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Method for separation and extraction of DNA and RNA in cells

A cell and cell lysis technology, applied in the field of separation and extraction of DNA and RNA in cells, which can solve problems such as the unsatisfactory effect of Trizol

Inactive Publication Date: 2018-11-13
苏州呼呼健康科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] However, when Trizol extracts DNA, the effect is often not ideal, and there is a lot of room for improvement in DNA concentration and purity.

Method used

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  • Method for separation and extraction of DNA and RNA in cells
  • Method for separation and extraction of DNA and RNA in cells
  • Method for separation and extraction of DNA and RNA in cells

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0096] This embodiment provides a method for extracting nucleic acid from hippocampal neurons, including:

[0097] ① Take out the hippocampal neuron cell sample, add 1mL Trizol reagent, pipette for 12-16 times, let stand at 18°C ​​for 30min, add 1μL linearized acrylamide solution, mix by inverting, let stand at 18°C ​​for 8min;

[0098] ② Add 100 μL of pre-cooled BCP reagent, shake vigorously by hand for 20-30 times, until the solution is milky white, let it stand at 18°C ​​for 18 minutes;

[0099] ③Centrifuge at 10000rpm for 20min at 4°C, and prepare the corresponding EP tube labeled with RNA;

[0100] ④ After centrifugation, the solution in the EP tube is divided into three layers, the upper layer is an aqueous solution containing RNA, the middle is a floc layer containing genomic DNA, and the lower layer is a red organic solution layer.

[0101] Carefully remove about 500 μL of the upper aqueous phase solution and transfer to a new EP tube marked with RNA;

[0102] Carefu...

Embodiment 2

[0114] This embodiment provides a method for extracting fibroblast nucleic acid, comprising:

[0115] ① Take out the fibroblast sample, add 1mL Trizol reagent, pipette for 12-16 times, let stand at 30°C for 15min, add 1μL linearized acrylamide solution, mix by inverting, let stand at 30°C for 3min;

[0116] ② Add 100 μL of pre-cooled BCP reagent, shake vigorously by hand for 20-30 times, until the solution is milky white, let it stand at 30°C for 12 minutes;

[0117] ③Centrifuge at 14000rpm for 10min at 4°C, and prepare the corresponding EP tube labeled with RNA;

[0118]④ After centrifugation, the solution in the EP tube is divided into three layers, the upper layer is an aqueous solution containing RNA, the middle is a floc layer containing genomic DNA, and the lower layer is a red organic solution layer.

[0119] Carefully remove about 500 μL of the upper aqueous phase solution and transfer to a new EP tube marked with RNA;

[0120] Carefully remove about 300 μL of the aq...

Embodiment 3

[0132] The present embodiment provides a kind of method of extracting PBMC nucleic acid, comprising:

[0133] ① Take out the PBMC sample, add 1mL Trizol reagent, pipette for 12-16 times, let stand at 20°C for 25min, add 1μL linearized acrylamide solution, mix by inverting, let stand at 20°C for 4min;

[0134] ② Add 100 μL of pre-cooled BCP reagent, shake vigorously by hand for 20-30 times, wait until the solution is milky white, and let stand at 20°C for 4 minutes;

[0135] ③Centrifuge at 13000rpm for 12min at 4°C, and prepare the corresponding EP tube labeled with RNA;

[0136] ④ After centrifugation, the solution in the EP tube is divided into three layers, the upper layer is an aqueous solution containing RNA, the middle is a floc layer containing genomic DNA, and the lower layer is a red organic solution layer.

[0137] Carefully remove about 500 μL of the upper aqueous phase solution and transfer to a new EP tube marked with RNA;

[0138] Carefully remove about 300 μL o...

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Abstract

The invention relates to the technical field of biology, in particular to a method for separation and extraction of DNA and RNA in cells. The method includes: a) adopting Trizol for splitting a cell sample to release nucleic acid to obtain a split product; b) contacting the split product with at least one organic extracting solvent, and centrifuging to layer the split product into an upper RNA aqueous phase layer, a middle DNA flocculent layer and a lower organic phase layer; c) respectively subjecting the RNA aqueous phase layer and the DNA flocculent layer to precipitation and washing, wherein a DNA precipitation agent for precipitation comprises 1,2-propanediol. By adoption of the method, problems of low yield and poor purity in simultaneous extraction of DNA and RNA can be effectivelyavoided.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to a method for separating and extracting DNA and RNA in cells. Background technique [0002] Prior art methods of concentrating, extracting, isolating and purifying nucleic acids from cellular source materials involve complex multi-step processes. When processing some biological samples or conducting clinical tests, the amount of samples obtained is often very scarce, so in order to make full use of the samples, it is usually necessary to extract DNA and RNA at the same time. [0003] In the prior art, the method for simultaneously extracting DNA and RNA from a sample is generally the Trizol lysis method. Trizol reagent is suitable for rapid isolation of RNA from cells and tissues, and is a mature RNA extraction reagent. Compared with other methods such as guanidine thiocyanate / phenol method, phenol / SDS method, guanidine hydrochloride method, guanidine thiocyanate method, etc., the T...

Claims

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Application Information

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IPC IPC(8): C12N15/10
CPCC12N15/1003
Inventor 江南李丹
Owner 苏州呼呼健康科技有限公司
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