CrRNA primer pair for PPV detection, CRISPRCas12a system and application method

An application method and primer pair technology, which is applied in the field of crRNA primer pair and CRISPRCas12a system, can solve the problems of invisible infection and infected pigs without obvious clinical symptoms, and achieve the effects of improving detection efficiency, reducing detection threshold, and saving detection time

Pending Publication Date: 2022-04-05
ANHUI AGRICULTURAL UNIVERSITY +2
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  • Description
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Problems solved by technology

Pregnant sows infected with porcine parvovirus generally show reproductive impairment symptoms, however, infected pigs at other stages have no obvious clinical symptoms and usually show asymptomatic infection

Method used

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  • CrRNA primer pair for PPV detection, CRISPRCas12a system and application method
  • CrRNA primer pair for PPV detection, CRISPRCas12a system and application method
  • CrRNA primer pair for PPV detection, CRISPRCas12a system and application method

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Embodiment Construction

[0060] The following will clearly and completely describe the technical solutions in the embodiments of the present invention with reference to the accompanying drawings in the embodiments of the present invention. Obviously, the described embodiments are only some, not all, embodiments of the present invention. Based on the embodiments of the present invention, all other embodiments obtained by persons of ordinary skill in the art without making creative efforts belong to the protection scope of the present invention.

[0061] The present invention provides a technical solution: a crRNA primer pair for PPV detection, a CRISPRCas12a system and an application method, comprising the following steps:

[0062] S1. Treatment of tissue samples: Weigh about 1 g of each tissue sample, add 1 mL of PBS, put it into a grinding bowl and grind it into a homogenate, transfer it to a sterile centrifuge tube, and centrifuge at 12000 rpm / min for 3 min in a centrifuge;

[0063] S2. Extraction o...

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Abstract

The invention relates to the technical field of virus detection, and discloses a crRNA primer pair for PPV detection, a CRISPRCas12a system and an application method.The method comprises the following steps that S1, tissue sample treatment is conducted, specifically, about 1 g of tissue samples are weighed, 1 mL of PBS is added, the mixture is put into a grinding bowl to be ground into homogenate, the homogenate is transferred into a sterile centrifugal tube, and centrifugation is conducted for 3 min in a centrifugal machine at the speed of 12000 rpm / min; s2, extraction of viral nucleic acid: carrying out nucleic acid extraction on the virus by using a viral genome DNA / RNA extraction kit of a certain biochemical science and technology Co., Ltd; s3, primer design: retrieving a porcine parvovirus whole genome (GenBank ID: NC001718.1) in GenBank, determining a highly conserved fragment in the porcine parvovirus whole genome as a VP2 gene through analysis and comparison by Meglign software, designing a full-length primer of the VP2 gene by using Primer Premier 5 software, and designing a full-length primer of the VP2 gene according to the highly conserved fragment VP2 gene in the porcine parvovirus whole genome; the crRNA primer of the porcine parvovirus VP2 gene is designed by using Cpf1 primer design software in a CRISPR-DT website, and is entrusted to be synthesized and produced by a biological company.

Description

technical field [0001] The invention relates to the technical field of virus detection, in particular to a pair of crRNA primers for PPV detection, a CRISPR Cas12a system and an application method. Background technique [0002] Porcine parvovirus (PPV) is a virus belonging to the Porcine Parvoviridae family. It is a virus that can replicate autonomously and is one of the main pathogens that cause reproductive disorders in sows. Sows infected with the virus can cause symptoms such as miscarriage, mummification and stillbirth, causing reproductive failure of pig mothers, which brings a serious problem to the pig industry. Infection in infected sows (particularly primiparous sows) is characterized by stillbirth, fetal malformations and mummification, but infection can also lead to neonatal death and piglet diseases including diarrhea and dermatitis. All kinds of pigs can be infected with porcine parvovirus, such as domestic pigs, wild boars, newborn piglets, and fattening pigs...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/6816C12Q1/70C12N15/11C12R1/93
Inventor 宋祥军涂健李雅男祁钊邵颖杨侃侃刘红梅姬凯元朱良强吴昊王立刘发全
Owner ANHUI AGRICULTURAL UNIVERSITY
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