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A kind of compound pcr detection method of rabbit skin fungus

A dermatophyte and detection method technology, applied in the field of biomedicine, can solve problems such as limited application and unsuitability for large-scale sample detection, and achieves the effect of small sample volume, suitable for large-scale sample detection, and reduction of PCR times

Inactive Publication Date: 2016-05-11
HEBEI UNIV OF ENG
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Problems solved by technology

Although this method is specific and sensitive, it requires the determination and analysis of gene sequences, which limits the application of this method, and only one fungus can be identified in one reaction, which is not suitable for large-scale sample detection

Method used

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  • A kind of compound pcr detection method of rabbit skin fungus
  • A kind of compound pcr detection method of rabbit skin fungus
  • A kind of compound pcr detection method of rabbit skin fungus

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Embodiment Construction

[0023] 1. DNA extraction

[0024] (1) Strain DNA extraction: Inoculate the bacterial strains of Trichophyton mentagrophytes, Microsporum canis and Epidermophyton flocculus in sand castle weak SDA culture medium, 27 ℃, shock culture for 8 days, harvest; Add 500 μL lysate (400 mM Tris-HCl (pH8.0), 60 mM EDTA (pH8.0), 150 mM NaCl, 1% sodium dodecyl sulfate); let stand at room temperature for 10 min, add 150 μL potassium acetate (pH4.8), vortex, Centrifuge at 12,000 g for 1 min; put the supernatant into a new test tube, add an equal volume of isopropanol; wash the DNA cluster with 70% ethanol, dry it, dissolve it in 50 μL of TE (10 mM Tris, 1 mM EDTA), and use 2 μL of DNA for subsequent PCR. The strain DNA template was used as a positive control group.

[0025] (2) DNA extraction of disease materials: Three groups of disease materials F1, F2, and F3 suspected of being infected with Trichophyton mentagrophytes, Microsporum canis, and Epidermophyton flocculus were subjected to the ...

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Abstract

The invention discloses a multiplex PCR detection method of rabbit skin fungi, which is characterized in that the method comprises the following steps: (1) extracting DNA from a diseased material suspected to be infected by trichophyton mentagrophytes, microsporum canis, and epidermophyton floccosum through a pyrolysis method; (2) synthesizing three pairs of specific primers of Pa and Pb, Pc and Pd, and Pe and Pf; (3) performing PCR amplification by using the diseased material DNA as a template and the mixture of the three pairs of primers of Pa-Pf as a primer; (4) detecting the PCR product by electrophoresis. The beneficial effects of the method of the invention are that a multiplex PCR detection method of rabbit skin fungi is provided, and three skin fungi can be identified through performing PCR once. When compared with conventional methods such as direct microscopic observation, pathogen isolation and culture, and the like, the method is strong in specificity, high in sensitivity, and time-saving; when compared with other molecular biological methods, the method reduces the times for PCR, needs no sequencing, has the characteristics of simplicity, rapidness, and economy, and is quite suitable for large-scale sample detection.

Description

technical field [0001] The invention relates to a detection method for rabbit skin fungus infection, in particular to a composite PCR detection method for rabbit skin fungus, which belongs to the technical field of biomedicine. Background technique [0002] Dermatophyte (Dermatophyte) is the pathogenic bacteria of skin mycosis. Dermatomycosis is a highly contagious zoonotic disease, which not only endangers the rabbit industry and causes heavy economic losses, but also infects humans and affects human health. The pathogens of dermatophytosis involve three genera, namely Trichophyton, Microsporum and Epidermophyton. The epidemiological survey of rabbit skin fungi showed that Trichophyton mentagrophytes, Microsporum canis and Epidermophyton flocculus were the most common skin fungi. Different skin fungi have different infection sites, cause different lesions, and take different medications and prevention and control measures. Therefore, it is very important to identify the t...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/68C12Q1/04C12R1/645
CPCC12Q1/686C12Q2537/143
Inventor 刘彦威刘娜刘利强杜鹃石玉祥张永英王慧真
Owner HEBEI UNIV OF ENG
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