Primer sequence and method for rapid identification of pseudosciaena crocea, larimichthys polyactis and collichthys lucidus

A primer sequence, technology of large yellow croaker, applied in biochemical equipment and methods, microbial determination/inspection, DNA/RNA fragments, etc., can solve the problems of breeding, production and sales confusion, scarcity of wild large yellow croaker, and high price. The effect of shortening the technical cycle, the identification method is simple and fast, and the steps are simple

Inactive Publication Date: 2016-11-09
INST OF OCEANOLOGY & MARINE FISHERIES JIANGSU
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Problems solved by technology

[0004] At present, due to the sharp decline in natural resources, wild large yellow croakers are extremely scarce, and most of the products sold in the market are artificially farmed products; while small yellow croakers and plum boy fish are mostly wild products, and the price of large individuals is higher
It is n

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  • Primer sequence and method for rapid identification of pseudosciaena crocea, larimichthys polyactis and collichthys lucidus
  • Primer sequence and method for rapid identification of pseudosciaena crocea, larimichthys polyactis and collichthys lucidus

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Embodiment 1

[0026] (1) Sample DNA was extracted and dissolved in double distilled water (100ng / ul), and stored at 4°C for later use.

[0027] (2) PCR amplification conditions

[0028] The numbers and sequences of the specific primers are:

[0029] Primer 1F sequence: 5'-GGAAAGAGCCAGGAAAGC;

[0030] Primer 1R sequence: 5'-GGCGGAACTCTGAGCAAA. (design according to EST sequence in GenBank)

[0031] The total volume of the PCR reaction is 25 μL, 10×Buffer 2.5 μl, dNTP 0.5 μl, Mg 2+ 1.5 μl, Taq enzyme 0.25 μl, each primer 1 μl (0.5 mmol / L), sample DNA template 1 μl, dH 2 O 17.25 μl.

[0032] The PCR amplification parameters were as follows: 94°C pre-denaturation for 5 minutes followed by 35 cycles: 94°C for 40 sec, 58°C for 40 sec, 72°C for 1 min; and finally 72°C for 10 min.

[0033] (3) Electrophoresis detection and map analysis of PCR amplification

[0034] Take 8 μL of PCR amplification products for detection on the electrophoresis instrument, the electrophoresis buffer is 0.5×TBE, t...

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Abstract

The invention relates to a primer sequence and a method for rapid identification of pseudosciaena crocea, larimichthys polyactis and collichthys lucidus. The primer sequence is as follows: 1F:5'-GGAAAGAGCCAGGAAAGC-3' and 1R:5'-GGCGGAACTCTGAGCAAA-3'. The method includes the steps of (1) extracting individual genomic DNA; (2) synthesizing a specific primer for sequence amplification and performing PCR (Polymerase Chain Reaction) amplification; and (3) detecting the PCR product through agarose gel electrophoresis, taking pictures and performing recording, obtaining respective specific DNA fragment spectrograms of the three kinds of fishes according to different migration distance of the amplification products, and performing distinguishing in accordance to attached spectrograms. The method can simply and accurately distinguish individual samples of the three kinds of fishes, and provides a new fast research method for germplasm identification of the three kinds of economic fishes with similar appearance.

Description

technical field [0001] The invention belongs to the field of molecular marker identification, in particular to a primer sequence and a method for rapidly identifying large yellow croaker, small yellow croaker and spiny-headed plum boy. Background technique [0002] Large yellow croaker (Pseudosciaena crocea Richardson 1846), belonging to Perciformes, Sciaenidae, Larimichthys, commonly known as yellow croaker, yellow croaker, etc., is mainly distributed in the southern Yellow Sea, the East my country Sea and the South China Sea. Warm-temp offshore group migratory fish, generally 20-40cm in length, the maximum can reach 75cm. Large yellow croaker is rich in protein, trace elements and vitamins, and has high economic value. It was once one of the four major marine economic fish in my country. [0003] The small yellow croaker (Larimichthys polyactis) is delicious, and the spiny head plum fish (Collichthyslucidus, commonly known as plum fish) is tender and soft. These two kinds ...

Claims

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Application Information

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IPC IPC(8): C12Q1/68C12N15/11
CPCC12Q1/686C12Q1/6888C12Q2565/125
Inventor 陈淑吟王思婷张志勇李鹏贾超峰祝斐尹绍武张曹进吴国均吴磊
Owner INST OF OCEANOLOGY & MARINE FISHERIES JIANGSU
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