Molecular marker method for identifying hybrid offspring of black sea bream and red sea bream by using ISSR (inter-simple sequence repeat) primer

A technology of molecular markers and progeny, applied in biochemical equipment and methods, microbiological determination/inspection, DNA/RNA fragments, etc., can solve problems such as no detection methods, to ensure rapid differentiation and application, and get rid of unreliable Factors, the effect of broadening the technical system

Pending Publication Date: 2022-04-15
INST OF OCEANOLOGY & MARINE FISHERIES JIANGSU
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AI Technical Summary

Problems solved by technology

So far, there is no report on the relevant detection methods for germplasm identification of black sea bream, backcross progeny and hybrid progeny

Method used

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  • Molecular marker method for identifying hybrid offspring of black sea bream and red sea bream by using ISSR (inter-simple sequence repeat) primer
  • Molecular marker method for identifying hybrid offspring of black sea bream and red sea bream by using ISSR (inter-simple sequence repeat) primer

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Embodiment 1

[0023] 1) Eight healthy individuals were randomly selected from the hybrid offspring (HF1) and backcross offspring (BF1) of black sea bream and red sea bream. Cut fish (or other body tissue) the size of a mung bean, cut it into pieces with scissors, and put it into a 1.5mL centrifuge tube; use the animal cell / tissue genomic DNA extraction kit to extract the total DNA, and store it at 4°C.

[0024] 2) ISSR primers: UBC899 in the ninth set of primers ISSR (http: / / www.biotech.ubc.ca / services / naps / primers / Primers.pdf) of the University of Columbia (UBC), the primer (UBC899 sequence: CAT GGT GTT GGT CAT TGT TCC A) Synthesized by Shanghai Sangong.

[0025] 3) UBC899 primers were used as ISSR primers to perform PCR amplification on the above-mentioned total DNA;

[0026] The PCR amplification system is: the total volume of the PCR reaction of 3 types of ISSR primers including black sea bream, HF1 and BF1 is 25μl, including: primer 0.5mmol / L, DNA template about 20ng, Buffer 2.5μl, dN...

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Abstract

The invention relates to a molecular marker method for identifying a black sea bream and red sea bream hybrid offspring by utilizing an ISSR (inter-simple sequence repeat) primer, which comprises the following steps of: performing PCR (polymerase chain reaction) amplification on individuals of black sea bream and two hybrid offspring new strains by utilizing an ISSR molecular marker, and distinguishing the individuals from the black sea bream and red sea bream hybrid offspring according to electrophoretic band detection of PCR products. The method disclosed by the invention is simple and convenient to operate, low in cost, high in accuracy and good in universality, can quickly and effectively identify a new hybrid strain obtained based on the parent combination of the black sea bream and the red sea bream, and provides technical help for new germplasm identification research of breeding research of new black sea bream hybrid varieties.

Description

technical field [0001] The invention belongs to the field of identifying black sea bream, in particular to a molecular marker method for identifying hybrid offspring of black sea bream and red sea bream by using ISSR primers. Background technique [0002] The basic principle of ISSR (inter-simple sequence repeat) molecular markers is: use the anchored microsatellite DNA as a primer, that is, add 2-4 random nucleotides to the 3' end or 5' end of the SSR sequence, In the reaction, the anchor primer can cause annealing at a specific site, resulting in PCR amplification of DNA fragments between repetitive sequences that are complementary to the anchor primer, and multiple bands of the amplified inter SSR region pass through the gel electrophoresis can be resolved. [0003] ISSR molecular marker technology has the characteristics of simple and fast operation, less DNA consumption, low technical requirements, and low cost. The primer design does not need to know the genome sequen...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/6888C12Q1/686C12N15/11
CPCC12Q1/6888C12Q1/686C12Q2600/158C12Q2525/151C12Q2565/125
Inventor 陈淑吟张志勇吉红九贾超峰孟乾祝斐徐大凤张志伟高波孙瑞健赵永超汤小建肖李霞周亚文
Owner INST OF OCEANOLOGY & MARINE FISHERIES JIANGSU
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