Mutated PCV2 (Porcine circovirus Type 2) virus free from being degraded by ubiquitination proteasome, and preparation method and application thereof

A proteasome and ubiquitination technology, applied in biochemical equipment and methods, botanical equipment and methods, applications, etc., can solve problems such as unresearched

CN108148816AActive Publication Date: 2018-06-12NORTHWEST A & F UNIV
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Patent Information

Authority / Receiving Office
CN · China
Current Assignee / Owner
Publication Date
2018-06-12

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Abstract

The invention provides a mutated PCV2 (Porcine circovirus Type 2) virus free from being degraded by ubiquitination proteasome, and a preparation method and application thereof. Three loci (164K, 179Kand 191K) of porcine circovirus 2 type (PCV2, GenBank and No. EU366323) strains Cap are mutated, the mutated PCV2 can not be subjected to MKRN1-mediated ubiquitination degrading, in addition, high speed copying can be carried out through infectious clone, and virus price is high than virus price of wild-type PCV2. Through mutated viruses obtained by the infectious clone, the pathogenic mechanism of the PCV2 can be more favorably researched, and a potential control measure is provided for PCV2 outbreak.
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Description

technical field

[0001] The invention relates to pMKRN1-mediated porcine circovirus type 2 (PCV2) strain Cap ubiquitination, in particular to the construction of PCV2 with amino acid site mutations related to pMKRN1 ubiquitin ligase action. Background technique

[0002] Porcine circovirus associated disease (PCVAD) is caused by porcine circovirus type 2 (PCV2), mainly including porcine dermatitis and nephropathy syndrome (PDNS) , piglet congenital tremors (congenital tremors, CT) and postweaning multisystemic wasting syndrome (postweaning multisystemic wasting syndrome, PMWS) and other diseases. Although PCV2 can replicate in PK-15 cells, it has not been able to reproduce at a high speed, which hinders the research on the pathogenic mechanism of PCV2. How to improve the replication ability of PCV2 is one of the urgent problems to be solved in the study of its pathogenic mechanism.

[0003] Studies have shown that human MKRN1 functions as an E3 ubiquitin ligase on a variety ...

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Embodiment Construction

[0040] The present invention will be further described in detail below in conjunction with the accompanying drawings and embodiments. What has been described is by way of explanation, not limitation, of the invention.

[0041] The present invention detects the effect of pMKRN1 on Cap degradation by co-expressing porcine MKRN1 (pMKRN1) and PCV2Cap to determine the effect of pMKRN1 on PCV2Cap degradation, then constructs a truncated mutant of PCV2Cap protein, and confirms the interaction region between pMKRN1 and PCV2Cap by co-immunoprecipitation assay , and further determined the interaction site between pMKRN1 and PCV2Cap by ubiquitination and degradation analysis. The PCV2 mutant constructed according to the site of action cannot be ubiquitinated, so that the replication ability of PCV2 in cells and in vivo is significantly improved. This discovery facilitates the study of the pathogenic mechanism of PCV2 and provides potential control measures for PCV2 outbreaks.

[0042] ...