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Method for extracting nucleic acid from paraffin-embedded tissue section

A technology of paraffin embedding and tissue sectioning, which is applied in the field of nucleic acid extraction, which can solve the problems affecting the purity and concentration of extracted nucleic acid, the limited amount of nucleic acid in the lysate, and the inability to study and test, etc., to protect from oxidation and avoid DNA self-dimerization Body formation, energy saving effect

Pending Publication Date: 2019-08-06
凡知医疗科技(江苏)有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0003] When using the magnetic bead method for nucleic acid extraction, the lysate is mostly directly added to the paraffin-embedded tissue section, which takes a long time and is time-consuming, and the amount of nucleic acid in the lysate is limited, which affects the purity and concentration of the final extracted nucleic acid and cannot be followed up. and the requirements of the experiment, and because RNA is more difficult to extract than DNA, we have designed a method for extracting nucleic acid from paraffin-embedded tissue sections to solve the above problems

Method used

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Examples

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Effect test

Embodiment 1

[0026] A method for extracting DNA from paraffin-embedded tissue sections, comprising the following steps:

[0027] S1, paraffin-embedded section: slice the paraffin-embedded tissue on a paraffin microtome, the thickness of the sliced ​​tissue is 4 μm, float the sliced ​​tissue on the warm water in the slide machine, flatten the tissue, and slide Pick up the tissue to be extracted, put it in an oven to bake, and take it out after the water is dried, and store it at room temperature as the sample to be tested for later use;

[0028] S2, lysing: take out the spare sliced ​​tissue in S1, add a new lysate, the new lysate is composed of dithiothreitol solution, guanidine isothiocyanate, CTAB, EDTA and potassium chloride, after the components are mixed, The final concentrations of dithiothreitol solution, guanidine isothiocyanate, CTAB, EDTA, and potassium chloride in the novel lysate are respectively: 4×10 -3 mol / L, 2mol / L, 2×10 -3 mol / L, 5×10 -3 mol / L, 0.4mol / L potassium chlori...

Embodiment 2

[0033] A method for extracting DNA from paraffin-embedded tissue sections, comprising the following steps:

[0034] S1, paraffin-embedded section: slice the paraffin-embedded tissue on a paraffin microtome, the thickness of the sliced ​​tissue is 5 μm, float the sliced ​​tissue on the warm water in the slide machine, flatten the tissue, and slide Pick up the tissue to be extracted, put it in an oven to bake, and take it out after the water is dried, and store it at room temperature as the sample to be tested for later use;

[0035] S2, lysing: take out the spare sliced ​​tissue in S1, add a new lysate, the new lysate is composed of dithiothreitol solution, guanidine isothiocyanate, CTAB, EDTA and potassium chloride, after the components are mixed, The final concentrations of dithiothreitol solution, guanidine isothiocyanate, CTAB, EDTA, and potassium chloride in the novel lysate are respectively: 6×10 -3 mol / L, 3mol / L, 3×10 -3 mol / L, 12×10 -3mol / L, 0.6mol / L, the pH value of...

Embodiment 3

[0040] A method for extracting RNA from paraffin-embedded tissue sections, comprising the following steps:

[0041] S1, paraffin-embedded section: slice the paraffin-embedded tissue on a paraffin microtome, the thickness of the sliced ​​tissue is 6 μm, float the sliced ​​tissue on the warm water in the slide machine, flatten the tissue, and slide Pick up the tissue to be extracted, put it in an oven to bake, and take it out after the water is dried, and store it at room temperature as the sample to be tested for later use;

[0042] S2, lysing: take out the spare sliced ​​tissue in S1, add a new lysate, the new lysate is composed of dithiothreitol solution, guanidine isothiocyanate, CTAB, EDTA and potassium chloride, after the components are mixed, The final concentrations of dithiothreitol solution, guanidine isothiocyanate, CTAB, EDTA, and potassium chloride in the novel lysate are respectively: 8×10 -3 mol / L, 2mol / L, 4×10 -3 mol / L, 10×10 -3 mol / L, 0.8mol / L, the pH value o...

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Abstract

The invention discloses a method for extracting nucleic acid from a paraffin-embedded tissue section. The method includes the following steps: S1, performing paraffin embedding slicing; S2, performingcracking, taking out a standby section tissue in S1, adding novel lysate into the section tissue, the volume ratio of the section tissue to the novel lysate being 1 : 50-100, performing centrifugal layering after performing mixing and uniform shaking for 10-15 min under a temperature of 65-75 DEG C, performing standing after layering, and taking an intermediate layer after standing so that mixedliquor can be obtained; S3, performing combination; S4, performing washing; and S5, performing elution. Cells can be fully and effectively cracked by using the novel lysate to perform the cracking ofnucleic acid on the section tissue, so that a cracking process can be accelerated, the nucleic acid can be protected from oxidizing, and the formation of DNA self-dimer can be avoided; and the methodhas the characteristics of being short in using time, high in extraction efficiency and high in extraction quality, so that the method is suitable for being popularized and used.

Description

technical field [0001] The invention relates to the field of nucleic acid extraction, in particular to a method for extracting nucleic acid from paraffin-embedded tissue sections. Background technique [0002] Nucleic acid includes deoxyribonucleic acid (RNA) and ribonucleic acid (DNA), which are the genetic material of all organisms. In cells, they mainly exist in the nucleus, and in viruses, they exist in the viral capsid. Cells are the basic unit of the structure and function of organisms. It is known that all organisms except viruses are composed of cells, but the life activities of viruses must also be reflected in cells. Biotechnology with nucleic acid as the research object includes a series of technologies such as nucleic acid extraction, cloning, amplification, detection, and sequencing. For nucleic acid extraction, compared with transmission xylene dewaxing, protease solution K incubation and other methods, The extraction method of the magnetic bead method has the...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/10
CPCC12N15/1013
Inventor 张建东宋克清孙诚田鸽杨宁王伟
Owner 凡知医疗科技(江苏)有限公司
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