A dynamic control system for near-infrared light control and its application

A near-infrared light and phytochrome technology, applied in the field of metabolic engineering, can solve problems such as uneven mass transfer of dissolved oxygen, affecting the growth and accumulation of acetoin in Escherichia coli, and achieve simple design, fewer system components, and strain growth load low effect

Active Publication Date: 2022-05-06
JIANGNAN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, the production of acetoin is limited by the specific surface area, causing the problem of uneven mass transfer of dissolved oxygen, which directly affects the growth and accumulation of acetoin in E. coli

Method used

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  • A dynamic control system for near-infrared light control and its application
  • A dynamic control system for near-infrared light control and its application
  • A dynamic control system for near-infrared light control and its application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0049] Example 1 Evaluation of near-infrared light-activated components under different illumination intensities and light pulse periods

[0050] Will P J23119 -Y 30 M-P J23119 -BudAB-NOX and P mrkA -BFP plasmid was transferred into E.coli F0601 (see Dong X, Chen X, Qian Y, et al.Metabolic engineering of Escherichia coli W3110to produce L-malate[J].Biotechnology&Bioengineering,2016,114(3):656 -664.), obtain engineering strains, place them in LB culture medium, and use SpectraMax M3 microplate microplate reader to carry out continuous fluorescence measurement on the above-mentioned engineering strains. When the strain is under near-infrared light conditions of 650nm, near-infrared phytochromes (including Bphs, Bpho, Yhjh and / or Mrkh) bind to the front of the PmrkA promoter to realize the expression of fluorescent protein BFP.

[0051] The experiment was carried out under the conditions of darkness or 650nm near-infrared light, and the results were as follows: figure 1 As s...

Embodiment 2

[0052] Embodiment 2 dynamic gene circuit introduces the shake flask fermentation performance of acetoin production strain

[0053] like image 3 shown in P J23119 -Y 30 M-P J23119 -BudAB-NOX and P mrkA -RpoS (see plasmid map figure 2 ) is an expression vector, and different dynamic gene circuits are introduced into the E.coli F0601 strain, and a total of 7 engineering strains Q1-Q7 are obtained, and the Q1 strain expresses only P J23119 -Y 30 M-P J23119 -Engineering strains obtained during BudAB-NOX, Q2 strains express only P mrkA -Engineering strains obtained during RpoS, Q3-Q7 respectively indicate that the near-infrared light intensity is 0.2W / cm 2 、0.25W / cm 2 、0.3W / cm 2 、0.4W / cm 2 , Engineering strains at 0.8W / cm.

[0054] like Figure 4 and 5 As shown, the strain growth and product synthesis were detected in NBS inorganic salt medium. 37°C, 200rpm, cultivate a single colony of the engineering strain in LB medium to OD 600 was 0.05, the initial inoculation...

Embodiment 3

[0055] Example 3 Fermentation performance of fermenter for introducing acetoin production strain into dynamic gene circuit

[0056] The fermentation performance of the Q7 strain was tested in a 5L fermenter. The temperature is constant at 37°C, 500rpm, and a single colony of the engineering strain is cultivated in LB medium to OD 600 0.05, the initial inoculum was controlled to be 5%, the ventilation rate was 1vvm, the fermentation cycle was 72h, the liquid filling volume was 3L, and the initial pH was controlled to be 6.8 with 4M sodium hydroxide and 2M hydrochloric acid. At the end of the fermentation, the accumulation of acetoin reached 66.8g / L, and the conversion rate reached 0.58g / g glucose.

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Abstract

The invention discloses a dynamic control system controlled by near-infrared light and its application, specifically discloses a dynamic control system related to combined control of near-infrared light and its application in the production of acetoin, which belongs to the technical field of metabolic engineering. The present invention selects the near-infrared photosensitive protein by means of molecular biology. Taking advantage of the photosensitive protein's specific response to light source characteristics of specific wavelengths and the advantages of not needing to add exogenous inducers, the production of acetoin in inorganic salt medium was realized by introducing a dynamic regulatory gene circuit targeting RpoS into engineered Escherichia coli. The yield of acetoin produced by the invention can reach 66.8g / L, and the conversion rate can reach 0.58g / g glucose.

Description

technical field [0001] The invention relates to a dynamic control system controlled by near-infrared light and its application, in particular to a dynamic control system controlled by near-infrared light and its application in the production of acetoin, belonging to the technical field of metabolic engineering. Background technique [0002] The dynamic control system is an emerging metabolic flow control method in the field of metabolic engineering. Its main feature different from the static control is that during the fermentation process, the engineered strains will adjust according to the fermentation time, physiological state, intracellular metabolite concentration, and changes in the extracellular environment. The corresponding adjustment of enzyme activity can affect the distribution of metabolic flux and improve the production capacity of products. The dynamic regulation system has the advantages of no need for artificial regulation in the fermentation process and no n...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N1/21C12N15/70C12P7/26C12R1/19
CPCC07K14/245C07K14/195C12N15/70C12P7/26C12N2800/60
Inventor 陈修来丁强刘立明李洋刘佳罗秋玲
Owner JIANGNAN UNIV
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