Method for extracting DNA of fertilized egg genome of ascaris suum

A technology of Ascaris suum and fertilized eggs, which is applied in the biological field, can solve the problems of the influence of genomic DNA extraction, the low content of genomic DNA in fertilized eggs of Ascaris suum, and the difficulty of extracting DNA, and achieve stable methods, reliable quality, and easy extraction. Effect

Inactive Publication Date: 2017-02-22
NANCHANG UNIV
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Problems solved by technology

[0004] For any PCR method, the quality of the genomic DNA template is one of the key factors to determine the success of DNA amplification. The special feature of fertilized eggs of Ascaris suum is that the eggs of Ascaris worms have a hard and dense protein shell And the impermeable lipid layer, so that the eggs have strong resistance to various physical and chemical substances and adverse external environments, but it has a certain impact on the extraction of genomic DNA from fertilized eggs
At the same time, the genome DNA content of fertilized eggs of Ascaris suum is very small, so it is very difficult to extract DNA

Method used

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  • Method for extracting DNA of fertilized egg genome of ascaris suum

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Embodiment 1

[0017] A method for extracting genomic DNA from fertilized eggs of Ascaris suum is as follows.

[0018] (1) Wash the eggs with distilled water.

[0019] Use a 1.5ml centrifuge tube, add distilled water and eggs, vortex at 3200rpm for 1min, then centrifuge at 10000r / min for 2min, discard the supernatant, repeat this process 3 times.

[0020] (2) Pretreatment of eggs.

[0021] Place the centrifuge tube alternately in boiling water at 100°C and -80°C for 1 min. After 6 times of repeated freezing and thawing, add PowerBead to the centrifuge tube, place it horizontally on a vortex shaker at 3200rpm for 10min, and centrifuge at 10000r / min for 30sec. , transfer the supernatant to a 1.5ml centrifuge tube.

[0022] (3) Proteinase K digestion.

[0023] Use Wizard ® Nucleic acid lysate and 0.5M EDTA (pH8.0) in the SV Genomic DNA Purification System kit: Add 220μl of nucleic acid lysate, 50μl of 0.5M EDTA (pH8.0), and 20μl of proteinase K (50mg / ml) to the above centrifuge tube , 56 ...

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Abstract

The invention provides a method for efficiently extracting DNA of a fertilized egg genome of ascaris suum. The method comprises the following five steps: washing eggs with distilled water; pre-treating the eggs; digesting protease K; removing impurities; and eluting DNA with high-purity water. The extracted DNA of the fertilized egg genome of ascaris suum has stable quality and can be used for effectively performing downstream PCR amplification, and an important technical method is provided to studying of molecular epidemiology, genetic diversity, genetic structure, systemic embryology and phylogeography of ascaris suum. The method has an important meaning to molecular diagnosis, prevention and control on porcine ascariasis.

Description

technical field [0001] The invention relates to the field of biotechnology, and relates to a method for extracting genomic DNA from fertilized eggs of Ascaris suum. Background technique [0002] Ascaris suum ( Ascaris suum Goeze, 1782) belongs to the phylum Nematodes, class Caudosensilla, order Ascaria, and genus Ascaris. It is the largest nematode parasitic in the small intestine of pigs, and the fresh worm body is light red or light yellow. The development of Ascaris suum includes three different developmental stages: eggs, larvae, and adults. The newly hatched eggs belong to the single-cell stage and are discharged to the outside with the feces. , forming infective eggs (ie third stage larvae). After the infectious eggs are swallowed by pigs, larvae hatch in the small intestine. Most of the larvae enter the blood vessels through the intestinal wall, enter the liver with the blood flow, and then migrate to the lungs. Finally, they undergo two molts in the small intesti...

Claims

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Application Information

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IPC IPC(8): C12N15/10
CPCC12N15/1003C12Q2521/537
Inventor 周春花邓仕标石琴华
Owner NANCHANG UNIV
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