ccaggcattt

[0096] 1741 agagaggttg cccggctggc ggaacctgct ggtccatggg cggacattac gcagggacca

[0097] 1801 tctgagtcct ttgttgattt tgccaatcgg cttataaagg cggttgaggg gtcagatctc

[0098]1861 ccgccttccg cgcgagctcc ggtgatcatt gactgcttta ggcagaagtc acagccagat

[0099] 1921 atccagcagc ttatacgggc agcaccctcc acgctgacca ccccaggaga gataatcaaa ...

Embodiment 2

[0102] Example 2. Expression of Avian Leukosis Virus P27 Protein

[0103] Induced expression of avian leukosis P27 gene in Escherichia coli: transform the recombinant plasmid into Escherichia coli BL21, inoculate it in LB solid medium containing ampicillin (Amp) resistance, culture overnight at 37°C, and take 3~ 4 single colonies were inoculated in LB liquid medium containing Amp, cultured to OD at 37°C with shaking (250rpm) 600 =0.6, add IPTG (isopropyl-β-D-thiogalactopyranoside) to a final concentration of 1mmol / l, and continue to culture at 37°C for 3 hours to obtain the expressed protein, the expression of avian leukemia P27 protein is 565mg / L. At the same time, an uninduced negative control was cultured. The expressed protein was identified by SDS-PAGE and Western-blot, and the results were as follows figure 1 shown.

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Abstract

The invention belongs to the technical field of animal epidemic disease serological diagnosis and relates to an avian leukosis P27 monoclonal antibody hybridoma cell strain and application thereof. The avian leukosis P27 monoclonal antibody obtained by secreting the hybridoma cell strain can be applied to detection of the avian leukosis P27, the 96-pore elisa plate is coated by the P27 specific polyclonal antibody, the P27 monoclonal antibody is labeled by alkaline phosphatase, and according to the avian leukosis P27 monoclonal antibody and the P27 monoclonal antibody labeled by alkaline phosphatase, the detection sensitivity, specificity and stability are effectively improved. A high-efficiency and sensitive detection mode is provided for a novel variety which is used for selecting, purifying and breeding avian leukosis positive chickens and has high genetic resistance, and the avian leukosis P27 monoclonal antibody hybridoma cell strain is low in cost, easy and convenient to operate and is suitable for popularization and application of animal husbandry production.

Description

technical field [0001] The invention belongs to the technical field of serological diagnosis of animal epidemic diseases, and relates to an avian leukosis P27 monoclonal antibody hybridoma cell line and application thereof. technical background [0002] Avian Leukosis (AL) is a general term for a variety of tumor diseases in poultry caused by Avian Leukosis Virus (ALV) belonging to the genus Alpharetrovirus in the Retroviridae family. Avian leukosis virus has been classified into 10 subgroups, named A subgroup to J subgroup respectively. Subgroups A to D are exogenous ALV, and subgroup E is endogenous ALV; subgroups A and B were once considered to be the most common exogenous virus subgroups in commercial layer chickens (especially Laihang chickens) , subgroup C and D infection reports are extremely rare, while subgroup E virus includes the extremely common low pathogenic endogenous leukemia virus; subgroup F is isolated from ring-necked pheasant, and subgroup G is isolated...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/20C07K16/10C07K16/18C07K14/465G01N33/574C12R1/91
Inventor 吴培星宋楠遇秀玲李纯玲宁海强韩涛张继瑜
Owner LANZHOU INST OF ANIMAL SCI & VETERINARY PHARMA OF CAAS
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