Anti-coccidiosis multivalent recombinant protein IgY (Immunoglobulin of yolk) as well as preparation method and application thereof

A technology of egg yolk antibody and recombinant protein, which is applied to the preparation method of peptide, immunoglobulin, botanical equipment and methods, etc., can solve the problems of low production cost and drug resistance, and achieve the reduction of oocyst excretion, survival rate and The effect of increased relative weight gain and good anticoccidial effect

Active Publication Date: 2020-07-31
CHINA AGRI UNIV
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0005] The purpose of the present invention is to provide an anti-coccidia multivalent recombinant protein egg yolk antibody, which aims to solve the problem of drug resistance in the current prevention and control of chicken coccidiosis drugs. The anti-chicken coccidia multivalent recombinant protein egg yolk antibody is immune The presence of specific maternal antibodies can be detected in the yolk of chicken queens, and the preparation of corresponding multivalent recombinant protein yolk antibodies can effectively provide immune protection for chicks. The multivalent recombinant protein yolk antibodies have good immune protection effects and low production costs, and can To a certain extent, improve the existing problems in the prevention and control of coccidiosis

Method used

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  • Anti-coccidiosis multivalent recombinant protein IgY (Immunoglobulin of yolk) as well as preparation method and application thereof
  • Anti-coccidiosis multivalent recombinant protein IgY (Immunoglobulin of yolk) as well as preparation method and application thereof
  • Anti-coccidiosis multivalent recombinant protein IgY (Immunoglobulin of yolk) as well as preparation method and application thereof

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0046] Expression and purification of embodiment 1 recombinant protein

[0047] In the present invention, the antigen of Eimeria tenella is EtROP. The accession number in the ToxoDB database is ETH_00005405 (https: / / toxodb.org / toxo / app / record / gene / ETH_00005405), the length of the gene fragment encoding the protein The accession number of EtGAM22 in the ToxoDB database is ETH_00035480 (https: / / toxodb.org / toxo / app / record / gene / ETH_00035480), and the length of the gene fragment encoding the protein is 597bp; the accession number of EmGAM56 in the ToxoDB database The number is EMWEY_00026710 (https: / / toxodb.org / toxo / app / record / gene / EMWEY_00026710), and the length of the gene fragment encoding the protein is 1422bp; the accession number of Ea3-1E in the ToxoDB database is EAH_00057690 (https: / / toxodb.org / toxo / app / record / gene / EAH_00057690), the length of the gene fragment encoding the protein is 1737bp.

[0048] 1. Cloning of genes and construction of recombinant plasmids

[0049]...

Embodiment 2

[0067] Embodiment 2 specific yolk antibody level monitoring

[0068] 1. Immunization procedure

[0069] Purchase 24-26 week-old Hainan brown hens, and randomly divide them into 3 groups, 15 in each group. The breeder hens were given the first immunization four weeks before the hen was born, and the four recombinant proteins were mixed with 100ug ( / feather) of Freund's complete adjuvant in a ratio of 1:1, and the chest was subcutaneously injected; two weeks later, the second immunization was carried out. After mixing 100ug ( / feather) of each of the four recombinant proteins with Freund's incomplete adjuvant 1:1, inject subcutaneously in the chest; four weeks after the second immunization, perform the third immunization, and mix 100ug ( / feather) of each of the four recombinant proteins with Freund's incomplete adjuvant. The adjuvant was mixed 1:1 and injected subcutaneously in the chest. The control group was treated in the same way as the protein immunization group. See Table ...

Embodiment 3

[0085] The evaluation of embodiment 3 egg yolk antibody preventive effect

[0086] 1. Evaluation of the preventive effect of egg yolk antibody

[0087] 1.1 Experimental design

[0088] The experiment was divided into 15 groups, with seven 1-day-old chicks in each group, among which 0.3ml, 0.5ml, and 1.0ml of recombinant protein yolk antibody were orally administered to groups 1-3 at the age of 11-13 days, and groups 4-6 were orally administered Blank yolk antibody 0.3ml, 0.5ml, 1.0ml, groups 7-9 were injected with recombinant protein yolk antibody 0.3ml, 0.5ml, 1.0ml respectively, groups 10-12 were injected with blank yolk antibody 0.3ml, 0.5ml, 1.0ml , the 13th group is the challenge control (7 feathers), and the 14th group is the blank control (7 feathers). After being weighed at 14 days old, the sporulated oocysts mixed with fresh Eimeria tenella, Eimeria giga and Eimeria pileus were orally infected with doses of 5000 / feather and 1.5×10 oocysts respectively. 5 pcs / feathe...

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Abstract

The invention discloses anti-coccidiosis multivalent recombinant protein IgY (Immunoglobulin of yolk) as well as a preparation method and an application thereof. The preparation method of the anti-coccidiosis multivalent recombinant protein IgY comprises the steps as follows: linking EtROP41, EtGAM22, EmGAM56 and Ea3-1E with Escherichia coli expression vectors respectively to obtain recombinant expression vectors, expressing recombinant proteins, mixing different recombinant proteins with adjuvants in a certain proportion, immunizing hens, collecting eggs to prepare IgY. Compared with the control group, after the prepared IgY is used for preventing and treating coccidiosis of chicks, the IgY can increase the survival rate and relative weight gain rate of the chicks and reduce excretion ofoocysts, and shows a good anti-coccidiosis effect and the application prospect in treatment and prevention of coccidiosis.

Description

technical field [0001] The invention relates to the technical field of biological and veterinary medicines, in particular to an anti-chicken coccidia multivalent recombinant protein yolk antibody and its preparation method and application. Background technique [0002] Chicken coccidiosis (Coccidiosis), an intestinal disease caused by one or several species of Eimeria coccidiosis, is also the most economically significant parasitic infection in poultry farming worldwide. Global economic losses due to coccidiosis in chickens are about billions of dollars each year, mainly including the cost of prevention and the loss of production performance (such as low feed conversion ratio, slow growth, death). At present, the prevention and treatment of coccidiosis mainly depends on anticoccidiosis drugs and live vaccines, but due to problems such as drug-resistant strains and drug residues, the use of anticoccidiosis drugs is limited; the preservation, transportation, application and M...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K16/20C07K16/02C07K14/455C07K1/34C12N15/30C12N15/70A61K39/395A61P33/02A23K50/75A23K20/195
CPCC07K16/20C07K16/02C07K14/455C12N15/70A23K50/75A23K20/195A61P33/02C07K2317/11A61K2039/505A61K2039/542A61K2039/54
Inventor 刘群杨旭宋星桔安同伟刘晶陈庆忠许建海
Owner CHINA AGRI UNIV
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