A kind of anti-goat pox virus vhh-4-1 based on Bactrian camel and its application

A goat pox virus and gene technology, applied in the field of biology, can solve the problems of low accuracy and sensitivity, high cost, and time-consuming, and achieve the effects of small molecular weight, easy expression, and good solubility

Active Publication Date: 2020-06-09
LANZHOU INST OF VETERINARY SCI CHINESE ACAD OF AGRI SCI
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0004] In order to solve the problems of time-consuming, laborious, high cost, and low accuracy and sensitivity in the prior art, the present invention has developed a key marker for anti-capillary pox virus, and provides a Bactrian camel-derived anti- Goat Poxvirus Single Domain Antibody VHH-4-1

Method used

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  • A kind of anti-goat pox virus vhh-4-1 based on Bactrian camel and its application
  • A kind of anti-goat pox virus vhh-4-1 based on Bactrian camel and its application
  • A kind of anti-goat pox virus vhh-4-1 based on Bactrian camel and its application

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Experimental program
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Effect test

Embodiment

[0061] 1. Screening of a Bactrian camel anti-goat pox virus single domain antibody VHH-4-1 gene

[0062] 1.1 Use camel-derived single-domain antibody homology arm primers to amplify the single-domain antibody gene from the constructed cDNA antibody library, 900bp is the VH-CH1-CH2 hinge region, 600bp is the VHH-CH2 hinge region, and VHH is about 400bp, and gel recovery 900 and 600bp bands, on this basis, amplified 400bp again, after gel recovery, ligated with T carrier, transformed JM109 competent cells, picked spots, shaken bacteria, identified, verified the appropriate sequence for sequencing. 50ul reaction system: 25ul Ex Taq mix 25ul, targeted upstream and downstream primers 1ul each, template 1ul (cDNA library plasmid), ddH 2 O 22ul, amplification conditions: 95°C for 3min, 95°C for 50s, 54°C for 50s, 35times, 72°C for 50s. The conditions of the three fragments are consistent.

[0063] 1.2 The obtained sequence and figure 1 The single-domain antibody framework is compa...

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Abstract

The present invention provides a two-humped camel-based anti-goat pox virus VHH-4-1 and a use thereof, and belongs to the field of biotechnology. The anti-goat pox virus VHH-4-1 is prepared by VHH-4-1screening, expressing and purifying, functionalized labeling and activity verifying, anti-goat pox virus concentrating and purifying, optimizing various indicators of a kit, etc. Due to the fact thatthe provided kit uses the functionalizedly labeled VHH-4-1, the kit avoids steps of primary antibodies, secondary antibodies, anti-antibodies, etc., improves sensitivity and stability, shortens time,saves resources and breaks through drawbacks that traditional Elisa methods are time-consuming, laborious, etc. The provided anti-goat pox virus VHH-4-1 provides a basis for establishing sensitive goat pox virus clinical diagnosis method and studying a virus infection mechanism, and also provides a key marker for the developed goat pox virus antibody solid phase competition Elisa kit. In addition, goat pox attenuated vaccine is used to be subjected to cell enlarge cultivation, concentration and purification as a coating antigen to improves the sensitivity.

Description

technical field [0001] The invention belongs to the field of biology, relates to Bactrian camel origin, in particular to an anti-goat pox virus single domain antibody VHH-4-1 based on Bactrian camel origin and its application. Background technique [0002] Sheep pox is an acute, febrile, highly contagious infectious disease caused by sheep pox / goat pox virus or mixed infection, causing fever, papules or herpes on the skin and mucous membranes of sheep and goats with less or no hair. It is the most serious of all animal pox diseases. Because of the similarity of clinical symptoms with sheep contagious pustule (commonly known as sheep mouth sore), the crossover of serological reactions, and the frequent occurrence of visceral type sheep pox have caused greater difficulties in the diagnosis of the disease. [0003] Single-domain antibody (VHH) is currently available as the smallest structural unit with stable structure, complete function, and ability to bind antigens. Due to i...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C07K16/08C07K1/34G01N33/535G01N33/569
CPCC07K16/081C07K2317/22C07K2317/569G01N33/535G01N33/56983G01N2333/065
Inventor 吴锦艳尚佑军田宏曹小安惠小婷关玉华王耀杰刘湘涛刘永生
Owner LANZHOU INST OF VETERINARY SCI CHINESE ACAD OF AGRI SCI
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