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Extracting kit for formalin soaked tissue DNA and extraction method

A formalin and kit technology, applied in the field of DNA extraction, can solve the problems of difficulty in obtaining, long extraction time, and inability to break cells well, and achieve high recovery rate, good DNA quality, and improved DNA recovery rate. Effect

Pending Publication Date: 2018-10-26
GENE CRAB BIOTECH CO
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

At present, the extraction time of these methods is long, which cannot meet the needs of clinical rapid detection, and cannot break cells well, and remove nucleic acid-protein cross-linking, so it is difficult to obtain high-quality DNA

Method used

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  • Extracting kit for formalin soaked tissue DNA and extraction method
  • Extracting kit for formalin soaked tissue DNA and extraction method
  • Extracting kit for formalin soaked tissue DNA and extraction method

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0052] Formalin Soaked Tissue DNA Extraction Kit

[0053] 1. Preparation of SDS buffer: Take 1g of SDS, add 50mM Tris-HCl (pH8.1) and mix well, and dilute to 100ml.

[0054] 2. Preparation of binding buffer: Weigh 31.84g GuHCl, 4.38g NaCl, 0.42g SDS, 70g Triton X-100, add 80ml ultrapure water, mix well, add 0.5ml EDTA, 5ml Tris-HCl, and dilute to 100ml.

[0055] 3. Preparation of 1.28mol / LEDTA: Weigh 18.61g EDTA into a 50mL beaker, and add 40mL ultrapure water. Place the beaker on a magnetic stirrer, add NaOH solid while stirring until the solution becomes clear, adjust the pH to 8.0, and dilute to 50 mL.

[0056] 4. 1mol / LTris-HCl (pH 8.1) solution preparation: Weigh 12.11g Tris into a 100mL beaker, and add 80mL ultrapure water to completely dissolve it, measure the pH with a pH meter, and adjust the pH with concentrated hydrochloric acid. Transfer the solution to a 100mL volumetric flask and add purified water to make the volume up to 100mL.

[0057] 5. Preparation of the first wash...

Embodiment 2

[0061] Pretreatment

[0062] 1. Select the lung tissue of a cancer patient soaked in formalin, take the tissue with an 18-gauge puncture needle, use a scalpel blade to cut the tissue into granules with a diameter of 1mm, and divide them into 5 samples with a weight of 4.2mg. . Open the screw cap of microTUBE, add 100μl SDS Buffer, add the minced tissue to the Buffer, tighten the cap, and number 1-5 respectively.

[0063] 2. Turn on the ultrasonic disruptor and set up the program. Under the condition that other parameters are the same, the fragmentation time of samples 1-5 increases in order, respectively, 120s, 180s, 240s, 300s, 360s. Samples No. 1 and No. 2 can be seen to have a small amount of tissue mass, which is not completely homogenized, and the other three groups are completely homogenized. See Table 1 for ultrasonic breaking parameters.

[0064] Table 1 Ultrasonic breaking parameters

[0065]

[0066] 3. Open the lid and pipette the sample from the microTUBE into a 1.5ml E...

Embodiment 3

[0088] 1. Select a lung tissue of a cancer patient soaked in formalin, take the tissue with an 18-gauge puncture needle, use a scalpel to cut the tissue into granules with a diameter of 1 mm, and divide them into 5 samples with a weight of 3.9 mg. . Open the screw cap of microTUBE, add 100μl SDS Buffer, add the minced tissue to the Buffer, tighten the cap, and number 1-5 respectively.

[0089] 2. Turn on the ultrasonic disruptor and set the program to disrupt the tissue. Under the condition that other parameters are the same, the disruption temperature of samples 1-5 increases sequentially, respectively 18℃, 20℃, 22℃, 24℃, 26℃. It can be seen that the 5 groups of samples are completely homogenized. See Table 3 for ultrasonic breaking parameters.

[0090] Table 3 Ultrasonic breaking parameters

[0091]

[0092] 3. Open the lid and pipette the sample from the microTUBE into a 1.5ml EP tube.

[0093] 4. Add 20μl proteinase K to the sample and mix well.

[0094] 5. Incubate at 56°C for ...

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Abstract

The invention provides an extracting kit for a formalin soaked tissue DNA and an extraction method, belonging to the technical field of DNA extraction. The kit contains the following components: an SDS buffer solution, a binding buffer solution, proteinase K, DNA purified magnetic beads, a first cleansing solution, a second cleansing solution and eluant. The extraction method comprises the following steps: (1) mixing a formalin soaked tissue with the SDS buffer solution, carrying out ultrasonic crushing so as to obtain a lysis solution, and carrying out two-step digestion on the lysis solutionby virtue of proteinase K; and carrying out extraction by virtue of the DNA purified magnetic beads, and carrying out elution by virtue of the eluant, so as to obtain the formalin soaked tissue DNA.According to the extracting kit and the extraction method, the extraction of the formalin soaked tissue DNA can be realized, and the extracted DNA is good in quality and high in recycling rate.

Description

Technical field [0001] The invention belongs to the technical field of DNA extraction, and in particular relates to a formalin-soaked tissue DNA extraction kit and an extraction method. Background technique [0002] In recent years, the incidence of cancer has increased worldwide, and most of the occurrence of tumors is the result of multiple gene mutations. Compared with traditional first-generation sequencing, next-generation sequencing (NGS) is a sequencing technology that can simultaneously detect multiple genes. The rapid development of second-generation sequencing has brought good news to the personalized diagnosis and precise treatment of tumors, and has greatly improved the quality of life of cancer patients. However, the first condition for these large-scale genetic tests is to obtain better quality DNA samples. The most direct object of tumor genetic testing is patient tissue specimens, but because fresh cancer tissue samples are difficult to sample and transport, and...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/10
CPCC12N15/1013
Inventor 林小静王海波侯军艳辛琳
Owner GENE CRAB BIOTECH CO
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