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Standard sample for calibration of PCR instrument, and construction method and application method thereof

A standard sample and construction method technology, applied in the field of PCR instrument, can solve the problems of no PCR instrument, aerosol pollution, and no unified PCR instrument calibration system and method, so as to avoid cross-contamination, easy storage, and avoid false positive or false Effects of Negative Results

Inactive Publication Date: 2019-07-26
杭州博度计量科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] However, standard samples and methods thereof are currently in short supply as PCR instrument calibration, and there is no standard sample for PCR instrument calibration in the prior art; there is no unified system and method for PCR instrument calibration; and the conventional PCR detection determines the presence of aerosol Contamination phenomenon, for this reason, we propose a standard sample, construction method and application method for PCR instrument calibration

Method used

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  • Standard sample for calibration of PCR instrument, and construction method and application method thereof
  • Standard sample for calibration of PCR instrument, and construction method and application method thereof
  • Standard sample for calibration of PCR instrument, and construction method and application method thereof

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Embodiment 1

[0038] Such as figure 1 In the shown embodiment one, a kind of standard sample for PCR instrument calibration, described a kind of PCR instrument calibration standard sample is composed of nucleotide (not limited to) fragments, adding common enzyme cutting sites, inserted into conventional cloning Prepared from a carrier, the standard sample for calibration of a PCR instrument is composed of specific animal, plant and microbial gene sequences.

[0039] A kind of construction method of PCR instrument calibration standard sample, a kind of construction method of PCR instrument calibration standard sample comprises the following steps:

[0040] (1) Select specific animal, plant and microbial gene sequences for splicing to obtain unique and specific gene sequences;

[0041] (2) Synthetic nucleotide fragments;

[0042] (3) Design PCR-specific primers;

[0043] (4) PCR amplification;

[0044] (5) The target sequence is obtained and verified by sequencing;

[0045] (6) insertin...

Embodiment 2

[0061] An application method of a standard sample for calibration of a PCR instrument comprises a specific primer probe sequence, a PCR reaction system, and a PCR reaction program.

[0062] 1. Prepare primer probes, standard samples and PCR materials.

[0063] 2. Use 10 μmol / L and 5 μmol / L of the primers and probes in the table below for use.

[0064] Primer probe name sequence Primer-F2 TGGCACGCAGGACAAGGT Primer-R2 CGCGTCCTTGACAAACCAA Probe-P2 5'6-FAM-CGATGTCGCAGTCGG-TAMRA

[0065] 3. Dilute the constructed standard sample according to the following table, and keep it for later use.

[0066]

[0067] 4. Configure the PCR reaction system as shown in the table below.

[0068]

[0069] 5. Transfer the prepared PCR reaction solution to a PCR machine for reaction. The PCR reaction program is: 1) 95°C, 10min; 2) 95°C, 30sec; 60°C, 1min; a total of 40cycles.

[0070] 6. After the PCR reaction is completed, the results can be analyzed.

[...

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Abstract

The invention discloses a standard sample for calibration of a PCR instrument. The standard sample for calibration of the PCR instrument is composed of (not limited to) nucleotide fragments and is prepared by adding a common enzyme digestion site and inserting into a conventional cloning vector. The standard sample and a method thereof for periodic calibration of the PCR instrument are used for monitoring the change of the measurement characteristics of the PCR instrument, greatly avoid the generation of false positive or false negative results in the process of PCR, and improve the accuracy and stability of a PCR test. The standard sample is easy to prepare, low in cost, stable in performance, easy to preserve, good in universality and high in accuracy. The standard sample and the methodthereof can effectively avoid cross-contamination, improve the accuracy of calibration detection, are simple to operate and short in use time, and have safe and non-toxic composition.

Description

technical field [0001] The invention relates to a PCR instrument, in particular to a standard sample for calibration of a PCR instrument, a construction method and an application method. Background technique [0002] Polymerase Chain Reaction Analyzer (Polymerase Chain Reaction Analyzer, hereinafter referred to as PCR instrument) is an instrument that uses DNA polymerase and the materials required for its reaction to replicate and amplify the gene DNA sequence under certain temperature conditions. It mainly includes two types: ordinary qualitative PCR instrument and fluorescent quantitative PCR instrument. Qualitative PCR usually consists of thermal cycle components, control components and power supply components. Fluorescence quantitative PCR instrument is mainly composed of sample carrier platform, thermal cycle components, transmission components, fluorescence detection optical components, microcircuit control components, computer and application software. [0003] Poly...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/686C12Q1/6806
CPCC12Q1/6806C12Q1/686C12Q2600/166C12Q2531/113
Inventor 金荣愉余笑波孙杨
Owner 杭州博度计量科技有限公司