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Method for detecting C135T single nucleotide polymorphism mark of turbot

A technology for single nucleotide polymorphism and detection method, which is applied in the field of detecting turbot C135T single nucleotide polymorphism markers, can solve problems such as cumbersome operation, and achieves simple operation, low cost, high-throughput closed tube effect of operation

Inactive Publication Date: 2013-07-17
YELLOW SEA FISHERIES RES INST CHINESE ACAD OF FISHERIES SCI
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Problems solved by technology

However, when they analyze EST sequences, the software they use is CAP3 and QualitySNP. The operation of this software requires the Linux operating system, and the operation is relatively cumbersome.

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  • Method for detecting C135T single nucleotide polymorphism mark of turbot
  • Method for detecting C135T single nucleotide polymorphism mark of turbot
  • Method for detecting C135T single nucleotide polymorphism mark of turbot

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Embodiment Construction

[0024] A method for detecting single nucleotide polymorphism markers of turbot C135T, comprising 1), extraction of genomic DNA; 2), screening of candidate SNPs; 3), design of primers and probes; 4), asymmetric PCR amplification 5), probe hybridization; 6), data acquisition; 7), genotyping, the specific steps are as follows:

[0025] 1. Genomic DNA extraction:

[0026]The samples in this experiment came from 42 individuals of 10 different trait groups in 8 families of Yantai Tianyuan Aquatic Products Co., Ltd., and the DNA was extracted by using the marine animal tissue genomic DNA extraction kit (spin column type, TIANGEN). a. Take 20 mg of fresh muscle tissue, put it into a centrifuge tube filled with 200 μL GA buffer, and vortex for 15 seconds. b. Add 20 μL of proteinase K (20 mg / ml) solution, vortex to mix, and centrifuge briefly to remove water droplets on the inner wall of the tube cap. Place at 56°C until the tissue is completely dissolved, and briefly centrifuge to re...

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Abstract

The invention discloses a method for detecting C135T single nucleotide polymorphism (SNP) of turbot. The method comprises the following steps of: extracting genome DNA of turbot and diluting for later use; analyzing a sequence of an expressed sequence tag (EST), screening a sequence containing a candidate SNP locus, designing specific primers at two ends of the sequence, and designing 3' end-closed unlabeled probes before and after the locus (including the locus); performing asymmetrical polymerase chain reaction (PCR) amplification on population genome DNA of turbot by using the primers; hybridizing an amplification product and the 3' end-closed unlabeled probes; and detecting a melting curve of the hybrid on a light scanner, and analyzing the melting curve to obtain a genetic polymorphism map of turbot. The method is suitable for detection technologies such as turbot population genetic markers, lineage authentication, genetic linkage map construction and the like; and by the method,a genetic variation map of the C135TSNP of turbot can be quickly, easily and accurately obtained, and the genotype of each individual of turbot can be intuitively detected.

Description

Technical field: [0001] The invention belongs to molecular genetic marker research technology, in particular to a method for detecting turbot C135T single nucleotide polymorphism marker (single nucleotide polymorphism, SNP). Background technique: [0002] Single nucleotide polymorphism (SNP) molecular marker is the most abundant mutation type in the genome, because of its large number, high coverage density, rich loci, almost distributed in the entire genome, with short fragments, easy amplification, and genetic stability Strong and other characteristics; easy to high-throughput, automated analysis, easy to study genetic mechanisms and other advantages, so it is a hot field in genetic breeding research. Regarding the research on the development of turbot SNP markers, before the present invention was made, there were some relevant reports at home and abroad, and the poplar candidate based on EST sequence published by Zhang Xinye, etc. SNPs marker analysis has reported a meth...

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/68
Inventor 马爱军刘庆明黄智慧王新安杨志曲江波商晓梅
Owner YELLOW SEA FISHERIES RES INST CHINESE ACAD OF FISHERIES SCI
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