Molecular marker for identifying different ecotypic species of coilia ectenes

A molecular marker, eco-type technology, applied in the determination/inspection of microorganisms, biochemical equipment and methods, DNA/RNA fragments, etc., can solve the problem of scarcity of SINE molecular markers

Inactive Publication Date: 2014-10-01
SHANGHAI OCEAN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] Currently, there are very few SINE molecular markers developed for fish. SINE markers have not been reported yet

Method used

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  • Molecular marker for identifying different ecotypic species of coilia ectenes
  • Molecular marker for identifying different ecotypic species of coilia ectenes
  • Molecular marker for identifying different ecotypic species of coilia ectenes

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0025] Experimental Materials

[0026] According to the source of the samples, the anchovy is divided into 6 populations: Xiangshan Port (XS) in Zhejiang, Chongming in Shanghai (CM), Jingjiang in Jiangsu (JJ), Lake Taihu (TH), Poyang Lake in Jiangxi (PY), and Dongting Lake in Hunan (DT). The sample size of each population is 10 tails. The samples from Xiangshan Port in Zhejiang came from the aquatic product market, and the collection place is unknown, and the rest of the samples were caught on board.

[0027] Genomic DNA Extraction

[0028] The extraction of genomic DNA from the sample was performed according to the instructions of Shanghai Sangon’s UNIQ-10 Column Genomic DNA Extraction Kit. The quality and concentration of the extracted DNA were detected by an ultraviolet spectrophotometer, and stored at -20°C for later use.

[0029] site segregation

[0030] Primers were designed according to the conserved sequence of SINE, and the partial sequence of SINE in the an...

Embodiment 2

[0049] experimental method

[0050] The specific primer of SINE insertion site, its sequence is:

[0051] The upstream primer of the Locus 5 site is shown in SEQ ID NO.1,

[0052] The downstream primer of the Locus 5 site is shown in SEQ ID NO.2,

[0053] The upstream primer at the Locus 29 site is shown in SEQ ID NO.3,

[0054] The downstream primer at the Locus 29 site is shown in SEQ ID NO.4,

[0055] The upstream primer at the Locus 60 site is shown in SEQ ID NO.5,

[0056] The downstream primer at the Locus 60 site is shown in SEQ ID NO.6.

[0057] Through PCR amplification, agarose gel electrophoresis, cloning, and sequencing, the population-specific DNA fragments of the SINE locus were obtained, and different populations of the anchovy were identified according to the differences in the number and size of the amplified fragments.

Embodiment 3

[0059] experimental method

[0060] The specific primer of SINE insertion site, its sequence is:

[0061] The upstream primer of the Locus 5 site is shown in SEQ ID NO.1,

[0062] The downstream primer of the Locus 5 site is shown in SEQ ID NO.2.

[0063] Through PCR amplification, agarose gel electrophoresis, cloning, and sequencing, the population-specific DNA fragments of the SINE locus were obtained, and the sequencing results of the Locus 5 locus were analyzed. Three bands with a size of 650 bp could be amplified but not The samples that amplified bands of 350 and 280 bp are the anchovy from Xiangshan Port, Chongming, Jingjiang or Taihu Lake, and the samples that can amplify bands of 350 and 280 bp but cannot amplify the band of 650 bp are Poyang Lake or Anchovy in Dongting Lake.

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Abstract

The invention discloses a molecular marker for identifying different ecotypic species of coilia ectenes. Polymorphic loci of retrotransponsons SINE of coilia ectenes are selected as targets, and a specific primer is designed according to flanking sequences inserted into the loci; DNA segments of the SINE insertion or deletion loci are obtained by applying the methods such as PCR amplification, agarose gel electrophoresis, molecular cloning and the like, and different species of the coilia ectenes are identified by using the specific DNA segments. By adopting the molecular marker, the species of the coilia ectenes can be quickly and accurately determined, and the molecular marker has extensive use in the aspects such as molecular marker-assisted breeding of the coilia ectenes, artificial discharge effect assessment of fries, dynamic species monitoring and the like.

Description

technical field [0001] The invention relates to the technical field of molecular biology of fish, in particular, it relates to a molecular marker used for identifying different ecotype populations of the common anchovy. Background technique [0002] The definition of fish species involves two levels, one is the species level, and the other is the population level. Species identification methods include external morphological judgment method, biochemical method, and a molecular marker method developed in the past ten years. The morphological judgment method is often used for identification at the species level; for the identification at the population level, biochemical methods have been used, but this method is cumbersome and has a high rate of misjudgment, so it is basically no longer used and replaced by a molecular marker method. Because of the good stability of molecular markers, they are now widely used in theoretical studies such as fish population identification, po...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/68C12N15/11
CPCC12Q1/6888C12Q2600/156
Inventor 刘东唐文乔郭弘艺
Owner SHANGHAI OCEAN UNIV
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