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Protein expression system regulated by arabinose-bacteria density binary signal and its application

A technology of arabinose and bacteria, applied in the field of protein expression system

Active Publication Date: 2020-08-04
EAST CHINA UNIV OF SCI & TECH +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] The components in the quorum sensing and quorum quenching circuits of bacteria play an important role in many artificial gene circuits and realize many biological functions. Researchers have rearranged the quorum sensing system itself or coupled it with other genes to construct Many complex man-made systems have been developed, but most of these systems only use the system to reproduce natural phenomena or simulate functions, and few of them are actually applied to production and life.

Method used

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  • Protein expression system regulated by arabinose-bacteria density binary signal and its application
  • Protein expression system regulated by arabinose-bacteria density binary signal and its application
  • Protein expression system regulated by arabinose-bacteria density binary signal and its application

Examples

Experimental program
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Effect test

Embodiment 5

[0095] Example 5. The "arabinose-bacteria density" binary system araQS is used as a tracer system in J774 macrophages

[0096] Animals and cells are typical environments lacking arabinose. After the verification of the previous examples, araQS will automatically turn on protein expression after entering the environment lacking arabinose in cells and proliferating.

[0097]J774 is a mouse mononuclear macrophage, isolated and screened from reticulocyte sarcoma of BALA / C mice, and participates in nonspecific and specific immunity in vertebrates. It can phagocytize or digest intracellular debris and pathogens in the form of fixed cells or free cells, and activate other immune cells to respond to pathogens. Edwardsiella tarda is a typical facultative intracellular parasite, which invades the epithelium and macrophages of the host body and expands therein, leading to systemic infection and hemorrhagic sepsis. Therefore, when Edwardsiella tarda containing the araQS expression system...

Embodiment 6

[0100] Example 6. Detection of the expression of the "arabinose-bacteria density" binary system araQS in zebrafish

[0101] Zebrafish is a small (4-6cm in body length) tropical freshwater fish of Cyprinidae. Cell labeling technology, tissue transplantation technology, mutation technology, haploid breeding technology, transgenic technology, and gene activity inhibition technology of zebrafish have matured. , is an excellent resource for studying the molecular mechanisms and genetics of embryonic development. Taking advantage of the characteristics of zebrafish embryos being transparent and juveniles being nearly transparent, we can simply add the host bacteria containing the araQS system into the zebrafish breeding environment. Through the swallowing and drinking of zebrafish, the water containing the host bacteria is sucked into the body, creating an internal environment for the host bacteria, and inducing its expression.

[0102] Put the juvenile zebrafish into the PBS suspe...

Embodiment 7

[0104] Example 7, Construction of WED multivalent carrier attenuated vaccine strain based on araQS system and immune evaluation of vaccine strain WED (araQS-G) in turbot

[0105] See Xiao J et al., Fish&shellfish immunology.2013 35:632-41 for the WED multivalent carrier attenuated vaccine strain.

[0106] Using the genome of Aeromonas hydrophila LSA34 (obtained from the Shanghai Ahua Bioengineering Institute) as a template, and using gap-F and gap-R as primers, the GAPDH protein-encoding gene gapA34 of Aeromonas hydrophila was amplified , replace it with pUTat-qs-katushka-TT-araC-P araBAD - katushka in aiiA, the expression of gapA34 gene is controlled by the araQS system, the plasmid map is as follows Figure 7 shown.

[0107] This in vivo induced expression plasmid was transformed into Edwardsiella tarda supervisory vaccine strain WED, making it a vector vaccine strain WED (araQS-G).

[0108] In order to evaluate the resistance effect of the multivalent vaccine to turbot a...

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Abstract

The invention relates to a protein expression system regulated by an arabinose-bacteria density binary signal and its application. The ''arabinose-bacteria density'' binary signal regulated protein expression system (araQS) is disclosed for the first time. A bacterial quorum-sensing system is tactfully applied to be combined with arabinose promoter (araC-ParaBAD) regulation expressed quorum quenching gene aiiA, thus making dual regulation expression of heterologous protein to be possible.

Description

technical field [0001] The invention belongs to the field of bioengineering, and more specifically, the invention relates to a protein expression system regulated by an arabinose-bacteria density binary signal and its application. Background technique [0002] Bacterial cells can release a small molecule chemical substance called autoinducer (AI), which gradually accumulates as the concentration of bacteria in the surrounding environment increases, and when its concentration reaches a certain threshold, it will be absorbed by the bacteria. Detect and generate a response, so that the bacteria uniformly produce changes in gene expression. This process is called quorum sensing (QuorumSensing, QS). The quorum sensing system of microorganisms is a quorum regulation mechanism for mutual communication between bacteria that has attracted much attention in recent years. Among them, the most notable is that the lux quorum sensing system in Vibrio fischeri regulates the bioluminescenc...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N15/74C12N1/21A61K39/116A61P31/04
Inventor 刘琴张元兴初腾王蓬勃王启要肖婧凡
Owner EAST CHINA UNIV OF SCI & TECH