Kit and method for extracting DNA of FFPE tissue sample and application thereof

A technology of tissue samples and kits, which is applied in the field of DNA extraction kits for FFPE tissue samples, can solve the problems of low DNA extraction, long dewaxing time, and low purity, so as to reduce manual operation procedures, shorten dewaxing time, The effect of high purity

Pending Publication Date: 2021-07-09
杭州康代思锐生物科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0008] The technical problem to be solved by the present invention is to overcome the disadvantages of low extraction amount and low purity of DNA extracted from FFPE tissue samples caused by the need for oil phase reagent

Method used

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  • Kit and method for extracting DNA of FFPE tissue sample and application thereof
  • Kit and method for extracting DNA of FFPE tissue sample and application thereof
  • Kit and method for extracting DNA of FFPE tissue sample and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0064] 1. Scrape FFPE tissue samples (5-10μm thick, 1×1cm 2 size) 1-4 pieces into a 1.5mL centrifuge tube, add 300μL deparaffinization buffer (2% SDS, 200mM Tris-HCl, 10mM EDTA-2Na, 1% Triton X-100, 0.1M NaCl , 1% Tween 20, 2% PEG6000), and then add 20 μL of proteinase K to shake and mix, and incubate in a metal bath at 55° C. for 15 minutes.

[0065] 2. Raise the temperature of the metal bath to 80°C and continue to incubate for 15 minutes (note: it is not necessary to take out the centrifuge tube, the temperature can be directly raised from 55°C to 80°C). The phase solution (about 250 μL) was aspirated and transferred to a new 1.5mL centrifuge tube.

[0066] 3. Add 30 μL of magnetic beads and 300 μL of binding solution (65% isopropanol, 0.2M NaCl) to the centrifuge tube in turn, mix by pipetting, and then mix every 2 minutes for 3 times to avoid magnetic Beads gather.

[0067] 4. Place the centrifuge tube on the magnetic stand for 30 seconds, so that the magnetic beads ar...

Embodiment 2

[0074] 1. Scrape FFPE tissue samples (5-10μm thick, 1×1cm 2 size) 1-4 pieces into a 1.5mL centrifuge tube, add 300μL deparaffinization buffer (3% SDS, 100mM Tris-HCl, 5mM EDTA-2Na, 0.5% Triton X-100, 0.2M NaCl , 0.1% Tween 20 and 0.1% PEG6000), 20 μL of proteinase K, mixed by shaking, and incubated at 56° C. for 15 min.

[0075] 2. Raise the temperature of the metal bath to 80°C, and continue to incubate for 15 minutes (Note: It is not necessary to take out the centrifuge tube, the temperature can be directly raised from 55°C to 80°C). The aqueous phase solution (about 250 μL) was aspirated and transferred to a new 1.5mL centrifuge tube.

[0076] 3. Add 20 μL of magnetic beads and 300 μL of binding solution (80% isopropanol and 0.4M NaCl) in sequence, shake and mix, and then mix every 3 minutes for 3 times to avoid aggregation of magnetic beads.

[0077] 4. Place the centrifuge tube on the magnetic stand for 30 seconds, so that the magnetic beads are completely absorbed. Ca...

Embodiment 3

[0084] 1. Scrape FFPE tissue samples (5-10μm thick, 1×1cm 2 size) 1-4 pieces into a 1.5mL centrifuge tube, add 300μL deparaffinization buffer (2% SDS, 200mM Tris-HCl, 10mM EDTA-2Na, 1% Triton X-100, 0.1M NaCl, 1% Tween 20, 2% PEG6000), and then add 20 μL of proteinase K to shake and mix, and incubate in a metal bath at 55° C. for 30 min.

[0085] 2. Raise the temperature of the metal bath to 80°C and continue to incubate for 30 minutes (Note: It is not necessary to take out the centrifuge tube, the temperature can be raised directly from 55°C to 80°C). The aqueous solution was aspirated and transferred to a new 1.5mL centrifuge tube.

[0086] Subsequent steps are the same as in Example 1.

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Abstract

The invention discloses a kit and a method for extracting DNA of an FFPE tissue sample and application thereof. The kit comprises a dewaxing and cracking buffer solution and protease K, wherein the dewaxing and cracking buffer solution is prepared from the following components in percentage by weight: 1 to 6 percent of lauryl sodium sulfate, 50 to 300 mM of Tris-HCl, 1 to 30 mM of EDTA-2Na, 0.1 to 4 percent of Triton X-100, 0.1 to 2 M of NaCl, 0.1 to 8 percent of Tween 20 and 0.1 to 10 percent of PEG 6000; the percentage is volume percentage. According to the kit, the unique dewaxing lysis buffer solution is adopted, dewaxing and tissue lysis of the FFPE tissue sample can be achieved without independently adding a dewaxing agent, the dewaxing step is simplified, meanwhile, the manual operation process is reduced, and the nucleic acid extraction and purification efficiency is improved.

Description

technical field [0001] The invention belongs to the field of biological tissue preservation, and in particular relates to a kit, method and application for DNA extraction, in particular to a kit, method and application for DNA extraction of FFPE tissue samples. Background technique [0002] Tumor tissue retains its unique genetic information, which is very precious and irreplaceable, and is an important research material for medical workers. However, tumor tissue must be stored under ultra-low temperature refrigeration after isolation, otherwise its original structure and Poor preservation will easily bring inconvenience to the follow-up research. FFPE (Formalin-Fixed and Parrffin-Embedded) is formalin-fixed and paraffin-embedded tissue samples, which can be stored at room temperature for a long time. The researchers processed the tumor tissue blocks for paraffin-embedding, and then processed them with an ultra-thin slicer Tissue samples with a thickness of 5-10 μm can achi...

Claims

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Application Information

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IPC IPC(8): C12N15/10
CPCC12N15/1003C12N15/1013
Inventor 雷小军高玉晓张瑞王琛
Owner 杭州康代思锐生物科技有限公司
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