IRAK-b gene of mytilus crassitesta and use of gene

A thick-shelled mussel and genetic technology, applied in the field of genetic engineering, can solve problems such as limited research, achieve strong recognition and induction capabilities, convenient operation, and reasonable design effects

Pending Publication Date: 2019-12-10
ZHEJIANG OCEAN UNIV
View PDF2 Cites 0 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, there is limited research on the response of this species to pathogen challenge

Method used

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
View more

Image

Smart Image Click on the blue labels to locate them in the text.
Viewing Examples
Smart Image
  • IRAK-b gene of mytilus crassitesta and use of gene
  • IRAK-b gene of mytilus crassitesta and use of gene
  • IRAK-b gene of mytilus crassitesta and use of gene

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0031] The IRAK-b gene of the thick-shelled mussel was obtained by the following method:

[0032] 1. Total RNA extraction;

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

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

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

[0036] (4) Use a pipette to repeatedly suck and suck the precipitate to resuspend it and leave it at room temperature for 5 minutes;

[0037] (5) Add 200 μL of chloroform, cover the centrifuge tube tightly, shake vigorously by hand for 15 seconds, and leave it 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 solution into another centrifuge tube, add 500μL of isopropanol, and mix gently;

[0040] (8) Place the mixed solution at -20°C for 30 min;

[0041] (9) Centrifuge at 1200...

Embodiment 2

[0102] Tissue-specific expression of IRAK-b gene in mussel

[0103] Collect 5 kinds of tissues (adductors, hemolymph, gills, gonads, and hepatopancreas) of mussels with thick shells, and extract total RNA from the collected samples according to the recommended method of Trizol Total RNA extraction kit from TaKaLa Proceed (refer to Example 1 for total RNA extraction for specific steps). Then use M-MLV (RNAase H - ) Reverse transcription kit, using total RNA for cDNA synthesis. According to the obtained IRAK-b gene sequencing results of the thick-shelled mussel, Primer 5.0 software was used to design the primers required for the IRAK-b gene fluorescence quantitative PCR. When designing primers, it is required that the annealing temperature of the forward and reverse primers is within 1℃, and the Tm is above 60℃. There is no complementation between the primers, and no primer dimer is formed during the PCR reaction. The primers were screened by gradient PCR and sequenced, and fina...

Embodiment 3

[0109] Expression of IRAK-b gene in the gill of the mussel Thick shell mussel after pathogen infection

[0110] According to the tissue-specific results of IRAK-b gene, gills were selected as candidate tissues to study the time expression profiles of McIRAK-b and McIRAK-a after infection. The specific method is as follows:

[0111] Choose healthy thick-shell mussels and keep them temporarily for two days at a breeding density of 40 / 21L, during which oxygenation is continued. Spirulina powder was fed once every morning and evening, and the water was changed 1 / 2 every 24h (before feeding at night), and the water was artificially configured seawater with a salinity of 30‰. Then the thick shell mussels were randomly divided into 3 groups, 30 in each group, and the adductor muscles of each group were injected with PBS (pH 7.4) resuspended Vibrio alginolyticus ( Vibrio parahemolyticus ) And Vibrio parahaemolyticus ( Vibrio Parahemolyticus ), the dose was 0.1 mL / head, the control grou...

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to view more

PUM

No PUM Login to view more

Abstract

The invention provides an IRAK-b gene of mytilus crassitesta and use of the gene, and belongs to the technical field of gene engineering. The nucleotide sequence of the gene is shown in SEQ ID NO: 1,with a total length of 1815 bp, the gene includes an ORF sequence of 1599 bp, encodes 532 amino acids, and plays an important role in immune signal pathway of mytilus crassitesta, and a correspondingimmune signal protein can be quickly synthesized to resist the invasion of external stressors on body self. Under the infection of Vibrio alginolyticus or Vibrio parahaemolyticus, the expression levelof the IRAK-b gene of mytilus crassitesta is up-regulated, and the gene can be used for preparing reagents for detecting the pathogen infection of mytilus crassitesta. According to the present invention, the amino acid sequence of the protein encoded by the IRAK-b gene of mytilus crassitesta is shown in SEQ ID NO: 2, and the IRAK-b gene of mytilus crassitesta can be used to produce a recombinantprotein of the IRAK-b gene of mytilus crassitesta, and the protein is used to prepare a composition for improving the immunity of marine molluscs.

Description

Technical field [0001] The invention belongs to the technical field of genetic engineering, and specifically relates to a thick-shelled mussel IRAK-b gene and uses thereof. Background technique [0002] The immune system (Immunity system) is an immune defense system formed by long-term evolution and evolution of organisms, including the innate immune system and the adaptive immune system. The innate immune system is the body's first natural barrier against the invasion of pathogenic microorganisms. It is regulated by genetic factors and is relatively stable. It has immune protection against many types of pathogens (such as parasites, bacteria, and viruses). Pattern recognition receptors (PRRs) play an important role in the innate immune response, and can specifically recognize pathogen-associated molecular patterns (PAMPs), thereby activating the innate immune system and activating the adaptive immune system , Trigger an immune response against the identified pathogen. Pathogen...

Claims

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to view more

Application Information

Patent Timeline
no application Login to view more
IPC IPC(8): C12N15/54C12N9/12C12Q1/6888A61K38/45A61P37/04
CPCC12N9/12C12Y207/10002C12Q1/6888A61P37/04C12Q2600/158A61K38/00Y02A50/30
Inventor 刘慧慧葛德隆张立
Owner ZHEJIANG OCEAN UNIV
Who we serve
  • R&D Engineer
  • R&D Manager
  • IP Professional
Why Eureka
  • Industry Leading Data Capabilities
  • Powerful AI technology
  • Patent DNA Extraction
Social media
Try Eureka
PatSnap group products

Browse by: Latest US Patents, China's latest patents, Technical Efficacy Thesaurus, Application Domain, Technology Topic.

© 2024 PatSnap. All rights reserved.Legal|Privacy policy|Modern Slavery Act Transparency Statement|Sitemap