Extraction method of total genomic DNA of fish

An extraction method and genome technology, applied in the field of genomic DNA extraction, can solve the problems of low genomic DNA content, dragging phenomenon, and inability to meet the needs of experiments, etc., and achieve the effect of improving the effect

Inactive Publication Date: 2015-11-11
NANCHANG UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Using fish scales as raw materials to extract the total genomic DNA of fish will not cause great harm to fish, but because fish scales contain fewer cells, less genomic DNA content, and more impurities, it is generally not used as experimental materials for extracting genomic DNA
like figure 2 As shown, if fish scales are used as raw materials and the phenol-chloroform method is used to extract DNA, the extracted DNA contains impurities and has a very serious dragging phenomenon, which cannot meet the needs of the experiment.

Method used

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  • Extraction method of total genomic DNA of fish
  • Extraction method of total genomic DNA of fish
  • Extraction method of total genomic DNA of fish

Examples

Experimental program
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Effect test

Embodiment 1

[0039] The method for extracting the total genomic DNA of fish in this embodiment requires tools such as scissors and tweezers to be sterilized and burnt before proceeding, and the extracted samples must not be contaminated. The specific steps are as follows:

[0040] A. Take 1 piece of fresh adult grass carp scale, about 0.15g, rinse it with sterilized deionized water for 4 times, and rinse the impurities on the scale. Cut the transparent parts of fish scales into pieces and put them into a 1.5mL sterilized centrifuge tube.

[0041] B. Add 400 μL of 5% Chelex-100 solution to the centrifuge tube in step A, and bathe in boiling water for 15 minutes; then add 20 μL of proteinase K solution with a concentration of 20 mg / mL, and bathe in water at 55°C for 4 hours.

[0042] C. Take out the digested fish scales in the centrifuge tube, draw the liquid in the centrifuge tube and place it in another 1.5mL sterilized centrifuge tube, at a temperature of 4°C, at a centrifugation speed of...

Embodiment 2

[0051] The method for extracting the total genomic DNA of fish in this embodiment requires tools such as scissors and tweezers to be sterilized and burnt before proceeding, and the extracted samples must not be contaminated. The specific steps are as follows:

[0052] A. Take 2 pieces of adult grass carp scales soaked in 85% ethanol, about 0.2g, rinse 5 times with sterilized deionized water, and rinse the alcohol and impurities on the scales. Cut the transparent parts of fish scales into pieces and put them into a 1.5mL sterilized centrifuge tube. The black parts of fish scales have more impurities, so try not to get them.

[0053] B. Add 500 μL of Chelex-100 solution with a concentration of 5% to the centrifuge tube in step A, and bathe in boiling water for 20 minutes; then add 20 μL of proteinase K solution with a concentration of 20 mg / mL, bathe in water at 55°C for 5 hours, and add an appropriate amount of proteinase K solution midway.

[0054] C. Take out the digested fi...

Embodiment 3

[0063] The method for extracting the total genomic DNA of fish in this embodiment requires tools such as scissors and tweezers to be sterilized and burnt before proceeding, and the extracted samples must not be contaminated. The specific steps are as follows:

[0064] A. Take 2 pieces of adult grass carp scales soaked in 85% ethanol, about 0.2g, rinse 5 times with sterilized deionized water, and rinse the alcohol and impurities on the scales. Cut the transparent parts of fish scales into pieces and put them into a 1.5mL sterilized centrifuge tube. The black parts of fish scales have more impurities, so try not to get them.

[0065] B. Add 500 μL of Chelex-100 solution with a concentration of 5% to the centrifuge tube of step A, and bathe in boiling water for 15 minutes; then add 20 μL of proteinase K solution with a concentration of 20 mg / mL, bathe in water at 55°C for 3 hours, and add an appropriate amount of proteinase K solution midway.

[0066] C. Take out the digested fi...

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Abstract

The invention relates to an extraction method of total genomic DNA of fish. The extraction method comprises the following specific steps: A, taking the fish tissue containing DNA, washing, grinding, and placing into a centrifuging tube; B, adding with Chelex-100 solution, carrying out boiling water bath for 15-20 min, adding with proteinase K solution, and carrying out water bath at 50-60 DEG C for more then 3h; C, sucking liquid in the centrifuging tube, placing the liquid into another centrifuging tube, and carrying out centrifugal separation; D, taking the supernate, adding with RNase solution, carrying out water bath at 37 DEG C for more than 45 min; E, adding with Tris saturated phenol solution, uniformly mixing, carrying out centrifugal separation, and taking the supernate; F, adding with a suitable amount of chloroform and isoamylol mixed solution, uniformly mixing, carrying out centrifugal separation, and taking the upper solution; G, adding with alcohol, and placing for 1h or a longer time; H, carrying out centrifugal separation, and discharging the supernate; I, adding with alcohol, carrying out centrifugal separation, and discharging the supernate; J, removing the alcohol, adding with sterile water, and storing for later use. According to the extraction method, a reagent is convenient to fetch, the DNA extraction effect is good, the integrity of DNA is high, especially the extraction can be carried out by taking scale as the raw material, and limitation from the quantity of a sample is avoided.

Description

technical field [0001] The invention relates to a method for extracting genome DNA, which belongs to the field of biotechnology. Background technique [0002] Fish are the oldest vertebrates. They inhabit almost all aquatic environments on Earth. From freshwater lakes and rivers to saltwater seas and oceans. Fish is a temperature-changing vertebrate that lives in water all year round, breathes with gills, and assists body balance and movement with fins. There are currently about 32,100 named fish species in the world. It is the most primitive and lowest group in the vertebrate subphylum. Fish meat is rich in protein, calcium, phosphorus, iron, vitamin B1, lecithin and other nutrients, which can enhance memory, thinking and analysis ability, and delay mental decline, so it plays an important role in the development of human physical strength and intelligence. Fish is suitable for a variety of cooking methods, not only delicious, but also easily digested and absorbed by t...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/10
Inventor 毛慧玲胡成钰方春林万正义龚瑞玥王芳
Owner NANCHANG UNIV
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