RDA method and kit for rapidly detecting feline parvovirus (FPV)

A feline parvovirus and kit technology, which is applied in the field of molecular biology, can solve problems such as cost and application range limitations, and achieve the effects of low detection cost, broad application prospects, and improved yield and stability

Pending Publication Date: 2021-02-02
GUANGZHOU PLUSLIFE BIOTECH CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] The detection of molecular biology is mostly based on PCR. The detection needs to rely on PCR machines or expensive real-time quantitative PCR machines, and

Method used

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  • RDA method and kit for rapidly detecting feline parvovirus (FPV)
  • RDA method and kit for rapidly detecting feline parvovirus (FPV)
  • RDA method and kit for rapidly detecting feline parvovirus (FPV)

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0082] Embodiment 1 A kind of feline parvovirus (FPV) RDA fluorescence detection kit

[0083] (1) Acquisition of recombinant enzyme KX and KY proteins

[0084] The reported recombinant enzyme UvsX has poor stability and is difficult to mass produce and store for a long time. In order to solve this problem, the research and development team used bioinformatics methods to analyze and simulate large quantities of protein structures, and finally found a new recombinant enzyme Enzyme KX and its accessory protein KY.

[0085] In this example, the R&D team extracted the key functional site information in the recombinase structure, such as DNA binding site, ATP hydrolysis site, etc., and mapped it to the three-dimensional structure of the protein to obtain secondary structure information and tertiary structure information , by integrating the functional residues of the primary structure sequence, secondary structure features and tertiary structure spatial distance, a data model for p...

Embodiment 2

[0135] Example 2 RDA fluorescence method detection reagent sensitivity test

[0136] The positive control is the pUC57-VP2 plasmid containing the VP2 gene of feline parvovirus (FPV), and the negative control is the empty vector pUC57 plasmid.

[0137] The specific operation is as follows:

[0138] Step 1: Dilute the positive control plasmid to 10^3c, and then dilute to 10^2c, 10^1c, and 10^0c by 10-fold serial dilution.

[0139] Step two, sample processing. Take 5 μL of each concentration of the plasmid in step 1 and put it in an EP tube, and take 5 μL of the negative control in another EP tube, add 20 μL BufferA respectively, shake and mix, and let stand at room temperature for 10-15 minutes;

[0140] Step 3, system preparation and testing. Add 25 μL of BufferB to each tube, shake and mix, add 50 μL of the mixed solution to the RDA constant temperature amplification reaction module, cover the tube cap, shake and centrifuge, and detect immediately; the reaction program is: ...

Embodiment 3

[0152] Example 3 RDA fluorescence method detection reagent specificity test

[0153] 3 cases of feline parvovirus (FPV), 1 case of feline chlamydia (CP), 1 case of feline calicivirus (FCV), and 1 case of feline herpes virus (FHV) will be clinically collected, and a total of 6 cases of 4 types have been verified by fluorescent quantitative PCR Test the specificity of the kit for the corresponding pathogen-positive samples.

[0154] The specific operation is as follows:

[0155] Step 1. Sample processing. Take 5 μL of each of the above 6 positive samples in EP tubes, and at the same time take 5 μL of the positive control and negative control of the kit into new EP tubes, add 20 μL BufferA respectively, shake and mix, and let stand at room temperature for 10-15 minutes;

[0156] Step 3, system preparation and testing. Add 25 μL of BufferB to each tube, shake and mix, add 50 μL of the mixed solution to the RDA constant temperature amplification reaction module, cover the tube c...

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Abstract

The invention discloses an RDA method and a kit for rapidly detecting feline panleukopenia virus (FPV). The kit for rapidly detecting the FPV comprises a specific primer pair and an RDA fluorescent label probe so as to realize safe, specific, sensitive, simple and convenient detection of the FPV, thereby making up the defects of the existing traditional detection technology. The kit provided by the invention can save the step of nucleic acid extraction, and is capable of detecting the FPV within 20 min at a constant temperature; and moreover, the kit has a specificity of 100%. Compared with acommon PCR method, the RDA fluorescence method has the advantages of being capable of carrying out reaction at a constant temperature, free of requirements for temperature change and complex instruments, and short in reaction time. Therefore, the kit is suitable for on-site rapid detection. The method and the kit thereof disclosed by the invention have the characteristics of being simple and rapidin operation, excellent in specificity, high in sensitivity, low in cost and the like; and thus, an effective technical means is provided for on-site rapid detection and screening of the FPV. The method and the kit have a wide application prospect.

Description

technical field [0001] The invention belongs to the technical field of molecular biology. More specifically, it relates to a primer pair, a probe and a related kit for detecting feline parvovirus (Feline panleukopenia virus, FPV) nucleic acid based on RDA fluorescence detection technology. Background technique [0002] Feline parvovirus (Feline panleukopenia virus, FPV) is also known as feline panleukopenia virus, feline infectious enteritis virus or feline distemper virus. FPV belongs to Parvoviridae, Parvoviridae, member of Bovine Canine Parvovirus, and is a single-strand non-enveloped NDA virus. FPV mainly infects a variety of animals such as cats and mustelidae, and is currently the virus with the widest infection range and the strongest pathogenicity among the carnivorous parvoviruses. Feline panleukopenia is an acute, highly contagious disease. The main clinical features of cats infected with FPV are high fever, vomiting, severe leukopenia and enteritis. The infect...

Claims

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Application Information

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IPC IPC(8): C12Q1/70C12Q1/6844C12N15/11C12R1/93
CPCC12Q1/701C12Q1/6844C12Q2521/507C12Q2563/107C12Q2531/119C12Q2522/101
Inventor 文荻琛刘华勇罗宝花陈翀
Owner GUANGZHOU PLUSLIFE BIOTECH CO LTD
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