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Kit for rapid detection of eperythrozoon, leptospira and toxoplasma in blood and application of kit

A technology of Eperythrozoon and Leptospira, which is applied to the kit of Toxoplasma gondii to quickly detect the field of Eperythrozoon and Leptospira in the blood, achieving the effect of simple operation, easy operation and reduced sample dose

Inactive Publication Date: 2014-05-14
ZHEJIANG UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

At present, this technology is mainly used in the rapid detection of various bacterial and viral diseases, but there is no report or publicity on the multiplex PCR rapid detection method for simultaneous detection of Eperythrozoon, Leptospira, and Toxoplasma gondii in blood

Method used

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  • Kit for rapid detection of eperythrozoon, leptospira and toxoplasma in blood and application of kit
  • Kit for rapid detection of eperythrozoon, leptospira and toxoplasma in blood and application of kit
  • Kit for rapid detection of eperythrozoon, leptospira and toxoplasma in blood and application of kit

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0031] Step (1). Sample DNA extraction

[0032] Take 0.5ml of pig blood from pigs with suspected disease, and extract DNA according to the instructions of the Universal Genomic DNA Extraction Kit to obtain the DNA sample to be tested.

[0033] Step (2). Preparation of PCR reaction system: The total reaction volume is 25 μl, and the PCR reaction solution is placed on ice to melt, and 22.38 μl of the PCR reaction solution, 0.12 μl of Taq DNA polymerase, and 2.5 μl of the DNA sample to be tested are placed in 96 Mix in a well PCR tube, blow and mix with a pipette, and then place in a centrifuge at 5000r / min for 2s. At the same time, a positive control substance and a negative control substance were set up.

[0034] Step (3). PCR amplification detection: place the mixed reaction tube in the PCR instrument, set the reaction conditions, and carry out the PCR reaction. The reaction conditions were set as follows: pre-denaturation at 94°C for 5 min; denaturation at 94°C for 30 s, an...

Embodiment 2

[0037] Step (1). Sample DNA extraction

[0038] Take 0.5ml of healthy pig blood, and extract DNA according to the instructions of the Universal Genomic DNA Extraction Kit to obtain the DNA sample to be tested.

[0039] Step (2). Preparation of PCR reaction system: The total reaction volume is 25 μl, and the PCR reaction solution is placed on ice to melt, and 22.38 μl of the PCR reaction solution, 0.12 μl of Taq DNA polymerase, and 2.5 μl of the DNA sample to be tested are placed in 96 Mix in a well PCR tube, blow and mix with a pipette, and then place in a centrifuge at 5000r / min for 2s. At the same time, a positive control substance and a negative control substance were set up.

[0040] Step (3). PCR amplification detection: place the mixed reaction tube in a PCR machine, set reaction conditions, and perform PCR reaction. The reaction conditions were set as follows: pre-denaturation at 94°C for 5 min; denaturation at 94°C for 30 s, annealing at 56.3°C for 30 s, extension at...

Embodiment 3

[0043] Step (1). Sample DNA extraction

[0044] Take 0.5ml of bovine blood from suspected diseased cattle, and extract DNA according to the instructions of the Universal Genomic DNA Extraction Kit to obtain the DNA sample to be tested.

[0045] Step (2). Preparation of PCR reaction system: The total reaction volume is 25 μl, and the PCR reaction solution is placed on ice to melt, and 22.38 μl of the PCR reaction solution, 0.12 μl of Taq DNA polymerase, and 2.5 μl of the DNA sample to be tested are placed in 96 Mix in a well PCR tube, blow and mix with a pipette, and then place in a centrifuge at 5000r / min for 2s. At the same time, a positive control substance and a negative control substance were set up.

[0046] Step (3). PCR amplification detection: place the mixed reaction tube in a PCR machine, set reaction conditions, and perform PCR reaction. The reaction conditions were set as follows: pre-denaturation at 94°C for 5 min; denaturation at 94°C for 30 s, annealing at 56....

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Abstract

The invention discloses a kit for rapid detection of eperythrozoon, leptospira and toxoplasma in blood and application of the kit. The kit consists of a detachable 96-pore polymerase chain reaction (PCR) tube, PCR reaction liquid, TaqDNA polymerase, standard and control, wherein the PCR reaction liquid contains a PCR buffer solution, dNTPs, MgCl2, KCl, gelatin and various detection primers; the various detection primers are respectively specific primers for eperythrozoon, leptospira and toxoplasma. The kit is applicable to detecting whether blood is infected by one or more pathogens in the eperythrozoon, leptospira and toxoplasma. The kit disclosed by the invention is rapid in detection speed, high in detection sensitivity, good in specificity, and simple and convenient to operate; and the kit can simultaneously identify and detect one or more specimen in mixed infection of the eperythrozoon, leptospira and toxoplasma, thus greatly improving detection accuracy and reduce false positive rate.

Description

technical field [0001] The invention relates to biotechnology, in particular to a kit for rapidly detecting Eperythrozoon, Leptospira, and Toxoplasma in blood, which can simultaneously detect single or multiple blood samples mixed with Erythrozoon, Leptospira, and Toxoplasma. Differential testing. Background technique [0002] Eperythrozoon, Leptospira, and Toxoplasma gondii are three zoonotic pathogens that are widely prevalent all over the world. The common feature of the three is that they all exist in the blood of animals and can cause The elevated body temperature of sick animals is one of the three important pathogens in the previously reported pig "high temperature hyperthermia syndrome". Humans can be infected through contact with sick animals and animal products such as pig blood. Eperythrozoon, also known as haematozoa, is a single-celled protozoa that parasitizes on the surface of human and livestock red blood cells, plasma and bone marrow. A variety of animals ...

Claims

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Application Information

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IPC IPC(8): C12Q1/68
CPCC12Q1/04C12Q1/686C12Q2537/143Y02A50/30
Inventor 徐定婷曲道峰沈连珠
Owner ZHEJIANG UNIV
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