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Application of AMO-miR-137 in preparation of broad-spectrum preparation for resisting white spot syndrome viruses

A white spot syndrome and preparation technology, applied in the direction of antiviral agents, applications, DNA / RNA fragments, etc., can solve the problems of unknown degradation difficulty, cumbersome steps, large molecular weight, etc., and achieve inhibition of further spread, inhibition of replication, Wide range of effects

Active Publication Date: 2019-06-21
SHANTOU UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The steps of the above-mentioned invention are cumbersome, or the required materials are various and the effect is slow
The molecular weight of the materials contained in it is relatively large, and the degree of difficulty of degradation is unknown. Whether its residue will affect human health is also unknown, and whether it will affect the ecological balance of breeding is also unknown.

Method used

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  • Application of AMO-miR-137 in preparation of broad-spectrum preparation for resisting white spot syndrome viruses
  • Application of AMO-miR-137 in preparation of broad-spectrum preparation for resisting white spot syndrome viruses
  • Application of AMO-miR-137 in preparation of broad-spectrum preparation for resisting white spot syndrome viruses

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0033] Detection test of miR-137 expression level in Scylla pseudocarpus after WSSV infection

[0034] The blue crabs weighing 30-40g and negative for WSSV by qRT-PCR were domesticated in an indoor seawater tank at a salinity of 10‰ and a water temperature of 25°C for one week, and then injected 200 μL from the fourth appendage of the crabs / only, 10 6 The number of copies of WSSV infection solution, blood cells were collected after infection 0h, 24h and 48h, and MagMAX TM mirVana TM Total RNAIsolation extraction kit was used to extract RNA, and then use PrimeScript TM II 1st Strand cDNA Synthesis kit was used for cDNA synthesis, the primers used were 5’-GTCGTATCCAGTGCAGGGTCCGAGGTCACTGGATACGACACGTGTAT-3’, and then the expression of miR-137 was detected by qRT-PCR, the primers used were P1 and P2.

[0035] P1:5'-CGCCGTTATTGCTTGAGA-3';

[0036] P2: 5'-TGCAGGGTCCGAGGTCACTG-3'.

[0037] see results figure 1 , after WSSV infection, the expression of miR-137 in the hemolym...

Embodiment 2

[0039] Experiment of miR-137 mimics up-regulating the expression of miR-137 in Scylla crab

[0040] According to the method of Example 1, the mud crabs were randomly divided into 3 groups, the experimental group was injected with 100 μL / miR-137 (10 nM), and the negative control group was injected with 100 μL / miR-137-scrambled (10 nM) or 100 μL / PBS respectively. . After 48 hours, the blood cells were collected and the expression level of miR-137 in the blood cells of the blue crab was detected according to the method of Example 1.

[0041] see results figure 2 , the results showed that injection of miR-137 mimics could significantly up-regulate the expression of miR-137 in mud crabs, which could be used for subsequent functional tests.

Embodiment 3

[0043] MiR-137 mimics promote the proliferation of WSSV in Scylla burrowing crabs

[0044] According to the method in Example 2, the pseudocave blue crabs were randomly divided into 3 groups, and the experimental group was injected with 100 μL / only 10 6 The mixed solution of WSSV infection solution with copy number and 100 μL / miR-137 (10 nM), and the negative control group was injected with 100 μL / only 10 6 The mixture of copy number of WSSV infection liquid and 100 μL / miR-137-scrambled (10nM) or 100 μL / 10 6 Infection solution of copy number WSSV. After 0h, 24h, and 48h, the appendage muscles of the blue crabs were cut, and the total DNA was extracted with a tissue gene DNA extraction kit (Tiangen), and then the virus copy number was detected by qRT-PCR. The primers used were P3 and P4.

[0045] P3: 5'-CAAATCTCCCCCTCATCTACTCAAC-3';

[0046] P4: 5'-AATAATTTTCCCGTTTCTGAATAGA-3'.

[0047] see results image 3 , the results showed that after the crabs were infected by WSSV, ...

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Abstract

The invention relates to application of AMO-miR-137 in preparation of a broad-spectrum preparation for resisting white spot syndrome viruses. The sequence of AMO-miR-137 for efficiently resisting WSSVis as follows: 5'-ACGUGUAUUCUCAAGCAAUAA-3'. The AMO-miR-137 provided by the invention can inhibit proliferation of WSSV in scylla paramamosain, penaeus japonicus and procambarus clarkii, remarkably improve the activity of blood lymphocytes of shrimps, enhance the resistance of the shrimps to viruses, and effectively inhibit transmission and infection of the WSSV viruses. The AMO-miR-137 disclosedby the invention has a wide application range and is not only suitable for marine cultured penaeus japonicus and scylla pseudocavernosa, but also suitable for freshwater cultured procambarus clarkii.The AMO-miR-137 is small in molecular weight and easy to degrade, does not destroy culture ecological balance, does not harm human health, and is environment-friendly. A potential medicine is provided for resisting the white spot syndrome virus in aquaculture, is not only suitable for prevention, but also suitable for a cultivation process of the shrimps and crabs with the outbreak of the white spot syndrome.

Description

technical field [0001] The invention relates to the field of prevention and treatment of viral diseases in aquaculture, in particular to the role of AMO-miR-137 in the preparation of anti-white spot syndrome virus inhibitors. Background technique [0002] Scylla paramamosain, referred to as blue crab, has the characteristics of fast growth, delicious meat, and high nutritional value. It is very popular in the seafood market. It is one of the important economic crabs in the southeast coast of my country, and it is also artificially cultivated in many countries. Major marine crab species. Japanese sac prawns (Marsupenaeus japonicus), commonly known as car prawns, are widely distributed, from the eastern waters of Africa, the Red Sea to Korea and Japan. In my country, they are mainly distributed in the East China Sea and South China Sea. It has the characteristics of delicious meat, high nutritional value, strong viability, etc., and is the most valuable variety of prawns. Pro...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/113A61K31/7088A61P31/20A23K20/153A23K20/195A23K50/80
CPCY02A40/81
Inventor 龚燚李升康林善梦
Owner SHANTOU UNIV
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