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A method and kit for detecting the expression of igf-1 alleles in Oni tilapia

A technology of Oni tilapia and IGF-1, which is applied in the field of biochemistry and molecular biology, can solve the problems of low detection efficiency, high cost, and high price of fluorescent quantitative reagents, and achieve the effect of low detection cost and easy operation

Active Publication Date: 2021-07-16
GUANGXI ACADEMY OF FISHERY SCI
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Problems solved by technology

However, these two methods have certain defects: AS-PCR method requires agarose gel electrophoresis detection, cumbersome operation, low detection efficiency, time-consuming and labor-intensive; TaqMan method is costly, and this method relies on fluorescent quantitative PCR, Fluorescent quantitative reagents are more expensive

Method used

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  • A method and kit for detecting the expression of igf-1 alleles in Oni tilapia
  • A method and kit for detecting the expression of igf-1 alleles in Oni tilapia
  • A method and kit for detecting the expression of igf-1 alleles in Oni tilapia

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Embodiment 1

[0037] Analysis of IGF-1 Allele Expression in 10 Tails of Oni Tilapia

[0038] 1. Preparation of materials

[0039] 10 Oni tilapia, PCR instrument, pyrosequencing instrument, pyrosequencing kit.

[0040] 2. Design and synthesis of primers

[0041] According to the sequence of IGF-1 in Nile tilapia and Oria tilapia, use the primer design software Primer Premier5.0 to design the specific primers and pyrophosphates for PCR amplification of IGF-1 allele expression in Oni tilapia The primers were sequenced, and the specific primer information was obtained as follows:

[0042] PS231-F: 5'-CAAGATTTCTCGCTCTGTGC-3';

[0043] PS231-BR: 5'-CGCTGTCCTACGCTCTGTC-3';

[0044] PS231-FS: 5'-ACCCAAGGTTAGTAGCAG-3'.

[0045] 3. RNA reverse transcription

[0046] 5 mg of tilapia liver was taken, crushed in a mortar after being snap-frozen in liquid nitrogen, and lysed with RNA trizol reagent to extract liver total RNA. The extracted RNA sample, reverse transcriptase, dNTP, Oligo(dT) 12-18 ...

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Abstract

The invention discloses a method and a kit for detecting the expression of IGF-1 alleles in tilapia oni, belonging to the fields of biochemistry and molecular biology. The detection method of the present invention comprises (1) RNA reverse transcription; (2) PCR amplification; (3) pyrosequencing reaction. The kit of the invention includes specific primers for PCR amplification and pyrosequencing primers. The method of the present invention can monitor the reaction in real time and perform quantitative and rapid detection, has the advantages of strong specificity, etc., brings convenience to the simple and rapid detection of IGF-1 allele expression in tilapia onis, and has low detection cost.

Description

technical field [0001] The invention belongs to the fields of biochemistry and molecular biology, and in particular relates to a method for detecting the expression of IGF-1 alleles in tilapia oni and a kit thereof. Background technique [0002] The growth of fish is regulated by growth hormone (GH) and insulin-like growth factor-1 (Insulin-like growth factor 1, IGF-1). GH is secreted by the pituitary gland, and by binding to its receptor (GHreceptor, GHR), it transmits a signal pathway to target cells, such as liver cells and muscle cells. Target cells activated by the GH signaling pathway secrete downstream hormones, such as IGF-1, to promote cell proliferation and animal growth. Due to its important growth-promoting role, IGF-1 has been extensively studied in aquatic animals. [0003] It has been reported that the hybrid offspring of hybrid brown-spotted grouper (Epinephelus fuscogutatus♀)×saddle-banded grouper (E.lanceolatus♂) has higher expression of IGF-1 in liver th...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/6888C12Q1/6869C12N15/11
CPCC12Q1/6869C12Q1/6888C12Q2600/124C12Q2600/158C12Q2531/113C12Q2565/301
Inventor 钟欢周毅罗永巨肖俊张孝瑾韩卓君于凡郑惠芳唐瞻杨郭忠宝
Owner GUANGXI ACADEMY OF FISHERY SCI
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