Microsatellite multi-PCR (Polymerase Chain Reaction) method for turbot paternity test

A paternity test, turbot technology, applied in biochemical equipment and methods, microbial determination/inspection, etc., can solve problems such as inability to high-throughput detection, large errors, etc., achieve low equipment requirements, improve efficiency and speed, A wide range of effects

Active Publication Date: 2010-11-10
YELLOW SEA FISHERIES RES INST CHINESE ACAD OF FISHERIES SCI
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0004] The purpose of the present invention is to construct a microsatellite multiplex PCR method for paternity identification of turbot, which is used for paternity identifica

Method used

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  • Microsatellite multi-PCR (Polymerase Chain Reaction) method for turbot paternity test
  • Microsatellite multi-PCR (Polymerase Chain Reaction) method for turbot paternity test
  • Microsatellite multi-PCR (Polymerase Chain Reaction) method for turbot paternity test

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Embodiment 1

[0033] Embodiment 1 (verify the accuracy of the present invention by the parent and progeny sample of known family lineage)

[0034] 1) DNA extraction from individual turbot

[0035] Collect 11 parents (3 parents missing) and 140 tail fin materials of corresponding offspring from 7 families of turbot, and use phenol-chloroform method to extract high-purity fin ray DNA: put a small amount of fin rays into a centrifuge tube and add DNA The extract solution TENS (10mM Tris-CI, pH8.0; 100mM EDTA, pH8.0; 100mM NaCl; 0.5% SDS) was digested by adding proteinase K and RNase for 2-3 hours, shaking several times to fully digest the fin rays, and then Add an equal volume of phenol / chloroform / isoamyl alcohol solution, mix until an emulsion is formed, centrifuge at room temperature, take the supernatant, and repeat the extraction twice. Add 2 volumes of ice-cold absolute ethanol to the supernatant and allow the DNA to precipitate for 10-30 minutes. The precipitate was rinsed with 70% ethan...

Embodiment 2

[0105] Example 2 Practical application of the multiplex PCR system in family paternity testing (pedigree analysis and paternity testing of individuals of unknown origin)

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Abstract

The invention relates to a microsatellite multi-PCR (Polymerase Chain Reaction) for a turbot paternity test, which comprises the steps of: extracting DNA from an individual turbot, screening pleomorphic microsatellite markers of the turbot, synthesizing a primer, constructing turbot multi-PCR systems and carrying out genealogy and paternity tests. In the method, two multi-PCR systems are established for the turbot genealogy and paternity tests, the steps of multi-PCR amplification, electrophoresis and silver staining are carried out on an individual to be tested so as to acquire microsatellite marker gene type data, cluster analysis for a genealogy test is carried out by adopting NTsys sofeware, and the paternity test is carried out by adopting Cervus2.0 software. The invention realizes that four microsatellite sites are simultaneously detected in one PCR, thus compared with the traditional PCR method, the efficiency of the method is improved by around 4 times, and the method can be subjected to the field test in a breeding base. The invention has the characteristics of high efficiency, economy, simplicity, convenience, easy operation and the like and can be popularized and applied in the aspects of germplasm identification, genealogy management and selective breeding of fine breeds for the turbots.

Description

technical field [0001] The invention belongs to the germplasm identification technology of aquatic animals in the field of aquaculture, and in particular relates to a microsatellite multiple PCR method for parentage identification of cultured turbot turbot, which is suitable for germ plasma identification and family management in turbot breeding and breeding . Background technique [0002] Turbot, which is called "Turbofish" in China, is a rare and precious flounder native to the coast of Europe. It has the advantages of fast growth, good meat quality, great breeding and market potential, and has become an excellent seawater cultured fish developed by various countries. one of the classes. Since it was introduced into my country in 1992, the turbot aquaculture industry has developed rapidly in my country, and it has now become an important fish culture species in the northern coast of my country. However, since my country is not the origin of turbot, the medium and long-te...

Claims

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Application Information

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IPC IPC(8): C12Q1/68
Inventor 陈松林苗贵东杜民杨景峰徐营
Owner YELLOW SEA FISHERIES RES INST CHINESE ACAD OF FISHERIES SCI
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