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Large yellow croaker scavenger receptor SCARA3 gene and application thereof

A scavenger receptor, large yellow croaker technology, applied in the field of genetic engineering, can solve the problems of economic loss, threat to large yellow croaker breeding, high mortality, etc., and achieve the effects of rapid response, reasonable design and convenient operation

Pending Publication Date: 2019-10-22
ZHEJIANG OCEAN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, with the rapid development of the aquaculture industry and the continuous expansion of the aquaculture scale, the problem of disease has become increasingly prominent. Among them, vibriosis caused by Vibrio Harvey is a relatively serious bacterial disease. It shows complex and diverse symptoms due to different parts, and causes high mortality, which has seriously threatened the cultivation of large yellow croaker and easily caused huge economic losses.

Method used

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  • Large yellow croaker scavenger receptor SCARA3 gene and application thereof
  • Large yellow croaker scavenger receptor SCARA3 gene and application thereof
  • Large yellow croaker scavenger receptor SCARA3 gene and application thereof

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0031] The complete ORF of the large yellow croaker scavenger receptor SCARA3 gene was obtained by the following method:

[0032] S1: Total RNA was extracted from the collected 8 tissue samples including liver, spleen, brain, heart, head kidney, muscle, skin, gills and intestine;

[0033] (1) Take 30mg of tissue samples in a 1.5mL centrifuge tube;

[0034] (2) Add 200 μL Trizol Regent to the centrifuge tube;

[0035] (3) Add 800 μL Trizol Regent after rapid and thorough grinding with an electric grinding rod;

[0036] (4) Blow and suck the precipitate repeatedly with a pipette gun to resuspend it, and place it at room temperature for 5 minutes;

[0037] (5) Add 200 μL of chloroform, cap the centrifuge tube tightly, shake vigorously by hand for 15 seconds, and place at room temperature for 10 minutes;

[0038] (6) Centrifuge at 12000×g for 15 minutes at 4°C to separate the solution into upper, middle and lower layers;

[0039] (7) Carefully pipette the upper aqueous phase sol...

Embodiment 2

[0082] Tissue-specific distribution and expression of scavenger receptor SCARA3 gene in large yellow croaker

[0083] Firstly, 9 tissues of liver, spleen, brain, heart, head kidney, muscle, gills, intestine and skin of healthy large yellow croaker (body length 20-30cm, weight 350-400g) were taken, according to the sequencing of the scavenger receptor SCARA3 gene As a result, fluorescent quantitative PCR primers (as shown in Table 1) were designed, and qRT-PCR method was used to analyze the tissue differential expression distribution of SCARA3 gene in various tissues with large yellow croaker β-actin as an internal reference. The 20μL amplification reaction system is as follows:

[0084]

[0085] After mixing and centrifuging, they were placed on a fluorescent quantitative PCR instrument and amplified by a two-step method: pre-denaturation at 95°C for 4 minutes; denaturation at 95°C for 10 seconds, extension at 60°C for 45 seconds, 40 cycles, and finally extension at 72°C for ...

Embodiment 3

[0089] Expression of Scavenger Receptor SCARA3 Gene of Large Yellow Croaker in Infection of Vibrio harvelii

[0090] Experimental large yellow croakers (body length 20-30cm, body weight 350-400g) were kept temporarily in clean seawater at 25°C for 1 week, and fresh seawater was changed every day. Subsequently, the large yellow croakers were randomly divided into two groups, 30 in each group, and the experimental group was intraperitoneally injected with 100 μL of PBS-resuspended Vibrio harveylius (pH 7.4, 1×10 8 CFU / mL), the control group was injected with 100 μL PBS (pH 7.4). Six tissues of liver, spleen, brain, head kidney, muscle, and skin were collected at 0h, 2h, 6h, 12h, 24h, 48h, and 72h after injection, and total RNA was extracted. The expression levels of the large yellow croaker scavenger receptor SCARA3 gene at 0, 6, 12, 24, 48 and 72 hours after infection with Vibrio harveyi were detected by fluorescent quantitative PCR. The primers, reagents and methods used were...

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Abstract

The invention provides a large yellow croaker scavenger receptor SCARA3 gene and application thereof, and belongs to the technical field of genetic engineering. The complete ORF of the large yellow croaker scavenger receptor SCARA3 gene includes 1938 basic groups, and the nucleotide sequence is shown as SEQ ID NO:1; under the infection of vibrio harveyi, the expression quantity of the gene is up-regulated, and the gene can be applied to detecting the vibrio harveyi resistance of large yellow croaker parents and / or breeding the vibrio-harveyi-resistant fine species and especially can be appliedto detecting the vibrio harveyi resistance of the large yellow croaker parents and / or breeding the vibrio-harveyi-resistant fine species. Recombinant expression protein of large yellow croaker scavenger receptor protein coded by the large yellow croaker scavenger receptor SCARA3 gene has remarkable vibrio-harveyi-resistant activity and can be used for preparing vibrio-harveyi-resistant biologicalproducts, and a gene sequence and method are provided for the research on the large yellow croaker vibrio-harveyi-resistant mechanism and the development of vibrio-harveyi-resistant feed additives.

Description

technical field [0001] The invention belongs to the technical field of genetic engineering, and in particular relates to a large yellow croaker scavenger receptor SCARA3 gene and its application. Background technique [0002] Large yellow croaker (Larimichthys crocea) belongs to Perciformes (Perciformes), Sciaenidae (Sciaenidae), and yellow croaker (Larimichthys), commonly known as "yellow croaker", "cucumber fish" and "big yellow croaker". Important fish species. Large yellow croaker has a golden body color, bright red lips, tender meat, delicious taste and high nutritional value. As my country's traditional "four seafood" and traditional "national fish", it occupies an important position in my country and the western Pacific ocean fishery. However, with the rapid development of the aquaculture industry and the continuous expansion of the aquaculture scale, the problem of disease has become increasingly prominent. Among them, vibriosis caused by Vibrio Harvey is a relative...

Claims

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Application Information

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IPC IPC(8): C12N15/12C12N15/10C07K14/46C12Q1/6888A61K38/17A61P31/04
CPCC07K14/461C12Q1/6888A61P31/04C12Q2600/124C12Q2600/158A61K38/00
Inventor 刘慧慧刘思佳
Owner ZHEJIANG OCEAN UNIV
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