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Reagent and kit for isothermal amplification detection of trypanosoma cruzi and application of reagent and kit

A technology of isothermal amplification and Trypanosoma cruzi, which is applied in the determination/inspection of microorganisms, resistance to vector-borne diseases, DNA/RNA fragments, etc., can solve the problems of high cost and inconvenient detection, and achieve high sensitivity and simplicity The effect of rapid detection and good on-site detection ability

Pending Publication Date: 2021-12-21
SHENZHEN CENT FOR DISEASE CONTROL & PREVENTION +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] The purpose of this application is to provide a reagent for detecting isothermal amplification and detection of Trypanosoma cruzi, a kit and its application, aiming to solve the problem of nucleic acid extraction and fluorescent PCR detection of samples to be tested in the prior art, which require professional operation and cost. High and inconvenient detection problems

Method used

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  • Reagent and kit for isothermal amplification detection of trypanosoma cruzi and application of reagent and kit
  • Reagent and kit for isothermal amplification detection of trypanosoma cruzi and application of reagent and kit
  • Reagent and kit for isothermal amplification detection of trypanosoma cruzi and application of reagent and kit

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Embodiment A1

[0070] This embodiment provides a reagent for detecting Trypanosoma cruzi by isothermal amplification, which includes: the first outer primer SEQ ID No.1, the second outer primer SEQ ID No.2, the first inner primer SEQ ID No.3, the second outer primer Two internal primers SEQ ID No.4, the first loop primer SEQ ID No.5, the second loop primer SEQ ID No.6, Bst polymerase, m-cresyl violet, KCL, MgSO 4 , Tween 20, dNTPs and nuclease-free water, the concentration of each component is as follows in Table 2:

[0071] Table 2

[0072]

[0073]

Embodiment A2

[0075] The embodiment provides a reagent for detecting Trypanosoma cruzi by isothermal amplification, which includes: the first outer primer SEQ ID No.1, the second outer primer SEQ ID No.2, the first inner primer SEQ ID No.3, the second Internal primer SEQ ID No.4, first loop primer SEQ ID No.5, second loop primer SEQ ID No.6, Bst polymerase, m-cresyl violet, KCL, MgSO 4 , Tween 20, dNTPs and nuclease-free water, the concentration of each component is as follows in Table 3:

[0076] table 3

[0077] components starting concentration use concentration Size (μL) first outer primer 0.3mM 1μM 0.12 μL second outer primer 0.3mM 1μM 0.12 μL first inner primer 2.4mM 45μM 0.48 μL second inner primer 2.4mM 45μM 0.48 μL first loop primer 1mM 10μM 0.4μL Second loop primer 0.3mM 10μM 0.12 μL Bst polymerase 8U / μl 0.3U / μl 1μL m-cresyl violet 50mM 0.1mM 0.16μL KCL 1M 50mM 0.2 μL ...

Embodiment A3

[0079] The embodiment provides a reagent for detecting Trypanosoma cruzi by isothermal amplification, which includes: the first outer primer SEQ ID No.1, the second outer primer SEQ ID No.2, the first inner primer SEQ ID No.3, the second Internal primer SEQ ID No.4, first loop primer SEQ ID No.5, second loop primer SEQ ID No.6, Bst polymerase, m-cresyl violet, KCL, MgSO 4 , Tween 20, dNTPs and nuclease-free water, the concentration of each component is as follows in Table 4:

[0080] Table 4

[0081] components starting concentration use concentration Size (μL) first outer primer 0.3mM 1.8μM 0.12 μL second outer primer 0.3mM 1.8μM 0.12 μL first inner primer 2.4mM 57.6μM 0.48 μL second inner primer 2.4mM 57.6μM 0.48 μL first loop primer 1mM 20μM 0.4μL Second loop primer 0.3mM 20μM 0.12 μL Bst polymerase 8U / μl 0.4U / μl 1μL m-cresyl violet 50mM 0.4mM 0.16μL KCL 1M 0.4mM ...

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Abstract

The invention relates to the technical field of molecular biological detection, and provides a reagent and a kit for isothermal amplification detection of trypanosoma cruzi and application of the reagent and the kit. The reagent for isothermal amplification detection of trypanosoma cruzi provided by the invention comprises a loop-mediated isothermal nucleic acid amplification primer group for detecting trypanosoma cruzi: a first outer primer SEQ ID No.1, a second outer primer SEQ ID No.2, a first inner primer SEQ ID No.3, a second inner primer SEQ ID No.4, a first loop primer SEQ ID No.5, and a second loop primer SEQ ID No.6. The loop-mediated isothermal nucleic acid amplification primer group is high in specificity; other plasmodium species in plasmodium can be effectively distinguished, so that the reagent is high in sensitivity, good in repeatability, low in false negative and false positive and high in detection accuracy when being used for detecting trypanosoma cruzi; and result judgment can be directly carried out by observing colour change through naked eyes.

Description

technical field [0001] The application belongs to the technical field of molecular biology detection, and in particular relates to a reagent, a kit and an application thereof for isothermal amplification detection of Trypanosoma cruzi. Background technique [0002] Chagas disease, also known as Chagas disease, is caused by Trypanosomacruzi and is transmitted by triatomine bugs. The disease mainly occurs in 21 Latin American countries, and it is more common in children. The acute phase is characterized by fever, generalized lymphadenopathy, and cardiac enlargement. The chronic phase is characterized by myocarditis, cardiac enlargement, and dilation of the esophagus or colon. In 2017, an estimated 6.2 million people worldwide had Chagas disease, with approximately 162,000 new infections and 7900 deaths each year. This results in an estimated global annual economic burden of US$ 7.2 billion. [0003] Although my country is not an endemic area of ​​American trypanosomiasis, t...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/6844C12Q1/6893C12N15/11
CPCC12Q1/6844C12Q1/6893C12Q2525/307C12Q2527/101C12Q2563/173Y02A50/30
Inventor 陈木新张仁利艾琳黄达娜张岩陈燕旌刘琴唐屹君边素莹林梓丹邓少玉张倩陈家旭苏川周晓农
Owner SHENZHEN CENT FOR DISEASE CONTROL & PREVENTION