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Screening method for adaptability SNP loci of Japanese eels

A screening method and adaptive technology, which are applied in the field of screening of adaptive SNP loci of Japanese eel, to achieve the effects of reducing library construction cost, reducing sequencing cost and consistent amplification efficiency

Inactive Publication Date: 2018-05-08
ZHEJIANG OCEAN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

It can be seen that different geographical populations of Japanese eel may have adaptive evolution to the local environment

Method used

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Experimental program
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Effect test

Embodiment 1

[0015] Embodiment 1: The screening method of Japanese eel adaptive SNP site, comprises the following steps:

[0016] 1) Genomic DNA extraction: Use phenol / chloroform extraction method to extract genomic DNA from Japanese eel muscle or fin ray tissue, the specific process is as follows: put the sample into a 1.5ml centrifuge tube and cut it into pieces, add 600μL STE (10mmol / L TriS- Cl, pH 8.0; 0.1mol / L EDTA, pH 8.0; 1%m / v SDS), shake evenly on a shaker, add RNase (4μg / μL) 6uL, final concentration 0.04μg / μL, incubate in a 37℃ oven During 1h, gently turn the centrifuge tube from time to time to mix the tissue with the extraction buffer evenly, then add 15μL of proteinase K (20μg / μL) with a final concentration of 0.5μg / μL, invert and mix well, and incubate in an oven at 50°C for 2h. Gently flip the centrifuge tube from time to time to mix the tissue with the extraction buffer evenly, then take it out of the oven and cool to room temperature, add 600 μL (pH 8.0) of equilibrated ph...

Embodiment 2

[0020] Embodiment 2: the screening method of Japanese eel adaptive SNP site, comprises the following steps:

[0021] 1) Genomic DNA extraction: Genomic DNA was extracted from Japanese eel muscle or fin ray tissue by phenol / chloroform extraction;

[0022] 2) RAD library construction and sequencing: the extracted genomic DNA was stored in ultrapure water, the dilution concentration was above 25ng / µL, the total mass of DNA was greater than 1µg, and the genomic DNA was digested with the restriction endonuclease EcoRI. The cutting reaction system was 50 μL (2 μL 10×NEB buffer, 0.5 μL endonuclease, 0.5 μL genomic DNA, and the rest was double distilled water). Add the first linker (complementary to the EcoRI cut DNA gap) to the fragment, mix the resulting fragments, randomly break into certain fragments (the average fragment length is about 500bp), and recover 200bp-400bp DNA by agarose gel electrophoresis Fragment, add 3'dA overhang after the end is blunt, and then add the second l...

Embodiment 3

[0025] Embodiment 3: the screening method of Japanese eel adaptive SNP site, comprises the following steps:

[0026] 1) Genomic DNA extraction: Genomic DNA was extracted from Japanese eel muscle or fin ray tissue by phenol / chloroform extraction;

[0027] 2) RAD library construction and sequencing: the extracted genomic DNA was stored in ultrapure water, the dilution concentration was above 25ng / µL, the total mass of DNA was greater than 1µg, and the genomic DNA was digested with the restriction endonuclease EcoRI. The cleavage reaction system was 50 μL (2 μL 10×NEB buffer, 0.5 μL endonuclease, 0.5 μL genomic DNA, and the rest was double distilled water). Before the reaction, add 0.2 ng tetrachloromandelic acid and 0.5 ng 2- Hydroxyphenylacetic acid, followed by reacting at 37°C for 10min, and then reacting at 72°C for 23min, the added tetrachloromandelic acid and 2-hydroxyphenylacetic acid have a synergistic effect with 10×NEB buffer, which improves the activity of the endonuc...

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Abstract

The invention discloses a screening method for adaptability SNP loci of Japanese eels. The method comprises the steps of extraction of a genome DNA, construction and sequencing of an RAD library, RADdata output and quality control and detection and screening of the SNP loci. The screening method for the adaptability SNP loci of the Japanese eels has the advantages that 37,700 SNP loci screened out through the method are widely distributed in the whole genome range of a Japanese eel sketch and totally contained in 10,710 contigs, each contig contains 3.5 SNP loci on average, genome informationresources of the Japanese eels which are endangered species are further supplemented and completed, and the important basis is provided for correlational research of the population genetic structureand population historical dynamic condition of the Japanese eels.

Description

technical field [0001] The invention relates to the field of molecular biology technology, in particular to a screening method for the adaptability SNP site of Japanese eel. Background technique [0002] The study of adaptive genetic differentiation is of increasing interest to researchers, especially in identifying and screening for molecular genetic markers of selection. Local adaptive evolution has drawn attention for the following reasons: First, in previous population genetics studies, only some neutral genetic data had been accumulated, and researchers realized that neutral variations detected in populations did not reveal The aspect of population adaptation evolution; secondly, under the current environment of global climate change, researchers are paying more and more attention to the biological microevolution caused by climate change. It is not easy to detect the adaptive differentiation of wild populations, especially the selection effect on spatial and temporal d...

Claims

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Application Information

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IPC IPC(8): C12Q1/6869G06F19/22
CPCC12Q1/6869G16B30/00C12Q2521/313C12Q2525/191C12Q2531/113
Inventor 刘炳舰刘立芹吕振明龚理柳意樊
Owner ZHEJIANG OCEAN UNIV
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