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Kit for detecting infectious laryngotracheitis virus, newcastle disease virus and infectious bronchitis virus

A technology for chicken Newcastle disease virus and chicken infectivity, applied in the direction of microbe-based methods, biochemical equipment and methods, microbiological determination/testing, etc., can solve the problem of inability to explain specific and sensitive simultaneous detection, lack of chip specificity and sensitivity To achieve good application prospects, rapid detection, and strong specificity

Active Publication Date: 2014-12-24
SICHUAN AGRI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0006] Patent applications with publication numbers 1616679 and 1616678 respectively disclosed gene chips for detecting 10 or 9 viruses including chicken Newcastle disease virus, chicken infectious bronchitis virus and chicken infectious laryngotracheitis virus, but there is no specificity of the chips in the documents. It cannot be proved that it can specifically and sensitively detect Newcastle disease virus, chicken infectious bronchitis virus and chicken infectious laryngotracheitis virus at the same time

Method used

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  • Kit for detecting infectious laryngotracheitis virus, newcastle disease virus and infectious bronchitis virus
  • Kit for detecting infectious laryngotracheitis virus, newcastle disease virus and infectious bronchitis virus
  • Kit for detecting infectious laryngotracheitis virus, newcastle disease virus and infectious bronchitis virus

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0036] The preparation of embodiment 1 kit of the present invention

[0037] 1. Materials and instruments

[0038] The same experimental materials and instruments as mentioned above.

[0039] 2. Experimental method

[0040] 2.1 Preparation of PCR primers and detection probes

[0041] (1) Design of pathogen-specific probes: by comparing and analyzing the nucleic acid sequences of chicken Newcastle disease virus, chicken infectious bronchitis virus, and chicken infectious laryngotracheitis virus included in GenBnak, a conserved region sequence was selected: NDV-F / HN, IBV-IBM / IBN, ILTV-TK / gB. Design and detect multiple pairs of probes for conserved sequences, and select the probe sequences with strong specificity.

[0042] (2) Design of specific primers for probe sequences: specific primers were designed for the above conserved probe sequences using bioinformatics software DNAman, Primer5.0, etc. Specific primers were synthesized by Shanghai Bioengineering Company.

[0043...

Embodiment 2

[0209] Embodiment 2 detection method of the present invention

[0210] 1. Nucleic acid extraction

[0211] Extraction of viral RNA: The kit method is used to extract viral RNA, and the test uses a small amount of virus / liquid sample RNA extraction kit. Extract RNA according to the instructions of the kit, the method is as follows:

[0212] a) Take 300 μL of the sample to be tested and place it in a centrifuge tube, add 500 μL of RV solution, shake vigorously for 2 minutes, and then let stand at room temperature for 5 minutes.

[0213] b) Add 750 μL of isopropanol and shake gently.

[0214] c) Pipette 800 μL into the adsorption column and centrifuge at 12000 rpm for 30 s at 4°C.

[0215] d) Discard the liquid in the collection tube, transfer the remaining lysate into an adsorption column, and centrifuge at 12000r for 30s at 4°C.

[0216] e) Add 500 μL RP solution after discarding the collected solution, and centrifuge at 12000 rpm for 30 seconds at 4°C to remove protein.

...

Embodiment 3

[0264] Embodiment 3 specificity test

[0265] 1. Test method

[0266] With the method of Example 2, five kinds of pathogens of NDV, IBV, ILTV, IBDV and AIV were detected to detect their specificity.

[0267] 2. Results

[0268] Experimental results such as Figure 4 As shown, the kit of the present invention can effectively detect NDV, IBV, and ILTV of the present invention, but will not detect other viruses, such as IBDV, AIV, indicating that the method of the present invention has strong specificity and will not amplify other viruses.

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Abstract

The invention discloses a kit for detecting infectious laryngotracheitis virus, newcastle disease virus and infectious bronchitis virus. The kit disclosed by the invention is capable of correctly and effectively detecting the infectious laryngotracheitis virus, the newcastle disease virus and the infectious bronchitis virus, and is strong in specificity, high in sensitivity, short in time, rapid in detection and good in application prospect.

Description

technical field [0001] The invention relates to a kit for detecting chicken infectious laryngotracheitis virus, chicken Newcastle disease virus and chicken infectious bronchitis virus. Background technique [0002] With the continuous improvement of the degree of intensification of my country's poultry industry, the incidence of various diseases is also increasing year by year. The overall level of poultry disease prevention and control in my country is not high, and the occurrence and harm of poultry diseases are still very serious. According to incomplete statistics, there are more than 80 kinds of diseases that endanger the production of poultry industry in my country, among which infectious diseases account for about 75% of the total number of poultry diseases. , the hazard is serious. At the same time, the occurrence of poultry diseases also has new characteristics, such as the continuous emergence of new infectious diseases; the increase in the types of new poultry dis...

Claims

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Application Information

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IPC IPC(8): C12Q1/70C12Q1/68C12N15/11C12R1/93
CPCC12Q1/701
Inventor 文心田曹三杰黄小波黄宁文翼平伍锐胡中凯赵松尹人杰邓静
Owner SICHUAN AGRI UNIV
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