Haemophilus paragallinarum fluorogenic quantitative PCR detection method

A haemophilus gallinarum and fluorescence quantitative technology, which is applied in the field of fluorescence quantitative PCR detection of Haemophilus paragallinarum, can solve the problems of poor growth of broilers, complicated separation and cultivation and animal experiment operations, and decreased egg production rate of laying hens.

Inactive Publication Date: 2015-10-28
SHANDONG BINZHOU ANIMAL SCI & VETERINARY MEDICINE ACADEMY
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Problems solved by technology

[0002] Haemophilus paragallinarum (H.gallinarum), also known as Avibacterium paragallinarum Apg, is the pathogen that causes infectious coryza IC in chickens. It mainly causes acute upper respiratory tract symptoms in chickens, mainly manifested as nasal passages. 1. A large amount of serous or mucous secretions appear in the sinuses, facial edema and conjunctivitis, which can lead to a decrease in the laying rate of laying hens (10%-40%), poor growth of broilers, and increased elimination rate. caused huge economic losses
[0003] At present, the laboratory diagnosis methods of Haemophilus paragallinarum mainly include the isolatio

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Embodiment Construction

[0016] The invention provides a simple, fast, sensitive and specific fluorescent quantitative PCR detection method for Haemophilus paragallinarum.

[0017] The dye used in the fluorescent quantitative PCR detection method for Haemophilus paragallinarum of the present invention is SYBR Green I fluorescent dye.

[0018] The primers used in the fluorescent quantitative PCR detection method for Haemophilus paragallinarum of the present invention are designed and synthesized using Premier 5.0 software according to the hagA gene sequence of Haemophilus paragallinarum published in GenBank (GenBank accession number: KF280261.1). The specific upstream primer P1 sequence is: 5'-GTAGCGAAGCAGCCAACTTA-3'; the specific downstream primer P2 sequence is: 5'-TGGGTTCGCTTCA CCATAAC-3', and the expected amplified product size is 123bp.

[0019] In one embodiment, the fluorescent quantitative PCR detection method of the present invention comprises the following steps:

[0020] (a) extracting the ...

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Abstract

The invention discloses a haemophilus paragallinarum fluorogenic quantitative PCR detection method. According to the haemophilus paragallinarum fluorogenic quantitative PCR detection method, a hagA gene with the high degree of specificity and conservative property is selected as an amplification target gene, SYBR Green I fluorochrome is taken as a detection target, through the continuous optimization of fluorogenic quantitative PCR reaction conditions, the preparation of a standard substance and the establishment of a standard curve, the haemophilus paragallinarum fluorogenic quantitative PCR detection method of the SYBR Green I fluorochrome jogged with DNA, and the DNA of a haemophilus paragallinarum hagA genetic template can be quantified accurately. The haemophilus paragallinarum fluorogenic quantitative PCR detection method has the advantages of being strong in specificity, high in sensibility, simple in operation, rapid in detection, accurate in quantification and the like, the haemophilus paragallinarum fluorogenic quantitative PCR detection method can be used for clinic rapid diagnosis of haemophilus paragallinarum and the quantitative analysis of the degree of infection by the haemophilus paragallinarum, and the haemophilus paragallinarum fluorogenic quantitative PCR detection method is suitable for popularization and application.

Description

technical field [0001] The invention relates to a method for detecting Haemophilus paragallinarum. Specifically, the present invention relates to a fluorescent quantitative PCR detection method for detecting Haemophilus paragallinarum. Background technique [0002] Haemophilus paragallinarum (H.gallinarum), also known as Avibacterium paragallinarum Apg, is the pathogen that causes infectious coryza IC in chickens. It mainly causes acute upper respiratory tract symptoms in chickens, mainly manifested as nasal passages. 1. A large amount of serous or mucous secretions appear in the sinuses, facial edema and conjunctivitis, which can lead to a decrease in the laying rate of laying hens (10%-40%), poor growth of broilers, and increased elimination rate. caused huge economic losses. [0003] At present, the laboratory diagnosis methods of Haemophilus paragallinarum mainly include the isolation and identification of bacteria, animal regression test and conventional PCR method. ...

Claims

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Application Information

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IPC IPC(8): C12Q1/68C12Q1/06
CPCC12Q1/6851C12Q2563/107C12Q2545/114
Inventor 苗立中王艳李峰孙翠平沈志强
Owner SHANDONG BINZHOU ANIMAL SCI & VETERINARY MEDICINE ACADEMY
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