Method utilizing microsatellite markers to identify polycultured Songpu mirror carp family

A microsatellite marker and polyculture technology, which is applied to the determination/inspection of microorganisms, biochemical equipment and methods, etc., can solve the problems of allelic differences, the inability of microsatellite markers to be universal and unsuitable for application, and achieve the effect of short cycle.

Inactive Publication Date: 2015-10-28
HEILONGJIANG RIVER FISHERY RES INST CHINESE ACADEMY OF FISHERIES SCI
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Problems solved by technology

However, in order to achieve stable appearance and target traits, farmed fish must undergo a certain degree of inbreeding and multiple generations of high selection pressure selection during the breeding process of new species. Many polymorphic sites are in this process. Homozygous, alleles are lost to varying degrees; moreover, selection and drift can cause the alleles of a microsatellite marker to vary greatly among different species or strains of the same fish species
Therefore, the microsatellite markers of fish cannot be used universally, and specific DNA markers must be selected for species or strains for genetic relationship identification
The research results also confirmed this point. Many marker sites with abundant polymorphisms in the German mirror carp breeding line (F4) or other carp species have been homozygous or have low polymorphism in Matsuura mirror carp, which is not suitable for Application in Pedigree Identification of Songpu Mirror Carp

Method used

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  • Method utilizing microsatellite markers to identify polycultured Songpu mirror carp family
  • Method utilizing microsatellite markers to identify polycultured Songpu mirror carp family
  • Method utilizing microsatellite markers to identify polycultured Songpu mirror carp family

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Experimental program
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specific Embodiment approach 1

[0015] Specific implementation mode one: this implementation mode identification polyculture Matsuura mirror carp family is carried out according to the following steps:

[0016] 1. Extract the individual genome DNA of the parents and offspring of each family of Matsuura mirror carp, and use the DNA of the parents and offspring as templates, and perform multiple PCR reactions with microsatellite marker primers;

[0017] 2. Load the PCR product of step 1 on a 10% polyacrylamide gel, perform polyacrylamide gel electrophoresis, and develop the color by silver staining. The electrophoretic pattern is analyzed by Gel-pro analyzer4.5 software, combining the length of the PCR product and the four bases The repeat unit confirms the size of the gene fragment and determines the genotype;

[0018] Three, set up the genotype data file of polyculture Matsuura mirror carp parent and progeny according to the analysis result of step 2;

[0019] 4. Use Cervus3.0 software to calculate the alle...

specific Embodiment approach 2

[0024] Embodiment two: the difference between this embodiment and embodiment one is: in step one, 9 pairs of microsatellite marker primers are divided into 4 groups to carry out multiple PCR reactions; the first group of PCR reactions consists of microsatellite marker 1, microsatellite marker 2 The primer pair of microsatellite marker 3 is carried out; the second group of PCR reactions is carried out by the primer pair of microsatellite marker 4 and microsatellite marker 5; the third group of PCR reactions is carried out by the primer pair of microsatellite marker 6 and microsatellite marker 7; The fourth set of PCR reactions was performed with primer pairs of microsatellite marker 8 and microsatellite marker 9 . Other steps and parameters are the same as those in Embodiment 1.

Embodiment 1

[0026] In 2013, 14 healthy males and 37 female broodstock were selected from the core population of Matsupu mirror carp in the Hulan Experimental Center of the Heilongjiang Fisheries Research Institute of the Chinese Academy of Fishery Sciences. Males mate with 2 to 3 females, and 37 full-sib families are established through artificial insemination. Collect fin ray samples of broodstock, record sex at the same time, and hatch fertilized eggs with different mating combination numbers separately. The offspring of Matsuura mirror carp were always cultured in the same pond and under the same conditions. At the 60th day of growth, all offspring of Matsuura mirror carp (1318 in total) were electronically marked and recorded, and fin rays were collected for genetic relationship identification.

[0027] 1. Extraction of individual genomic DNA from parents and offspring of Matsuura mirror carp:

[0028] (1) Take the fin ray tissue (about 60 mg per tail) of Matsuura mirror carp parent ...

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Abstract

The invention discloses a method utilizing microsatellite markers to identify polycultured Songpu mirror carp family, and relates to a method utilizing microsatellite markers to identify mirror carp family. The method for identifying polycultured Songpu mirror carp family includes the following steps: I, extracting DNA of parent and polycultured offspring individuals and performing multiplex PCR; II, performing gel electrophoresis to the PCR product to confirm the size of gene segments and genotypes; III, establishing genotype data files; IV, calculating allele frequency of all the loca to obtain related genetic parameters and integrated exclusion probability of all the loca; V, calculating the analytical procedure through utilizing the parents provided by the software, and confirming the source of the offspring individual family according to the calculated LOD value of candidate parents. The microsatellite marker combination and method provided by the invention can be used for identifying the existing Songpu mirror carp family.

Description

technical field [0001] The invention relates to a method for identifying carp family by using microsatellite markers. Background technique [0002] Matsuura mirror carp is a new species of carp bred by the Heilongjiang Fisheries Research Institute on the basis of the German mirror carp breeding line (F4). If the generation is not indicated, it is not a new variety of Matsuura mirror carp). The growth rate and meat yield of this fish are much higher than other carp of the same species, and it has fewer scales and is easy to process. It has been promoted to more than 20 provinces and cities across the country for breeding, bringing significant economic and social benefits. However, on the other hand, Matsuura mirror carp, like other carps, has low heritability for some important traits such as disease resistance, overwintering weight loss rate, carcass quality, etc., and it is difficult to obtain ideal genetic progress by population selection. The breeding practice of a larg...

Claims

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Application Information

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IPC IPC(8): C12Q1/68
CPCC12Q1/6888C12Q1/6827C12Q2600/156
Inventor 胡雪松石连玉李池陶贾智英
Owner HEILONGJIANG RIVER FISHERY RES INST CHINESE ACADEMY OF FISHERIES SCI
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