Nondestructive in-vivo squid DNA sample acquisition method and extraction agent

A sample collection, non-damaging technology, applied in the field of DNA collection, can solve the problems of lack of non-damaging live cephalopod DNA collection and individual identification methods, and achieve the effects of strong protein digestion, high DNA purity, and high molecular weight

Inactive Publication Date: 2018-05-18
ZHEJIANG OCEAN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Cephalopods are species that have been artificially propagated and developed in recent years. Objectively, it is necessary to establish live DNA sampling and identification techniques for broodstock breeding, establishment of family lines, and identification of proliferated and released individuals. However, there is still a lack of effective non-invasive methods. DNA collection and individual identification method of live cephalopods

Method used

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Examples

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Comparison scheme
Effect test

Embodiment 1

[0016] The invention discloses a non-damaging living squid DNA sample collection method, scraping tissue cells by rubbing the squid mantle cavity, and using them as samples to extract DNA.

[0017] The DNA extraction method is as follows: pretreat the tissue cells obtained by scraping, add lysis buffer and proteinase K to incubate to obtain the incubation solution; use the phenol-chloroform-isoamyl alcohol mixture to extract the incubation solution, and centrifuge to take the upper supernatant; add chloroform-isoamyl alcohol-petroleum ether mixture to the supernatant, centrifuge to take the supernatant after fully reacting; add absolute ethanol to the supernatant, centrifuge to take the precipitate after sufficient precipitation; add Ethanol solution, fully dissolved to obtain squid DNA, and stored at low temperature. The above-mentioned DNA extraction method is an improved CTAB method, which utilizes the strong denaturation and complexation properties of the lysis buffer, the...

Embodiment 2

[0025] The invention discloses a non-damaging living squid DNA sample collection method, scraping tissue cells by rubbing the squid mantle cavity, and using them as samples to extract DNA.

[0026] The DNA extraction method is as follows: pretreat the tissue cells obtained by scraping, add lysis buffer and proteinase K to incubate to obtain the incubation solution; use the phenol-chloroform-isoamyl alcohol mixture to extract the incubation solution, and centrifuge to take the upper supernatant; add chloroform-isoamyl alcohol-petroleum ether mixture to the supernatant, centrifuge to take the supernatant after fully reacting; add absolute ethanol to the supernatant, centrifuge to take the precipitate after sufficient precipitation; add Ethanol solution, fully dissolved to obtain squid DNA, and stored at low temperature. The above-mentioned DNA extraction method is an improved CTAB method, which utilizes the strong denaturation and complexation properties of the lysis buffer, the...

Embodiment 3

[0034] The invention discloses a non-damaging living squid DNA sample collection method, scraping tissue cells by rubbing the squid mantle cavity, and using them as samples to extract DNA.

[0035] The DNA extraction method is as follows: pretreat the tissue cells obtained by scraping, add lysis buffer and proteinase K to incubate to obtain the incubation solution; use the phenol-chloroform-isoamyl alcohol mixture to extract the incubation solution, and centrifuge to take the upper supernatant; add chloroform-isoamyl alcohol-petroleum ether mixture to the supernatant, centrifuge to take the supernatant after fully reacting; add absolute ethanol to the supernatant, centrifuge to take the precipitate after sufficient precipitation; add Ethanol solution, fully dissolved to obtain squid DNA, and stored at low temperature. The above-mentioned DNA extraction method is an improved CTAB method, which utilizes the strong denaturation and complexation properties of the lysis buffer, the...

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PUM

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Abstract

The invention discloses a nondestructive in-vivo squid DNA sample acquisition method. By friction of a squid mantle cavity, tissue cells are obtained through scraping to serve as a sample for DNA extraction. The DNA extraction method is an improved CTAB method, and by strong denaturation and complexation of pyrolysis buffer solution, phenol-chloroform-isoamyl alcohol repeated extraction and chloroform-isoamyl alcohol-petroleum ether reaction, high protein digestion capacity and removal of impurities including phenols, saccharides and the like with serious interferences on enzyme functions arerealized while high yield is achieved. In addition, high extracted DNA molecular weight is realized, evident degradation is avoided, nucleic acid completeness is kept, a measured OD value range is 1.6-1.9, and high DNA purity is realized.

Description

technical field [0001] The invention relates to the field of DNA collection, in particular to a method for collecting a DNA sample of a non-damaging live squid and an extraction reagent. Background technique [0002] Squid, whose real name is squid, also known as squid, cuttlefish or cuttlefish, is an animal of the mollusk phylum Cephalopoda squid. What is different from squid is that squid has a boat-shaped calcareous hard sheath. When squid encounters a strong enemy, it will use "ink jet" as a means of escape, waiting for an opportunity to leave, so it has names such as "squid" and "cuttlefish". Pigmented sacs in the skin that change color and size in response to 'moods'. Eggs are laid in late spring on wood chips or seaweed. Squids leap out of the water and have amazing aerial flight abilities. [0003] DNA sample collection is a very critical step for individual identification or related molecular biological analysis. In actual operation, DNA samples are generally ex...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/10C12Q1/6806
CPCC12N15/1003C12Q1/6806
Inventor 吕振明朱科桦龚理刘炳舰刘立芹
Owner ZHEJIANG OCEAN UNIV
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