Looking for breakthrough ideas for innovation challenges? Try Patsnap Eureka!

Primer, probe and kit for identifying serum I-type Mardivirus gene deletion vaccine, attenuation vaccine and wild virus

A technology of gene deletion vaccine and Marek virus, applied in the direction of microorganism-based methods, microorganisms, biochemical equipment and methods, etc., can solve the problems of indistinguishability, achieve the effect of promoting immune prevention and control, and broad market application value

Pending Publication Date: 2018-05-15
SHANDONG AGRICULTURAL UNIVERSITY
View PDF4 Cites 7 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, this method cannot distinguish between the serum type I gene deletion vaccine and the passage attenuated vaccine in the serum type I vaccine virus, and the serum type I gene deletion vaccine and wild virus

Method used

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
View more

Image

Smart Image Click on the blue labels to locate them in the text.
Viewing Examples
Smart Image
  • Primer, probe and kit for identifying serum I-type Mardivirus gene deletion vaccine, attenuation vaccine and wild virus
  • Primer, probe and kit for identifying serum I-type Mardivirus gene deletion vaccine, attenuation vaccine and wild virus
  • Primer, probe and kit for identifying serum I-type Mardivirus gene deletion vaccine, attenuation vaccine and wild virus

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0070] Embodiment 1: the design of fluorescence quantitative PCR primer and probe

[0071] Through the detailed comparison and analysis of the MDV genome sequence that has been published, especially the cloned and purified MDV genome (the accession numbers of the reference sequences of the present invention in NCBI are respectively: CVI988 / Rispens-DQ530348, GX0101-JX844666, 584A-EU627065, RB1B-EF523390, Md5-AF243438, GA-AF147806, Md11-AY510475, LMS-JQ314003), especially all current MD serum type I gene deletion vaccine genome sequences designed to be able to differentially diagnose serum type I gene deletion vaccine, serum type I Primers and probes of vaccine CVI988 / Rispens and MD wild virus.

[0072] The primer and probe sequences designed to distinguish chicken Marek's disease serum type I gene deletion vaccine from wild virus are as follows:

[0073] Upstream primer: ALMDV-F:5'-CCTACAGTCCCGCTGACGAT-3' (SEQ ID NO.1)

[0074] Downstream primer: ALMDV-R: 5'-CTGTTGGGGATGTCGTG...

Embodiment 2

[0079] Embodiment 2: Establishment of fluorescent quantitative PCR identification method

[0080] 1. Culture of serum type I MDV

[0081] Serum type I vaccine CVI988 / Rispens was purchased from Beijing Lingyu Biotechnology Co., Ltd. Serum type I gene deletion vaccine SC9-1 (recorded in patent CN102628053B), serum type I MDV Md5, GX0101 were all kept by our laboratory. Melt the serum type I MDV frozen in liquid nitrogen quickly in a 37°C water bath, put it into a high-pressure sterilized 1.5ml centrifuge tube in an ultra-clean bench, centrifuge at 2000 rpm for 3 minutes at 4°C, and remove as much as possible. Cell cryopreservation solution, 1ml of cell maintenance solution was gently resuspended and added to the cell bottle in an appropriate amount, placed in a 37°C, 5% carbon dioxide (CO2) incubator for 3-5 days, and after the typical plaques of MDV appeared, follow the Cells were collected by digesting the cells.

[0082] 2. Extraction of MDV DNA

[0083] about 10 7 Add 50...

Embodiment 3

[0097] Example 3: The composition of the kit, the optimization of experimental parameters and the investigation of specificity, sensitivity and repeatability

[0098] 1. The composition of the kit:

[0099] The kit of this embodiment is used for differential diagnosis of serum type I gene deletion vaccine, serum type I vaccine CVI988 / Rispens and MD wild virus, and contains ALMDV detection primers and probes (SEQ ID NO.1-3), FIMDV Detect primers and probes (SEQ ID NO.4-6), positive standard plasmids and fluorescent quantitative PCR reaction reagents.

[0100] Wherein, the positive standard plasmid is pMP recombinant plasmid. The tandem sequence 1324bp of the first 450bp of the MDV meq gene synthesized by a commercial sequencing company and the MDV wild virus pp38 gene was cloned into the pMD19 vector and constructed.

[0101] Fluorescent quantitative PCR reaction reagents include: Gene Expression Master Mix buffer and ddH 2 O.

[0102] 2. Optimization of experimental para...

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to View More

PUM

No PUM Login to View More

Abstract

The invention discloses a primer, a probe and a kit for identifying serum I-type Mardivirus gene deletion vaccine, attenuation vaccine and wild virus. The primer and the probe comprise a first pair ofprimer and probe and a second pair of primer and probe, wherein the nucleotide sequence of the first pair of primer and probe is shown as SEQ ID NO. 1 to SEQ ID NO. 3; the nucleotide sequence of thesecond pair of primer and probe is shown as SEQ ID NO. 4 to SEQ ID No. 6. A fluorogenic quantitative PCR reaction is carried out by using the primers and the probes provided by the invention, the serum I-type Mardivirus gene deletion vaccine, attenuation vaccine and wild virus can be identified and diagnosed quickly, accurately, scientifically and objectively, and the immune state of chicken vaccines and the infection progress of wild virus are definite through accurate detection for vaccine virus and wild virus copy number.

Description

technical field [0001] The invention relates to the technical field of biological products, in particular to primers, probes and kits for identifying serotype I Marek virus gene-deleted vaccines, passaged attenuated vaccines and wild virus. Background technique [0002] Chicken Marek's disease (Marek`s Disease, MD) is caused by Marek's virus (Marek`s Disease Virus, MDV), one of the most common T lymphocyte proliferative diseases in chickens, peripheral nerve, iris, skin, muscle and Lymphocyte infiltration, hyperplasia, and tumor formation in various internal organs are characteristic. [0003] MDV is divided into 3 serotypes. Serum type I is the pathogenic type. According to the strength of virulence, MDV strains are divided into attenuated, virulent, super-virulent and super-virulent. The commercialized MD vaccine commonly used at present is the attenuated type I strain of CVI988 / Rispens. The immunization of MD vaccine effectively controlled the outbreak of MD, but with ...

Claims

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to View More

Application Information

Patent Timeline
no application Login to View More
Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/70C12Q1/6851C12N15/11C12R1/93
CPCC12Q1/6851C12Q1/701C12Q2600/16C12Q2531/113C12Q2537/143C12Q2561/101
Inventor 苏帅崔宁孙淑红
Owner SHANDONG AGRICULTURAL UNIVERSITY
Who we serve
  • R&D Engineer
  • R&D Manager
  • IP Professional
Why Patsnap Eureka
  • Industry Leading Data Capabilities
  • Powerful AI technology
  • Patent DNA Extraction
Social media
Patsnap Eureka Blog
Learn More
PatSnap group products

Browse by: Latest US Patents, China's latest patents, Technical Efficacy Thesaurus, Application Domain, Technology Topic, Popular Technical Reports.

© 2024 PatSnap. All rights reserved.Legal|Privacy policy|Modern Slavery Act Transparency Statement|Sitemap|About US| Contact US: help@patsnap.com