Storage method of pcr reaction reagent and reaction kit

A technology for reaction reagents and preservation methods, applied in biochemical equipment and methods, microorganism determination/inspection, etc., can solve problems such as high melting point, influence experimental results, influence UNG enzyme anti-pollution and other problems, achieve low cost and simple manufacturing method Effect

Active Publication Date: 2012-02-01
深圳生科原生物有限公司
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

In order to solve the above problems, there are currently two methods, one is to use vegetable oil as a steam barrier in the hot-start PCR reaction to reduce solvent evaporation, but this method cannot solve the problem of maintaining Taq enzyme activity for a long time, and the other It is to use solid paraffin to isolate the enzyme in the PCR reaction from other components, creating more practical and effective hot-start PCR methods, but the existing solid paraffin melting point is too high, about 50 ~ 70 ° C, and at -20 ° C Under certain conditions, PCR reagents can only be stored for 1 to 2 weeks
In addition, if the PCR reaction system also contains UNG enzyme, the suitable temperature for UNG enzyme is 37-50°C. The melting point of existing solid paraffin is too high. To melt, it needs to be heated to above 50°C, which affects the UNG enzyme in the PCR reaction. play an anti-pollution role in the liquid, which may affect the experimental results

Method used

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  • Storage method of pcr reaction reagent and reaction kit
  • Storage method of pcr reaction reagent and reaction kit
  • Storage method of pcr reaction reagent and reaction kit

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0026] Solid paraffin (Shanghai Specimen Model Factory) and liquid paraffin (Tianjin Fuchen Chemical Reagent Co., Ltd.) were heated and melted at a ratio of 1:4 (W / V), mixed evenly, and mixed paraffin was prepared.

[0027] Carry out the experiment according to the method for the determination of paraffin wax in "Chinese Pharmacopoeia (Second Edition)", use a capillary tube with openings at both ends, about 10mm in height, place it in a refrigerator at 4°C for 24 hours, and bind the capillary tube tightly with a rubber ring after the mixed paraffin wax solidifies On a thermometer, the contents of the capillary are attached to the middle of the thermometer bulb. Immerse the capillary together with the thermometer in the temperature transfer liquid. The upper end of the mixed paraffin should be about 10mm below the temperature transfer liquid surface; heat carefully. When the temperature exceeds 0.5°C and the mixed paraffin begins to rise in the capillary, read the temperature d...

Embodiment 2～10

[0029]Basically the same as Example 1, the difference is that the ratios of solid paraffin and liquid paraffin in Examples 2 to 10 are successively 1:5, 1:6, 1:7, 1:8, 1:8.5, 1:9, 1:10 , 1:11, 1:12 (W / V). The melting points of mixed paraffins are shown in Table 1.

[0030] Table 1

[0031] Example

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Abstract

The invention provides a mixed paraffin, a method for storing PCR reaction reagents using the mixed paraffin, and a PCR reaction kit containing the mixed paraffin. The mixed paraffin is uniformly mixed with liquid paraffin and melted solid paraffin, wherein , the solid paraffin is selected from the solid paraffin whose melting point is higher than 50° C., and the ratio of the solid paraffin to the liquid paraffin is 1:4 to 1:12 (w / v). The melting point of the mixed paraffin of the present invention is between 27-38 DEG C, and is used for isolating enzymes and other components in the PCR reaction solution. The melting point is moderate and does not affect the effect of fluorescent quantitative PCR reaction. Moreover, the method for producing mixed paraffin in the embodiment of the present invention is simple and low in cost.

Description

technical field [0001] The invention relates to a method for storing PCR reaction reagents by using mixed paraffin and a PCR reaction kit containing the mixed paraffin. Background technique [0002] Real-time fluorescent quantitative PCR (Real Time Polymerase Chain Reaction, Real Time PCR real-time PCR) technology is a method for quantitative or qualitative analysis of the starting template by real-time detection of the fluorescence signal of each cycle product in the PCR amplification reaction. At present, it is widely used in basic scientific research, clinical diagnosis, disease research and drug development and other fields. The basic principle is: a fluorescent chemical substance is introduced into the PCR reaction system, and as the PCR reaction progresses, the PCR reaction product accumulates continuously, and the fluorescence signal intensity also increases proportionally. After each cycle, a fluorescence signal intensity is collected, so that the change of the prod...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/68
Inventor 张鹏汪再兴吴炳义彭论严琼英
Owner 深圳生科原生物有限公司
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