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PCR (polymerase chain reaction) detection kit for hacmophilus parasuis

A technology of Haemophilus suis and detection kit, which is applied in the field of Haemophilus parasuis PCR detection kit

Inactive Publication Date: 2013-03-20
GUIZHOU INST OF ANIMAL HUSBANDRY & VETERINARY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Now the market lacks a kit that can detect Haemophilus parasuis in clinical samples simply, quickly, sensitively and accurately

Method used

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  • PCR (polymerase chain reaction) detection kit for hacmophilus parasuis
  • PCR (polymerase chain reaction) detection kit for hacmophilus parasuis
  • PCR (polymerase chain reaction) detection kit for hacmophilus parasuis

Examples

Experimental program
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Effect test

Embodiment 1

[0056] Embodiment 1 of the present invention: Haemophilus parasuis PCR detection kit, it comprises 10 μ L of PCR premix buffer, PCR premix buffer consists of MgCl of 3 mmol / ul 2 , 500pmol / ul dNTP 2.0 μL, 25 mmol / μL Tris-HCl and 0.2 μL Taq enzyme, the pH value is pH8.3; 25 μL ultrapure water; 2 μL Marker DL2000; 1 μL concentration of 12 pmol / μL Primers and 1 μL lower primer with a concentration of 12 pmol / μL, the sequence number of the upstream primer is LPF 5′-GTGATGAGGAAGGGTGGTGT-3′, the sequence number of the downstream primer is LPR 5′-GGCTTCGTCACCCCTCTGT-3′; 25 μL of protease with a concentration of 20 mg / ml K, 30 μL SDS with a mass concentration of 10%, 500 μL TE buffer solution, TE buffer solution including a mixed solution composed of Tris-HCL10mM and EDTA1mM, the pH value of the mixed solution is 7.3; use 2μL non-Haemophilus parasuis DNA fragment as Negative control; use 2 μL of the standard PCR product of Haemophilus parasuis as a positive control.

Embodiment 2

[0057] Embodiment 2 of the present invention: Haemophilus parasuis PCR detection kit, it comprises 8 μ L of PCR premix buffer, PCR premix buffer consists of MgCl of 3mmol / ul 2 , 500pmol / ul dNTP2.0μL, 25mmol / μL Tris-HCl and 0.2μL Taq enzyme composition, the pH value of which is pH8.3; 25μL ultrapure water; 3μL Marker DL2000; 0.75μL concentration of 12pmol / μL The upper primer and 0.75 μL lower primer with a concentration of 12 pmol / μL, the sequence number of the upstream primer is LPF 5′-GCTAACTCCGTGCCAGCAG-3′, the sequence number of the downstream primer is LPR 5′-GTCATCATGGCCCTTACGAG-3′; the concentration of 25 μL is 20 mg / ml Proteinase K, 20 μL SDS with a mass concentration of 10%, 500 μL TE buffer, TE buffer includes a mixed solution composed of Tris-HCL10mM and EDTA1mM, the pH value of the mixed solution is 7.0; use 2μL DNA of non-Hemophilus parasuis The fragment was used as a negative control; 2 μL of the standard PCR product of Haemophilus parasuis was used as a positive ...

Embodiment 3

[0058] Embodiment 3 of the present invention: Haemophilus parasuis PCR detection kit, it comprises 12.5 μ L PCR premix damping fluid, PCR premix buffering fluid is made of the MgCl of 3mmol / ul 2 , 500pmol / ul dNTP2.0μL, 25mmol / μL Tris-HCl and 0.2μL Taq enzyme composition, its pH value is pH8.3; 25μL ultrapure water; 3μL Marker DL2000; 2μL concentration of 12pmol / μL upper primer And 2 μL of the lower primer with a concentration of 12 pmol / μL, the sequence number of the upstream primer is LPF 5′-GCTAACTCCGTGCCAGCAG-3′, the sequence number of the downstream primer is LPR 5′-GTCATCATGGCCCTTACGAG-3′; 25 μL of proteinase K with a concentration of 20 mg / ml , 50 μL SDS with a mass concentration of 10%, 500 μL TE buffer, TE buffer includes a mixed solution composed of Tris-HCL10mM and EDTA1mM, the pH value of the mixed solution is 8.0; 2μL DNA fragments of non-Hemophilus parasuis were used as negative Control; use 2 μL of the standard PCR product of Haemophilus parasuis as a positive co...

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Abstract

The invention discloses a PCR detection kit for hacmophilus parasuis. The kit comprises 8-12.5 microliters of PCR premix buffer, 25 microliters of ultrapure water, 2-3 microliters of Marker DL2000, 0.75-2.0 microliters of two primers with a concentration of 12pmol per microliter, 2 microliters of negative control and 2 microliters of positive control. In the invention, the conventional PCR methodis employed to replace a biochemical test and a serological surveillance which are labor and time consuming, and the PCR detection kit for detecting hacmophilus parasuis is developed. Able to detect hacmophilus parasuis, the kit is applicable to detection, diagnosis and epidemiological survey of hacmophilus parasuis. Errors caused by human judgments can be avoided, and harms on pigs are also reduced. With the kit, hacmophilus parasuis can be detected within 24 hours, which greatly shortens the detection time of hacmophilus parasuis. Thus, the kit of the invention is characterized by speediness, high sensitivity, strong specificity and the like.

Description

technical field [0001] The invention relates to a kit, in particular to a PCR detection kit for Haemophilus parasuis. Background technique [0002] Haemophilus parasuis (HPS) is a Gram-negative, non-hemolytic, NAD-dependent bacterium with 15 serotypes. HPS can cause fibrinous polyserositis, polyarthritis, pleurisy and meningitis in pigs. This bacterium has been recognized as the causative agent of Glasser's disease in swine since 1910. HPS exists in the upper respiratory tract of healthy pigs and clinically mainly infects piglets, especially weaned piglets aged 4 to 6 weeks. The clinical feature is acute sepsis and often dies 2 days after infection. The disease can also cause significant economic losses to the swine industry. Haemophilus parasuis disease exists in pig herds of some pig breeding bases, professional pig farmers and farmers in my country, which seriously endangers the health of pig herds. The current market lacks a kit that can detect Haemophilus parasuis i...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/68C12R1/21
Inventor 杨茂生吴位珩徐景峨杨莉
Owner GUIZHOU INST OF ANIMAL HUSBANDRY & VETERINARY
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