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Paraffin-based preservation method for nucleic acid isothermal amplification reaction reagent, and reaction reagent

A technology of isothermal amplification and preservation method, which is applied in the field of preservation of warm amplification reaction reagents and their mixtures, can solve the problems of large energy consumption, and achieve the effects of reduced storage and transportation costs, low cost, and simple process

Inactive Publication Date: 2012-10-03
YELLOW SEA FISHERIES RES INST CHINESE ACAD OF FISHERIES SCI
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

The invention patent "Method for Preserving Reagent Mixture of Loop-Mediated Isothermal Amplification (200810159414.3)" discloses a method of adding a desiccant protection agent to the mixture of Loop-Mediated Isothermal Amplification Reagent and then vacuum drying or rapid air-drying to realize Loop-Mediated Isothermal Amplification The method of long-term preservation of the amplification reaction reagent mixture at room temperature, but this method requires complex equipment such as a special vacuum freeze dryer or a fast air dryer, and consumes a lot of energy. It is not a very economical method.

Method used

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  • Paraffin-based preservation method for nucleic acid isothermal amplification reaction reagent, and reaction reagent
  • Paraffin-based preservation method for nucleic acid isothermal amplification reaction reagent, and reaction reagent
  • Paraffin-based preservation method for nucleic acid isothermal amplification reaction reagent, and reaction reagent

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Effect test

Embodiment 1

[0023] The preparation of embodiment 1 different melting point paraffins

[0024] The paraffin mentioned in the present invention refers to a mixture of straight-chain alkanes, branched-chain alkanes or naphthenes with a melting point of 20°C-90°C, which can be solid paraffin (paraffin wax), or a combination of solid paraffin and liquid paraffin (liquid paraffin). ) mixed in proportion to meet the requirements of the above melting point of the mixed paraffin.

[0025] In the experiment, it was found that the melting point of the purchased commercial paraffin wax does not completely correspond to its label. For example, the paraffin wax with the label 60 needs a constant temperature of 60°C-62°C to melt well, so the examples did not classify according to the label of the paraffin wax. Experiment, but the melting point range of paraffin is used as the basis for classification experiments, such as the solid paraffin wax of the label 60 used at 60°C-62°C, the solid paraffin wax of...

Embodiment 2

[0028] Example 2 Using paraffin to seal Bst DNA polymerase and loop-mediated isothermal amplification (loop-mediated isothermal amplification, LAMP) reaction reagent separately, analysis of the influence on the storage effect of LAMP reaction reagent

[0029] 1. Use paraffins with different melting points to seal, isolate and store the mixture of Bst DNA polymerase and LAMP reaction reagent

[0030]Add 1 μL of Bst DNA polymerase (8 U / μL) to 288 0.2 mL centrifuge tubes, and divide the 288 0.2 mL centrifuge tubes into 16 groups as shown in column 1 of Table 2 (18 in each group, Every 2 as a unit, will be used as a repeat in the future random inspection), the first 15 groups will be used as the treatment group, and the last 1 group will be used as the control group. At the same time, as shown in the first column of Table 2, 15 kinds of paraffins with different melting points were sequentially melted respectively, and 54 μL of each kind of paraffins with melting points were melted...

Embodiment 3

[0040] Example 3 Using Paraffin to Block Bst DNA Polymerase and Isolate the LAMP Reaction Reagent Influence Analysis on the Preservation Effect of the LAMP Reaction Reagent

[0041] 1. Use paraffins with different melting points to block Bst DNA polymerase to isolate and store it from the LAMP reaction reagent mixture

[0042] Add 1 μL of Bst DNA polymerase (8 U / μL) to 288 0.2 mL centrifuge tubes, and divide the 288 0.2 mL centrifuge tubes into 16 groups as shown in column 1 of Table 3 (18 in each group, Every 2 as a unit, will be used as a repeat in the future random inspection), the first 15 groups will be used as the treatment group, and the last 1 group will be used as the control group. At the same time, as shown in the first column of Table 3, take 15 kinds of paraffins with different melting points and melt them respectively. After melting the paraffins of each melting point, take 27 μL of Bst DNA polymerase in the 9 centrifuge tubes corresponding to the treatment group...

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Abstract

The invention relates to a paraffin-based preservation method for nucleic acid isothermal amplification reaction reagent, and reaction reagent. The preservation method is as follows: biological enzymes, used for nucleic acid isothermal amplification, are covered or embedded and then separated from one or more components (except biological enzymes) in the nucleic acid isothermal amplification reaction liquid, so that long term preservation of the nucleic acid isothermal amplification reagent at normal temperature can be realized. The isothermal amplification reaction reagent or the mixture thereof both treated by the method provided by the invention can be stored and transported at normal temperature, and can be used directly without any special treatment. And further, the method provided by the invention has the characteristics of simple process and low cost. The isothermal amplification reaction reagent or the mixture thereof treated by the method provided by the invention has the advantage of stable and enduring activity under the convenient storage condition, so that the storage and transportation cost of the isothermal amplification reaction reagent is greatly lowered and great conveniences are brought to actual operation of isothermal amplification reaction at the same time.

Description

technical field [0001] The invention belongs to the technical field of preservation of biochemical reagents, and in particular relates to a paraffin-based method for preserving nucleic acid isothermal amplification reaction reagents and reaction reagents, that is, a method for preserving isothermal amplification reaction reagents and mixtures thereof in molecular biology. Background technique [0002] In the past 20 years, nucleic acid amplification technology represented by polymerase chain reaction (PCR) has been rapidly developed and applied; in recent years, a series of new nucleic acid amplification technologies have been produced. Among them, nucleic acid isothermal amplification (isothermal nucleic acid amplification) can expand the copy number of a specific DNA or RNA fragment at a specific temperature. Nucleic acid isothermal amplification utilizes biological enzymes with special functions to amplify target nucleic acids under constant temperature conditions; isothe...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/10C12Q1/68
Inventor 张庆利黄倢李美霞杨冰刘庆慧李晨
Owner YELLOW SEA FISHERIES RES INST CHINESE ACAD OF FISHERIES SCI
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