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Feline coronavirus fluorescence EMA detection primer group, kit and detection method

A cat coronavirus and detection kit technology, applied in the biological field, can solve the problems of not being widely used, unable to use grassroots and on-site detection, high price, etc. Effect

Inactive Publication Date: 2018-09-21
SUZHOU CLICKGENE BIOTECH CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, both PCR technology and real-time fluorescent quantitative PCR technology need to be equipped with expensive PCR instruments or real-time fluorescent quantitative PCR instruments, so they cannot be used for grassroots and on-site detection, and have not been widely used at present

Method used

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  • Feline coronavirus fluorescence EMA detection primer group, kit and detection method
  • Feline coronavirus fluorescence EMA detection primer group, kit and detection method
  • Feline coronavirus fluorescence EMA detection primer group, kit and detection method

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0067] Embodiment 1, feline coronavirus fluorescent EMA detects

[0068] Feline coronavirus fluorescent EMA detection kit was used to detect feline coronavirus. The feline coronavirus fluorescent EMA detection kit comprises:

[0069] 1. 500 μl of complex solution;

[0070] Each 30ml reconstitution solution contains:

[0071]

[0072] 2. 48 pieces of freeze-dried enzyme tubes containing primers;

[0073] Each 4mL freeze-dried mixture contains the following components:

[0074]

[0075] Wherein, the sequences of primer pairs and probes are as follows:

[0076] Upstream primer: 5'-TCAAGGTCTGGTTCTCAGTCTAG-3';

[0077] Downstream primer: 5'-GTTTTTCTTCCAGGTGTGTTTGT-3';

[0078] Probe: 5'-TTTTTCTTCCAGGTGTGTTTG / i6-FAMdT / T / idSp /

[0079] / iBHQ1dT / TAGGTGT-3'C3Spacer.

[0080] 3. The negative control substance is the RNase-free water blank control.

[0081] 4. The positive control substance is feline pseudocoronavirus, and the concentration is 1 μg / mL.

[0082] Feline cor...

Embodiment 2

[0088] Embodiment 2: the detection of clinical sample:

[0089]Feline coronavirus fluorescent EMA detection kit was used to detect feline coronavirus clinical samples. The feline coronavirus fluorescent EMA detection kit comprises:

[0090] 1. 500 μl of complex solution;

[0091] Each 30ml reconstitution solution contains:

[0092]

[0093]

[0094] 2. 48 pieces of freeze-dried enzyme tubes containing primers;

[0095] Each 4mL freeze-dried mixture contains the following components:

[0096]

[0097] Wherein, the sequences of primer pairs and probes are as follows:

[0098] Upstream primer: 5'-TCAAGGTCTGGTTCTCAGTCTAG-3';

[0099] Downstream primer: 5'-GTTTTTCTTCCAGGTGTGTTTGT-3';

[0100] Probe: 5'-TTTTTCTTCCAGGTGTGTTTG / i6-FAMdT / T / idSp /

[0101] / iBHQ1dT / TAGGTGT-3'C3Spacer.

[0102] 3. The negative control substance is the RNase-free water blank control.

[0103] 4. The positive control substance is feline pseudocoronavirus, and the concentration is 1 μg / mL. ...

Embodiment 3

[0111] Embodiment 3, cross reaction detection:

[0112] Feline calicivirus, feline distemper virus, feline herpes virus, canine feline fever virus, canine parvovirus and rabies vaccine nucleic acid, with feline coronavirus positive samples as contrast, the specificity detection of the present invention was carried out, and the results showed that feline coronavirus Virus-positive samples are positive results, and other samples are negative results, which proves that the detection of feline coronavirus of the present invention is compatible with feline calicivirus, feline distemper virus, feline herpes virus, canine febrile fever virus, canine parvovirus and rabies vaccine nucleic acid. No cross-reactivity.

[0113] The invention uses normal temperature nucleic acid amplification technology, through multiple enzymatic reactions such as M-MLV reverse transcriptase, RNase inhibitor, DNA helicase, single-stranded DNA binding protein, DNA polymerase, etc. In 10-30 minutes, the RNA...

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Abstract

The invention relates to a feline coronavirus fluorescence EMA detection primer group, a kit and a detection method. The feline coronavirus fluorescence EMA detection primer group comprises an upstream primer, a downstream primer and a probe, the kit comprises a reconstitution fluid, a lyophilized enzyme tube containing primers, a positive quality control material and a negative quality control material. The detection method comprises the steps that the kit is used for performing a normal-temperature nucleic acid amplification technology, through the multiple enzymatic reactions of M-MLV reverse transcriptase, Rnase inhibitor, DNA helicase, single stranded DNA binding protein, DNA polymerase and the like, to-be-detected DNA is reversely transcribed to cDNA within 10-30 minutes under the constant temperature of 37-45 DEG C, the cDNA is amplified by millions of times, and rapid verification of the to-be-detected DNA can be achieved by cooperating with a fluorescence detection technology.The feline coronavirus fluorescence EMA detection primer group, the kit and the detection method have the advantages that the operation is simple, the time is short, the requirements for an instrument are low, and the feline coronavirus fluorescence EMA detection primer group, the kit and the detection method are suitable for the rapid diagnosis for pet infectious diseases.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to a feline coronavirus fluorescent EMA detection primer set, a kit and a detection method. Background technique [0002] Feline Coronavirus (FCoV): According to statistics, 25% to 40% of domestic cats are positive for coronavirus, while in large cattery or breeding cattery, the positive rate reaches 80% to 100%. Although there may be no clinical symptoms at all after infection, feline infectious peritonitis may occur with a very high mortality rate. This disease often occurs in young adult cats under the age of four, especially in cats housed in groups. Currently, there is no effective vaccine for the prevention of feline infectious peritonitis, nor is there an effective drug treatment. Therefore, accurate diagnosis is crucial for the management and control of FCoV. [0003] At present, there are a few detection methods for feline coronavirus (FCoV) nucleic acid, mainly including PCR...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/70C12N15/11C12R1/93
CPCC12Q1/6844C12Q1/701C12Q2521/107C12Q2521/513C12Q2521/101C12Q2522/101C12Q2563/107
Inventor 谭卓胡振新郜安国
Owner SUZHOU CLICKGENE BIOTECH CO LTD
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