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Primer and probe for detecting infectious bronchitis virus as well as detection method and application

A technology for bronchitis and chicken infectivity, applied in biochemical equipment and methods, microbiological determination/inspection, DNA/RNA fragments, etc., can solve immune failure, decline in egg production and egg quality, and economic losses in aquaculture And other issues

Pending Publication Date: 2021-06-22
HEBEI AGRICULTURAL UNIV.
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Chickens suffering from the disease will have symptoms of coughing, sneezing, and decreased egg production and egg quality, which will bring greater economic losses to the breeding industry
There are many serotypes of IBV, and the cross-protection between different serotypes is basically no or very weak, which can easily cause immune failure and bring great difficulties to the prevention and treatment of IBV
At the same time, IB is similar to a variety of poultry respiratory diseases such as Infectious laryngotracheitis (Infectious laryngotracheitis), avian influenza (Avian influenza), Newcastle Disease (Newcastle Disease) and other respiratory symptoms, which makes clinical diagnosis more difficult, delays the best treatment time, and leads to treatment fail
The traditional PCR method requires agarose gel electrophoresis to observe the results, which has disadvantages such as low sensitivity, long time-consuming, and the need to use carcinogenic EB dyes.

Method used

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  • Primer and probe for detecting infectious bronchitis virus as well as detection method and application
  • Primer and probe for detecting infectious bronchitis virus as well as detection method and application
  • Primer and probe for detecting infectious bronchitis virus as well as detection method and application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0026] 1. Materials and methods

[0027] 1.1. Extraction and preservation of DNA / RNA samples

[0028] Chicken infectious bronchitis virus IBV (AV1511) and chicken infectious laryngotracheitis virus ILTV (AV195) were provided by the China Veterinary Drug Administration; avian influenza virus AIV and chicken Newcastle disease virus NDV were stored in our laboratory after clinical testing.

[0029] Viral genome DNA / RNA extraction kit: purchased from Tiangen Biochemical Technology Co., Ltd.

[0030] 2. Design of primers and probes

[0031] According to the conserved and stable NA gene sequence (Sequence ID: HM159258.1) in the IBV genome, primers and probes were designed. The designed primer sequences are shown in Table 1, and were synthesized by Shanghai Sangon Bioengineering Co., Ltd.

[0032] Table 1 Primers and Probes

[0033]

[0034]

[0035] 1.3. Extraction of viral DNA / RNA

[0036] Extract the DNA / RNA of IBV, AIV, NDV and ILTV according to the instruction manual o...

Embodiment 2

[0067] A kind of primer and probe that detects chicken infectious bronchitis virus, the sequence of described primer is:

[0068] Upstream primer IBV-RAA-F: 5'-CAGGTAAAGGTGGAAGAAAACCAGTCCCAG-3' (SEQ ID NO: 1);

[0069] Downstream primer IBV-RAA-R: 5'-AGGTCCCCCATCTGAAAATCGTAGTGGATATTG-3' (SEQ ID NO: 2);

[0070] Probe IBV-T: 5'-TGATTCTCAGGATGGTATAATGTGGGTTGC / i6FAMdT / G / idSp / / iBHQ1dT / AAGGGTGCTGATGTA-3'C3 Spacer.

[0071] The kit for detecting chicken infectious bronchitis virus comprises the above primers and probes, fluorescent basic buffer solution, magnesium acetate solution, RAA dry powder reagent, chicken infectious bronchitis virus positive control gene and deionized water.

[0072] Utilize above-mentioned kit to detect the method for chicken infectious bronchitis virus: take the genome of 62 parts of clinical samples extracted in embodiment 1 respectively as template, take chicken infectious bronchitis virus positive control gene as positive control, take deionized water ...

Embodiment 3

[0080] A kind of primer and probe that detects chicken infectious bronchitis virus, the sequence of described primer is:

[0081] Upstream primer IBV-RAA-F: 5'-CAGGTAAAGGTGGAAGAAAACCAGTCCCAG-3' (SEQ ID NO: 1);

[0082] Downstream primer IBV-RAA-R: 5'-AGGTCCCCCATCTGAAAATCGTAGTGGATATTG-3' (SEQ ID NO: 2);

[0083] Probe IBV-T: 5'-TGATTCTCAGGATGGTATAATGTGGGTTGC / i6FAMdT / G / idSp / / iBHQ1dT / AAGGGTGCTGATGTA-3'C3 Spacer.

[0084] The kit for detecting chicken infectious bronchitis virus comprises the above primers and probes, fluorescent basic buffer solution, magnesium acetate solution, RAA dry powder reagent, chicken infectious bronchitis virus positive control gene and deionized water.

[0085] Utilize above-mentioned kit to detect the method for chicken infectious bronchitis virus: take the genome of 62 parts of clinical samples extracted in embodiment 1 respectively as template, take chicken infectious bronchitis virus positive control gene as positive control, take deionized water ...

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Abstract

The invention relates to the technical field of virus detection, and particularly discloses a primer and a probe for detecting an infectious bronchitis virus as well as a detection method and application. In the primers and the probe for detecting the infectious bronchitis virus, the sequence of an upstream primer is shown as SEQ ID NO: 1, the sequence of a downstream primer is shown as SEQ ID NO: 2, and the sequence of the probe is shown as SEQ ID NO: 3. By utilizing the combination of the primer and the probe, the real-time fluorescence RAA detection of the infectious bronchitis virus can be realized, and the detection method has the advantages of high specificity, good sensitivity, accuracy and high efficiency.

Description

technical field [0001] The invention relates to the technical field of virus detection, in particular to a primer, a probe, a detection method and an application for detecting chicken infectious bronchitis virus. Background technique [0002] Infectious bronchitis (IB) is a respiratory disease of chickens caused by infectious bronchitis virus (IBV), and is a common contact and acute respiratory disease. Chickens suffering from the disease will suffer from symptoms of coughing, sneezing, decreased egg production and egg quality, which will bring greater economic losses to the breeding industry. There are many serotypes of IBV, and the cross-protection between different serotypes is basically no or very weak, which may easily cause immune failure and bring great difficulties to the prevention and treatment of IB. At the same time, IB is similar to a variety of poultry respiratory diseases such as Infectious laryngotracheitis (Infectious laryngotracheitis), avian influenza (Av...

Claims

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Application Information

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IPC IPC(8): C12Q1/70C12Q1/6844C12N15/11
CPCC12Q1/701C12Q1/6844C12Q2521/507C12Q2522/101
Inventor 张铁王文静王春光李馨月石建存李希明曹立辉
Owner HEBEI AGRICULTURAL UNIV.
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