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Method for extracting fecal microbial DNA of pandas

A technology of fecal microorganisms and giant pandas, which is applied in the field of DNA extraction of giant panda feces microorganisms, can solve the problems of poor repeatability and low DNA concentration, and achieve the goal of ensuring repeatability, increasing the concentration of DNA, and improving the repeatability of DNA extraction Effect

Inactive Publication Date: 2019-10-22
CHENGDU RES BASE OF GIANT PANDA BREEDING
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Problems solved by technology

[0004] Aiming at the problems of low DNA concentration and poor repeatability in the extraction process of fecal microbial DNA in the prior art, the present invention provides a method for extracting giant panda fecal microbial DNA, the purpose of which is to fully break the cell walls of bacteria in large particles , to further increase the DNA concentration and improve the repeatability of DNA extraction

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  • Method for extracting fecal microbial DNA of pandas

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Experimental program
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Embodiment

[0052] 1. Experiment preparation:

[0053]Prepare silicon beads (zirconia / silica beads, 0.1mm, 1mm), medicine spoon, 500ml beaker, glass rod, 500ml blue cap bottle, centrifuge tube (1.5ml, 2ml, 100ml), 2.0ml screw-top cryopreservation tube, gauze , pipette tips (200 μL, 1000 μL) have been sterilized; the water bath is adjusted to 99°C; Liquid gun (100 μL, 1000 μL); stool kit (QIAamp DNA Stool MiniKit); normal saline in a 4°C refrigerator to pre-cool; electronic balance; oscillator (Analytik Jena AG SpeedMill PLUS) and vortex mixer.

[0054] Weigh the silicon beads in advance: Weigh 0.2g of silicon beads with a diameter of 0.1mm, and weigh 0.5g of silicon beads with a diameter of 1.0mm, and put them into the same 2ml cryopreservation tube for later use.

[0055] 2. Sample collection and processing

[0056] Step 1. Feces collection: Collect 200g of fresh giant panda feces in sampling bags or tubes, freeze them immediately with liquid nitrogen, and then transport them back to t...

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Abstract

The invention relates to a method for extracting bacterial DNA, in particular to a method for extracting fecal microbial DNA of pandasby taking the fecal microbial DNA of the pandas as a specific object. In view of the problems of low DNA concentration and poor repeatability in the fecal microbial DNA extraction processin the prior art, according to the technical scheme, the method includes the following steps of 1, making a fecal sample into a bacterial suspension to conduct cryopreservation; 2, taking the cryopreserved bacterial suspension out, adding beads with the diameter of 0.9-1.1 mm, conducting high-speed shaking and beating through a shaker or a homogenizer to break large particles in the bacterial suspension, adding beads with the diameter of 0.09-0.11 mm into the bacterial suspension, and conducting high-speed shaking and beating through the shaker or the homogenizer to cell walls of bacteria; 3, extracting DNA from the bacterial suspension treated in step 2 by a kit method.The method for extracting the fecal microbial DNA of the pandas has the advantages of being high in DNA concentration in the extraction process, high in extracted DNA concentration, and good in extraction repeatability.

Description

technical field [0001] The invention relates to a method for extracting bacterial DNA, takes giant panda feces microbial DNA as a specific object, and specifically relates to a method for extracting giant panda feces microbial DNA. Background technique [0002] At present, there are many methods and kits for extracting fecal microbial DNA at home and abroad, but the concentration of DNA extracted by these methods is low. In order to improve this shortcoming, researchers have added a method of mechanically breaking the bacterial cell wall, that is, adding 0.1mm or 1mm beads before DNA extraction, and then using an oscillator / homogenizer to perform high-speed oscillations to beat the bacterial cell wall (such as doi: 10.1128 / mBio.00022-15, 10.1038 / s41396-018-0051-y), destroy the bacterial cell wall structure to release DNA, and then use the kit method to extract DNA. [0003] Although these methods increase the concentration of fecal microbial DNA, the DNA concentration is st...

Claims

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Application Information

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IPC IPC(8): C12N15/10
CPCC12N15/1006C12Q2563/149
Inventor 张文平谢军金侯蓉张志和张亮沈富军刘佳文王娟韩富瑶刘红王洁
Owner CHENGDU RES BASE OF GIANT PANDA BREEDING
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