Multiplex PCR detection primer set and kit used for mycoplasma gallisepticum, mycoplasma gallisepticum and parabacterium bacillus and application of kit

A technology for Mycoplasma synovialis and Mycoplasma gallisepticum, which is applied in the field of molecular detection, can solve the problems of the difficulty in diagnosis of the three diseases, the inability to realize the simultaneous detection of Mycoplasma gallisepticum, Mycoplasma gallisepticum and Mycobacterium gallisepticum, and achieve high sensitivity good, good specific effect

Active Publication Date: 2019-11-15
INNER MONGOLIA AUTONOMOUS REGION ACAD OF AGRI & ANIMAL HUSBANDRY SCI
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] Chickens infected with MG and APG showed respiratory symptoms, and some chickens infected with MS also showed respiratory symptoms, which brought great difficulties to the diagnosis of the three diseases
[0006] At present, the existing technology can only realize the single or double detection of Mycoplasma gallisepticum, Mycoplasma gallisepticum and Avibacterium paragallinarum, but cannot realize the simultaneous detection of Mycoplasma gallisepticum, Mycoplasma gallisepticum and Avibacterium paragallinarum

Method used

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  • Multiplex PCR detection primer set and kit used for mycoplasma gallisepticum, mycoplasma gallisepticum and parabacterium bacillus and application of kit
  • Multiplex PCR detection primer set and kit used for mycoplasma gallisepticum, mycoplasma gallisepticum and parabacterium bacillus and application of kit
  • Multiplex PCR detection primer set and kit used for mycoplasma gallisepticum, mycoplasma gallisepticum and parabacterium bacillus and application of kit

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0052] Example 1 The specificity verification of the multiplex PCR detection of Mycoplasma gallisepticum, Mycoplasma gallisanum and Avibacterium paragallinarum for non-diagnostic purposes

[0053] Double-distilled water, Mycoplasma gallisepticum, Mycoplasma gallisepticum and Avibacterium paragallinarum multiplex (MG\MS\APG) mixed DNA, Escherichia coli DNA, Salmonella pullorum DNA, Pasteurella multocida DNA, Staphylococcus aureus DNA was used as a template, and the following reaction system was used for multiplex PCR amplification.

[0054] The reaction system is based on 25 μL:

[0055] A: MS-F / R each 0.25μL, MG-F / R each 0.5μL, APG-F / R each 0.5μL, Mix 12.5μL, H 2 O 10 μL;

[0056] B: MS-F / R each 0.25 μL, MG-F / R each 0.5 μL, APG-F / R each 0.5 μL, Mix 12.5 μL, H 2 O 7 μL, DNA 3 μL (MG, MS, APG each 1 μL);

[0057] C: MS-F / R each 0.25 μL, MG-F / R each 0.5 μL, APG-F / R each 0.5 μL, Mix 12.5 μL, H 2 O 9 μL, MGDNA 1 μL;

[0058] D: MS-F / R each 0.25 μL, MG-F / R each 0.5 μL, APG-F / R...

Embodiment 2

[0067] Example 2 Sensitivity Verification of Multiplex PCR Detection of Mycoplasma gallisepticum, Mycoplasma gallisanum and Avibacterium paragallinarum for non-diagnostic purposes

[0068] Adjust the initial concentration of MG, MS, and APG genomic DNA to 5×10 2 ng / μL, take 100 μL MG, MS and APG genomic DNA in equal volumes and mix them, then perform 10-fold serial dilution (10 2 ~10 -7 ), then take 1 μL of mixed DNA for each dilution as a template, perform multiplex PCR amplification according to the following reaction system, calculate the minimum detection amount of MG, MS, and APG by this multiplex PCR method, and evaluate its sensitivity.

[0069] Reaction system: 25 μL includes: 0.25 μL each of MS-F and MS-R, 0.5 μL each of MG-F and MG-R, 0.5 μL each of APG-F and APG-R, 12.5 μL of PCR Mix, H 2 07 μL, 1 μL each of the genomic DNA of Mycoplasma gallisepticum, Mycoplasma gallisepticum and Avibacterium paragallinarum.

[0070] For experimental results, see figure 2 , ...

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Abstract

The invention provides a multiplex PCR detection primer set and kit used for mycoplasma gallisepticum, mycoplasma gallisepticum and parabacterium bacillus and application of the kit, and belongs to the technical field of molecular detection. The primer set comprises MS-F, MS-R, MG-F, MG-R, APG-F and APG-R; the nucleotide sequences of MG-F, MG-R, MS-F, MS-R, APG-F and APG-R are shown in SEQ ID NO:1-SEQ ID NO:6 respectively. It is verified that the primer set has high specificity and sensitivity.

Description

technical field [0001] The invention relates to the technical field of molecular detection, in particular to a primer set, a kit and an application for multiplex PCR detection of Mycoplasma gallisepticum, Mycoplasma gallisynovus and Avibacterium paragallinarum. Background technique [0002] Mycoplasma gallisepticum (Mycoplasma Gallisepticum, MG), also known as Mycoplasma gallisepticum or Mycoplasma gallisepticum, was first isolated from infected chickens by Delaplane et al. in 1943. Mycoplasma gallisepticum disease caused by this pathogen is distributed worldwide , the infection rate of the disease in American flocks can reach 73%, and the seropositive rate of the disease in Malaysian flocks is 26%. The disease is widespread in our country, and the data show that the positive rate of Mycoplasma gallisepticum infection in my country has reached 50% to 80%, and it has become one of the important diseases threatening the poultry industry in our country. Different strains of My...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/689C12Q1/04C12N15/11C12R1/35C12R1/01
CPCC12Q1/689C12Q2600/16
Inventor 杨斌赵世华钱琳娜萨娜宋爱军张帆陈伟卢岩王娜戴伶俐张月梅宋越刘威达来宝力格
Owner INNER MONGOLIA AUTONOMOUS REGION ACAD OF AGRI & ANIMAL HUSBANDRY SCI
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