Method for rapidly detecting feline calicivirus as well as primer and kit for detection
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A feline calicivirus and kit technology, which is applied in the detection field of feline calicivirus, can solve the problems that monoclonal antibodies cannot be universal, time-consuming, and cannot meet the detection requirements, etc.
Active Publication Date: 2021-06-08
HARBIN VETERINARY RES INST CHINESE ACADEMY OF AGRI SCI
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[0003] Currently, FCV detection methods include serological detection and virological detection. Serological detection includes serum neutralization test, agar diffusion test, immunofluorescence or enzyme-linked immunoassay, etc. These methods are relatively time-consuming
Virus detection methods, including electron microscopy, RT-PCR, fluorescent quantitative RT-PCR and antigen-capturing colloidal gold detection methods, etc.; however, because fe
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specific Embodiment approach 1
[0026] Embodiment 1: The sequence of the upstream primer ERA-F of the primers and probes for rapid detection of feline calicivirus in this embodiment is: 5'-GTCTCAAACTCTGAGCTTCGTGCTTAAAAC-3';
[0027] The sequence of the downstream primer ERA-R for rapid detection of feline calicivirus is: 5'-Biotin-ACTTAGGACATACATCATAACTAGCACAAG-3';
[0028] The sequence of the probe Probe for rapid detection of feline calicivirus is: 5'-FAM-AAACTCTGAGCTTCGTGCTTAAAACTCACA[THF]TGTCCGTAAGGACTT-C3Spacer-3'.
[0029] The present invention couples biotin (Biotin) to the 5' end of the downstream primer; couples a FAM group to the 5' end of the probe, replaces a base with tetrahydrofuran (THF) in the middle of the probe sequence, and The 3' end of the needle is treated with a C3-Spacer group.
specific Embodiment approach 2
[0030] Specific embodiment two: the RT-ERA nucleic acid test strip amplification kit for the rapid detection of feline calicivirus in this embodiment includes the primers and probes described in specific embodiment one, positive control samples, isothermal nucleic acid amplification kit and test strips;
[0031] Wherein, the positive control sample contains the RNA sequence of feline calicivirus ORF1 region;
[0032] The isothermal nucleic acid amplification kit includes recombinant enzyme lyophilized enzyme powder, dissolving agent, activator, RNase inhibitor and RNase-free H 2 O.
[0033] In the present invention, RNase-free H 2 O was used as a negative control sample.
specific Embodiment approach 3
[0034] Embodiment 3: The difference between this embodiment and Embodiment 2 is that the test strip contains anti-biotin antibody, anti-FAM antibody, anti-mouse antibody and gold-labeled particles. Other steps and parameters are the same as those in Embodiment 2.
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Abstract
The invention discloses a method for rapidly detecting feline calicivirus as well as a primer and a kit for detection, and relates to a method for detecting the feline calicivirus as well as a primer and a kit for detection. According to the invention, an upstream primer ERA-F, a downstream primer ERA-R and a probe Probe are protected. The kit comprises the primer, the probe, a positive control sample, an isothermal nucleic acid amplification kit and a test strip. The detection method comprises the following steps: 1, preparing a detection solution; and 2, detecting a reaction product in the step 1 by using a test strip to obtain a detection result. According to the invention, the feline calicivirus is rapidly detected through an RT-ERA technology by adopting a primer and probe combination mode. The primer and probe combination disclosed by the invention is designed aiming at a feline calicivirus ORF1 conserved region, has the advantages of high sensitivity, strong specificity, good repeatability and the like, and has no cross reaction with feline herpes virus, feline parvovirus and feline infectious peritonitis virus.
Description
technical field [0001] The invention relates to a detection method for feline calicivirus, primers and a kit for detection. Background technique [0002] Feline Calicivirus (FCV) is a highly prevalent pathogen that causes respiratory diseases in cats. It often infects kittens under one year old. Cats infected with FCV usually cause clinical manifestations such as oral ulcers, fever, sneezing, and rhinitis. , conjunctivitis, atypical symptoms can cause claudication, diarrhea or fatal systemic disease (Virulent SystemicDisease, VSD). After recovery, cats with clinical disease and asymptomatic carriers can still shed the virus for a long time, which is conducive to the spread of FCV in cats and is an important source of infection. [0003] Currently, FCV detection methods include serological detection and virological detection. Serological detection includes serum neutralization test, agar diffusion test, immunofluorescence or enzyme-linked immunoassay, etc. These methods are ...
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