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Toxoplasma gondii rapid detection kit and preparation method thereof

A detection kit, a technology for Toxoplasma gondii, which is applied in the field of temperature amplification, can solve the problems of affecting specificity and sensitivity, and long reaction time, and achieves the effects of rapid reaction, simple operation, and long detection time.

Active Publication Date: 2011-02-16
LANZHOU INST OF VETERINARY SCI CHINESE ACAD OF AGRI SCI
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Problems solved by technology

However, the PCR method requires an expensive PCR instrument to be used, and the reaction time is long. After the reaction, agarose gel electrophoresis is required to observe the results with a gel imager, and there may be some leftovers during the extraction process of animal genome DNA. Inhibitors of PCR reactions affect their specificity and sensitivity

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  • Toxoplasma gondii rapid detection kit and preparation method thereof
  • Toxoplasma gondii rapid detection kit and preparation method thereof
  • Toxoplasma gondii rapid detection kit and preparation method thereof

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Embodiment 1

[0027] (1) Preparation of LAMP reaction solution:

[0028] ① Configure Tris-HCl: Weigh 121.1g Tris in 800mLddH 2 Dissolve in O, adjust the pH to 8.8 with HCl, set the volume to 1000mL, and set aside;

[0029] ② Weigh 1.49g KCl, 3.96g MgSO 4 ·7H 2 O, 2.64g (NH4) 2 SO 4 , 187.4g betaine, and add 40mL of Tris-HCl and 2mL Tween 20 configured in ①, add dd H 2 O is fixed to 900mL, for subsequent use;

[0030] ③ Take 900 μL of the solution prepared in ②, add 100 μL of 25mM dNTPs, and mix well.

[0031] (2) Prepare the primer mixture:

[0032] ①Aiming at a highly conserved region of Toxoplasma gondii nucleoside triphosphate hydrolase gene, primers were designed using PrimerExpolorer V4 online software;

[0033] ② Mixed primers include 22.4 μL FIP and BIP, 2.8 μL F3 and B3, and 11.2 μL LF and LB, mix well.

[0034] (3) Preparation of standard positive template:

[0035] ① Each mouse with a weight of 20-25 grams weighed 1×10 5 ~1×10 6 Toxoplasma gondii tachyzoites were inocu...

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Abstract

The invention relates to a Toxoplasma gondii rapid detection kit and a preparation method thereof. The kit is based on the molecular biology and the nucleoside triphosphate hydrolase (NTPase) gene with high conservation of Toxoplasma gondii to adopt the loop-mediated isothermal amplification (LAMP) method to perform the rapid detection of Toxoplasma gondii. The kit contains LAMP reaction solution, primer, Bst DNA polymerase, color reagent, standard positive template and dd H2O. The kit of the invention is characterized by high sensitivity and specificity, rapid reaction, simple operation and the like, does not require special instrument and equipment and solves the defects of the prior art such as long detection time of toxoplasmosis, large workload and complicated operation. The kit of the invention can be used to detect whether a clinical sample contains Toxoplasma gondii and used in the early diagnosis and the molecular epidemiological study of toxoplasmosis.

Description

technical field [0001] The invention relates to a circle-mediated isothermal amplification technology for detecting toxoplasma genome DNA, specifically a toxoplasma rapid detection kit, and the invention includes a preparation method of the kit. Background technique [0002] The commonly used methods for diagnosing toxoplasmosis include agglutination test, enzyme-linked immunosorbent assay and PCR, etc., and these methods have certain shortcomings. The agglutination test is mainly the indirect hemagglutination method. The disadvantage of this method is that the detection sensitivity is low, and there may be cases of missed detection; the disadvantage of the enzyme-linked immunosorbent assay is that the required equipment is expensive, and it needs to be used in conjunction with a special microplate reader and operation. The process is relatively complicated and the detection time is relatively long; moreover, these two methods are for the detection of Toxoplasma gondii antib...

Claims

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Application Information

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IPC IPC(8): C12Q1/68
Inventor 张德林王萌王艳华付宝权李文卉
Owner LANZHOU INST OF VETERINARY SCI CHINESE ACAD OF AGRI SCI
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