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Dual RT-PCR (reverse transcription-polymerase chain reaction) detection for CDV (canine distemper virus) and CPIV (canine parainfluenza virus) and special primer for detection

An RT-PCR and primer pair technology, applied in the biological field, can solve problems such as poor sensitivity, and achieve the effect of simple method and saving detection cost.

Inactive Publication Date: 2015-03-18
山东大学生命科学学院 +2
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  • Abstract
  • Description
  • Claims
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Problems solved by technology

However, the minimum nucleic acid detection amount of these three pairs of primers for CDV, CPV, and CPIV is 1ng / μL; their sensitivity is poor

Method used

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  • Dual RT-PCR (reverse transcription-polymerase chain reaction) detection for CDV (canine distemper virus) and CPIV (canine parainfluenza virus) and special primer for detection
  • Dual RT-PCR (reverse transcription-polymerase chain reaction) detection for CDV (canine distemper virus) and CPIV (canine parainfluenza virus) and special primer for detection
  • Dual RT-PCR (reverse transcription-polymerase chain reaction) detection for CDV (canine distemper virus) and CPIV (canine parainfluenza virus) and special primer for detection

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Experimental program
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Effect test

Embodiment 1

[0057] Construction of dual RT-PCR detection method for canine distemper virus (CDV) and canine parainfluenza virus (CPIV)

[0058] 1. Primer design

[0059] Detected CDV (Genbank No. AF112189), CPIV (Genbank No. EF543648) in GenBank

[0060] part of the known sequence. Homology analysis was carried out on each gene region of the virus, and the H gene of CDV and the NP gene of CPIV were respectively selected as the target sequences amplified by each virus. Primer Premier 5 was used to design primers for the above conserved regions, and DNAstar was used for primer-dimer analysis of the designed two viral primers to avoid the formation of stable primer-dimers between primers. Select the following 2 pairs of primers whose amplified sequences are CDV (593bp) and CPIV (1530bp), and analyze the homology or complementarity of the amplified sequences of each pair of primers. Avoid high homology or complementarity between them, so as to determine the double RT-PCR primers of the two...

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Abstract

The invention discloses dual RT-PCR (reverse transcription-polymerase chain reaction) detection for a CDV (canine distemper virus) and a CPIV (canine parainfluenza virus) and a special primer for the detection. The invention provides the special primer for virus detection. The primer comprises a primer pair A and a primer pair B, wherein the primer pair A comprises a primer 1 and a primer 2; the primer pair B comprises a primer 3 and a primer 4; and nucleotide sequences of the primer 1, primer 2, primer 3 and the primer 4 are sequence 1, sequence 2, sequence 3 and sequence 4 respectively in a sequence table. Experiments prove that the special primer for dual RT-PCR detection for the CDV and the CPIV has good sensibility, can at least detect the RNA (ribonucleic acid) content to be about 1*10<-3>ng / mu L, the sensitivity is far higher than that of 1 ng / mu L. A dual RT-PCR detection method is simple, convenient, fast, economical and efficient, and has good application prospect.

Description

technical field [0001] The invention relates to double RT-PCR detection of canine distemper virus and canine parainfluenza virus and special primers thereof, belonging to the field of biotechnology. Background technique [0002] The RT-PCR detection method of canine distemper virus (CDV) and canine parainfluenza virus (CPIV) is a common method for rapid detection of these two pathogens at present, and is widely used in laboratory pathogen diagnosis. Compared with the method of virus isolation, The RT-PCR method is faster and easier to diagnose, and the virus isolation takes too long, which is not conducive to rapid clinical diagnosis. Therefore, the current laboratory diagnosis of viral diseases mainly uses the PCR / RT-PCR method. At present, some RT-PCR kits for CDV have been commercialized and put into clinical application. However, there are no commercialized kits for RT-PCR methods for single detection of CPIV or one-time detection of CDV and CPIV. [0003] At present,...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/70C12Q1/68C12N15/11C12R1/93
CPCC12Q1/686C12Q1/70C12Q2537/143C12Q2531/113
Inventor 王贵升田夫林王金宝尹斐斐徐鸿李玉杰孙圣福蔺晓月宗晓晨
Owner 山东大学生命科学学院
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