Kit for extracting DNA through magnetic bead method and extraction method

A magnetic bead method and kit technology, applied in the field of molecular biology, can solve problems such as the inability to rule out the influence of polysaccharides, the inability to detect the target DNA, and the interference of the test system. It achieves simple and efficient operation methods, high DNA extraction rates, and reduced interference effect

Active Publication Date: 2018-10-19
浙江迪恩生物科技股份有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Although most of the fungal DNA extraction by the magnetic bead method is easy to operate, its disadvantage is that the influence of polysaccharides on DNA adsorption cannot be ruled out.
[0005] In addition, although there are many classic methods for the DNA extraction of organisms, these traditional methods cannot meet the needs of some tests increasingly. Some tests require DNA with high purity and do not want to contain any ot

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  • Kit for extracting DNA through magnetic bead method and extraction method
  • Kit for extracting DNA through magnetic bead method and extraction method
  • Kit for extracting DNA through magnetic bead method and extraction method

Examples

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Example Embodiment

[0065] Example 1: DNA extraction kit for sclerotium produced by magnetic beads

[0066] 1.1. The magnetic bead method for sclerotium-producing fungus DNA extraction kit includes (A7-A8 samples):

[0067] Step 1: Under aseptic conditions, clip the sclerotia of the cultured fungus (yeast) into a 1.5ml centrifuge tube, the sclerotia mass is not less than 500mg, and freeze-grind with liquid nitrogen. Add 250μL solution I, then add 50μL RNase A, mix well.

[0068] Step 2: Add 50μL of proteinase K, mix thoroughly and then bath in a constant temperature water bath at 60°C for 20 minutes. Centrifuge at 12000 rpm for 5 minutes, and transfer the supernatant to a new tube.

[0069] Step 3: Add 150μL of solution II, gently flip up and down 5-6 times. Centrifuge at 12000 rpm for 5 minutes, and transfer the supernatant to a new tube.

[0070] Step 4: Add 350μL of solution III and gently turn it up and down to fully lyse the bacterial solution into a transparent solution. Centrifuge at 12000 rpm f...

Example Embodiment

[0099] Example 2: DNA extraction from samples using processed magnetic beads

[0100] According to the extraction process described in Example 1, the difference is: before using magnetic beads for DNA extraction, the purchased magnetic beads are processed, and the processed magnetic beads are used for DNA extraction. The practicality and implementation of other methods and reagents The magnetic bead extraction method in Example 1 is the same.

[0101] To process the magnetic beads, the processing method is as follows:

[0102] The magnetic beads were purchased from Shanghai Jiaofei Bio-Pharmaceutical Technology Co., Ltd. (the same batch number as in Example 1). The magnetic beads were specially used for DNA extraction. The magnetic beads were treated with glyphosate solution (5mol / L) and Soak in guanidine isothiocyanate solution (8mol / L). The treatment method is as follows: Take 2g of magnetic beads and soak in 10ml of Glyphosate solution for 5-10 hours, rinse with PBS buffer (pH=7...

Example Embodiment

[0109] Example 3: Determination of DNA adsorption capacity using treated magnetic beads

[0110] Use an ultrasonic cell disruptor to disrupt the cell wall of the bacteria to be tested (Staphylococcus aureus). The specific methods of disruption are as follows:

[0111] Scrape the bacteria and prepare a suspension with distilled water (10 7 Pieces / mL), then place the beaker containing the suspension in the disintegrator, put the probe of the disintegrator into the beaker and insert the spore suspension into the spore suspension, set the disintegration time to 10min in an ice bath environment, and take out the sample after cell disintegration. Magnetic beads for DNA detection.

[0112] Add a certain weight of the magnetic beads (but 2-5 grams) of the present invention (the treated magnetic beads in Example 2) to the broken sample liquid, and place the beaker on the magnetic stand for DNA magnetic bead adsorption. After 10 minutes, take out the magnetic beads, rinse with washing solutio...

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Abstract

The invention discloses a kit for extracting DNA through a magnetic bead method and a rapid and simple method for extracting polysaccharide fungus DNA. The kit comprises a solution (I), a solution (II), a solution (III), a solution (IV), a washing solution, eluent, and magnetic bead suspension. The solution (II) is innovatively used to remove polysaccharides of fungi, and the interference of extracellular polysaccharides is eliminated during the DNA extraction process. During the test process, the magnetic beads of the kit are preprocessed, DNA is taken as the molecular target, the adsorbing performance of magnetic beads on RNA and proteins is weakened, and the operation has the characteristic of high specificity. The kit is used to extract DNA of polysaccharide fungi, the detection is simple and convenient, and the interference of extracellular polysaccharides on DNA extraction is reduced.

Description

technical field [0001] The invention belongs to the field of molecular biology, and in particular relates to a DNA extraction kit for sclerotia-producing fungi by a magnetic bead method and a method for quickly extracting DNA of sclerotia-producing fungi with strong specificity. Background technique [0002] Sclerotia is a dormant body composed of closely intertwined hyphae, and its main function is to resist adverse environments. Common sclerotia-producing fungi include Sclerotinia and Rhizoctonia, which contain a large amount of exopolysaccharides. The fungal exopolysaccharide has the characteristics of high adsorption and high viscosity, which is one of the difficulties in the separation and extraction of high-purity DNA from sclerotia-producing fungi. Molecular biological methods are now one of the essential technologies for fungal identification, and the separation and purification of DNA is the basis for molecular biological research. At present, the traditional CTAB...

Claims

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Application Information

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IPC IPC(8): C12N15/10
CPCC12N15/1013
Inventor 黄宵戴明雁徐华莉高颖张明洲
Owner 浙江迪恩生物科技股份有限公司
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