A kind of multiplex immunofluorescence analysis primer, kit and method for detecting chicken Marek's disease virus and chicken infectious anemia virus
A technology for chicken Marek's disease and chicken infectivity, which is applied in the field of virus detection in the breeding industry, can solve the problems of increased experimental difficulty and higher requirements, and achieves the effects of less sample amount, lower detection cost, and strong specificity.
- Summary
- Abstract
- Description
- Claims
- Application Information
AI Technical Summary
Problems solved by technology
Method used
Image
Examples
Embodiment 1
[0079] Example 1 Primer Design
[0080] After screening a large number of designed primers, it was found that the primer pair C1 and C2, M1 and M2 have the best effect on simultaneous detection of chicken infectious anemia virus (CAV) and chicken Marek's disease virus (MDV) for multiplex immunofluorescence analysis , whose base sequence is shown below.
[0081] Primer C1: 5'-CGACATCGGAGGAGACAG-3' (SEQ ID NO: 1),
[0082] Primer C2: 5'-GGAAGCGGATAGTCATAGTAGA-3' (SEQ ID NO: 2);
[0083] Primer M1: 5'-CCCATTCCCTCTTCTGCC-3' (SEQ ID NO: 3),
[0084] Primer M2: 5'-GCTGAGCGTAAACCGTC-3' (SEQ ID NO: 4).
[0085] The invention adopts the method of multiple immunofluorescence analysis to distinguish chicken infectious anemia virus and chicken Marek's disease virus, so the above primers are further modified to meet corresponding operation requirements. The 5' ends of primers C1 and M1 are connected with tag sequences, and the connected tag sequences are respectively:
[0086] The tag...
Embodiment 2
[0089] Example 2: The establishment of multiple fluorescent immunoassay kits for detecting CAV and MDV
[0090] The kit includes the following components:
[0091] (1) Multiplex fluorescent immunoassay primers designed in Example 1;
[0092] (2) Two kinds of fluorescently encoded microspheres containing anti-tag sequences that encode different fluorescent colors, and the anti-tag sequences can be complementary to the tag sequences in the primers for multiple fluorescent immunoassays; both types of microspheres can be purchased From luminex company, the numbers of fluorescently encoded microspheres corresponding to CAV and MDV are MTAG-A019 and MTAG-A065, respectively.
[0093] (3) Streptavidin-phycoerythrin complex.
Embodiment 3
[0094] Example 3 Establishment of Multiple Fluorescence Immunoassay Detection Method for CAV and MDV
[0095] (1) Construction of plasmids
[0096] The DNA of CAV and MDV pathogens were extracted with Tiangen’s automatic nucleic acid extractor, PCR amplification was performed with primer pairs C1 and C2, M1 and M2 respectively, and the amplified products were detected by agarose gel electrophoresis and purified by gel cutting. The purified cDNA was connected to the pMD-19T vector with a kit from TaKaRa Company, the connected product was transformed into DH5a competent cells, single clones were selected, colony PCR identification was carried out, and the colonies identified as positive bacteria were subjected to plasmid extraction. Send for sequencing.
[0097] (2) Plasmid PCR amplification
[0098] Use the primers described in Example 1 to carry out single-fold and double-fold PCR amplification of CAV and MDV primers, respectively.
[0099] Preparation of upstream primer mi...
PUM
Abstract
Description
Claims
Application Information
- R&D Engineer
- R&D Manager
- IP Professional
- Industry Leading Data Capabilities
- Powerful AI technology
- Patent DNA Extraction
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