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Screening method of scylla paramamosain SNPs molecular marker

A technology of mud crabs and molecular markers, which is applied in the field of genetic marker detection of mud crabs, can solve the problems of limited population genetic diversity evaluation molecular marker-assisted breeding, and achieve the effects of short experimental period, simple operation and low cost

Inactive Publication Date: 2013-04-17
EAST CHINA SEA FISHERIES RES INST CHINESE ACAD OF FISHERY SCI
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Problems solved by technology

At present, studies on mitochondrial DNA and microsatellite molecular markers have been carried out in Scylla syringae, but there are few studies on SNPs markers, which limits the research of population genetic diversity assessment, genetic map construction and molecular marker-assisted breeding. carry out

Method used

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  • Screening method of scylla paramamosain SNPs molecular marker
  • Screening method of scylla paramamosain SNPs molecular marker
  • Screening method of scylla paramamosain SNPs molecular marker

Examples

Experimental program
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Effect test

Embodiment 1

[0052] 1. Acquisition of Genomic DNA Sequence of Scylla pseudocarpus

[0053] In the early stage, our laboratory carried out the research on the sequencing of the transcriptome of the mud crab, from which the present invention randomly selects different sequencing sequences as the target sequences for screening SNPs.

[0054] 2. Sequencing and identification of candidate SNPs

[0055] Design PCR primers for the sequence selected above, and the expected size of the product is between 500bp and 900bp. Using the DNA of 5 different individuals as templates, PCR amplification was carried out. Send the PCR products with clear results, high yield and no non-specific amplification to the sequencing company for bidirectional and direct sequencing. The sequencing results were spliced ​​and compared to identify candidate SNPs.

[0056] 3. SNPs verification and genotyping

[0057] Three primers were designed for each candidate SNP site, including two allele-specific upstream primers (...

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Abstract

The invention relates to a screening method of a scylla paramamosain SNPs molecular marker, which comprises the following steps of: (1) obtaining a scylla paramamosain genome sequence through a genomic library or a GenBank database obtained by sequence determination; (2) carrying out primer design on the obtained genome sequence, carrying out sequence determination after PCR (Polymerase Chain Reaction) amplification, contrasting the sequences of different individuals and verifying candidate SNPs (Single Nucleotide Polymorphisms); and (3) designing a primer with a specific allete for a candidate SNPs site and carrying out verification and genetic typing on the SNPs by adopting a real-time fluorescent quantitative PCR dissolving curve method. The screening method has the advantages of short experimental period, low cost, simplicity in operation and the like, and the genetic types of the different individuals can be accurately detected to provide a novel genetic marker for scylla paramamosain genetic variation analysis, population genetic diversity evaluation and molecular marker-assisted breeding.

Description

technical field [0001] The invention belongs to the field of genetic marker detection of Scylla pseudomaculata, in particular to a screening method for SNPs molecular markers of Scylla pseudomaculata. Background technique [0002] Single nucleotide polymorphisms (Single Nucleotide Polymorphisms, SNPs) molecular markers exist in almost all eukaryotic genomes, and are the most abundant genetic variation resources in biological genomes. SNPs have typical biallelic nature, and there is one SNP in every 200bp~1000bp nucleotides in most biological genomes. Because of its many advantages, SNPs are considered to be the most potential genetic markers in the field of genetics and evolution. At present, SNPs molecular markers have been reported in a variety of aquaculture animals, such as: Litopenaeus vannamei, Japanese flounder, rainbow trout, Atlantic salmon, oyster, etc. [0003] Scylla paramamosain belongs to Arthropoda, Crustacea, Decapoda, Portunidae, and Scylla. It is mainly d...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/68
Inventor 马洪雨冯娜娜马凌波马春艳徐真
Owner EAST CHINA SEA FISHERIES RES INST CHINESE ACAD OF FISHERY SCI
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