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Cell lysis solution for rapid gene detection

A cell lysate and rapid technology, applied in the field of molecular biology, can solve problems such as obstruction, low gene amplification efficiency, and incomplete cell lysis

Inactive Publication Date: 2015-06-03
重庆京因生物科技有限责任公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Therefore, directly adding cells as templates to the PCR reaction mixture for reaction will result in incomplete cell lysis during the PCR process, and cell fragments after cell lysis and some proteases will hinder the PCR reaction, resulting in low or low gene amplification efficiency. experiment failed

Method used

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  • Cell lysis solution for rapid gene detection
  • Cell lysis solution for rapid gene detection
  • Cell lysis solution for rapid gene detection

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 2

[0056] (1) Configure cell lysate:

[0057] Use a 1.5ml centrifuge tube, add 100ulSDS and 18.78ulTriton X-100, then add 881.22ul nuclease-free water to a total volume of 1000ul, make the final concentration of SDS reach 1%, and make the final concentration of Triton X-100 reach 2% , that is, 200x cell lysate, then shake and mix well, and store at -4°C.

[0058] The present invention is illustrated by taking the final concentration of Example 2 in the above-mentioned Table 1 and Table 2 as an example, wherein, dNTPs, MgCl 2 , 5x Colorless Reaction Buffer, and DNA polymerase using GoTaq DNA Polymerase were purchased from Shanghai Promega Biological Products Co., Ltd.

[0059] (2) Prepare a 23ul PCR reaction mixture: the amount of each raw material added is shown in Table 3:

[0060] table 3

[0061]

[0062] Add each raw material according to the amount added in each kit in Table 3 to make a PCR reaction mixture.

[0063] Among them: CYP2C19*2 upstream primer, CYP2C19*2 do...

Embodiment 1

[0094] The difference from Example 2 lies in the configuration of the cell lysate and the PCR reaction mixture, and the specific operations are as follows:

[0095] (1) Configure cell lysate:

[0096] Use a 1.5ml centrifuge tube, add 10ul SDS and 1.88ul Triton X-100, then add 988.12ul nuclease-free water to a total volume of 1000ul, make the final concentration of SDS reach 0.1%, and make the final concentration of Triton X-100 reach 2%, that is, 200x cell lysate, then shake and mix, and store at -4°C.

[0097] The present invention is illustrated by taking the final concentration of Example 1 in the above-mentioned Table 1 and Table 2 as an example, wherein, dNTPs, MgCl 2 , 5x Colorless Reaction Buffer, and DNA polymerase using GoTaq DNA Polymerase were all purchased from Shanghai Promega Biological Products Co., Ltd.

[0098] (2) Prepare 23ul of PCR reaction mixture: the amount added is shown in Table 4:

[0099] Table 4

[0100]

[0101] Add each raw material accordi...

Embodiment 4

[0118] The difference from Example 2 lies in the configuration of the cell lysate and the PCR reaction mixture, and the specific operations are as follows:

[0119] (1) Configure cell lysate:

[0120] Use a 1.5ml centrifuge tube, add 300ul SDS and 56.34ul Triton X-100, then add 643.66ul nuclease-free water to a total volume of 1000ul, so that the final concentration of SDS reaches 3%, and the final concentration of Triton X-100 reaches 6 %, which is the 200x cell lysate, then shake and mix well, and store at -4°C.

[0121] The present invention is illustrated by taking the final concentration of Example 4 in the above Table 1 and Table 2 as an example, wherein, dNTPs, MgCl2, 5x Colorless Reaction Buffer, and DNA polymerase are all purchased from Shanghai Promega Biological Products Co., Ltd. using GoTaq DNA Polymerase .

[0122] (2) Prepare 23ul of PCR reaction mixture: the amount added is shown in Table 5:

[0123] table 5

[0124]

[0125] Add each raw material accord...

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Abstract

The invention discloses a cell lysis solution for rapid gene detection. The cell lysis solution comprises a PCR mixed solution, wherein the PCR mixed solution is prepared from the following raw materials: DNA polymerase, dNTPs, an upstream primer, a downstream primer, a mutant-type molecular beacon, a wild-type molecular beacon, MgCl2, a PCR buffer solution and the cell lysis solution. The invention also provides the cell lysis solution which can realize gene amplification or detection by using a cell as a template in a manner of cooperating with the PCR mixed solution and an application method of the cell lysis solution.

Description

technical field [0001] The invention belongs to the field of molecular biology, and in particular relates to a cell lysate for rapid gene detection. Background technique [0002] Polymerase Chain Reaction (PCR) is a method for enzymatically synthesizing specific DNA fragments in vitro, and is one of the most commonly used molecular biology techniques. Since Muller invented the polymerase chain reaction (PCR) in 1985, the way biological scientists conduct research has gradually changed. This technology, which can obtain a large number of DNA fragments in a short time, plays a huge role in forensic science, DNA cloning, genome analysis, genetic disease and infectious disease diagnosis. [0003] Once the PCR technology came out, it was rapidly and widely used in various fields of molecular biology. It can be used not only for the isolation, cloning and nucleotide sequence analysis of genes, but also for the construction of mutants and recombinants, the study of gene expressio...

Claims

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Application Information

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IPC IPC(8): C12Q1/68
Inventor 任晓东罗志超
Owner 重庆京因生物科技有限责任公司
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