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Method for optimizing synthetic route of fatty acid in escherichia coli

A technology for fatty acid synthesis and Escherichia coli, applied in biochemical equipment and methods, botanical equipment and methods, applications, etc., can solve problems such as staying at the theoretical basis, and achieve the effect of optimizing metabolic pathways

Active Publication Date: 2014-12-10
SHANGHAI JIAOTONG UNIV
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Problems solved by technology

[0011] The first purpose of the present invention is to provide a method for optimizing the fatty acid synthesis pathway of Escherichia coli, so as to solve the defect that the relevant research on metabolic pathway optimization in the prior art mainly stays at the theoretical basis level
[0012] The second object of the present invention is to provide an E. coli fatty acid production system constructed by a method for optimizing the fatty acid synthesis pathway of E. coli, so as to solve the defect that the relevant research on the optimization of metabolic pathways in the prior art mainly stays at the theoretical basis level
[0013] The third object of the present invention is to provide the application of the fatty acid production system of Escherichia coli in the synthesis of fatty acids in Escherichia coli, so as to solve the defect that the relevant research on metabolic pathway optimization in the prior art mainly stays at the theoretical basis level

Method used

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  • Method for optimizing synthetic route of fatty acid in escherichia coli
  • Method for optimizing synthetic route of fatty acid in escherichia coli
  • Method for optimizing synthetic route of fatty acid in escherichia coli

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Embodiment 1

[0035] The raw materials used in this embodiment are as follows:

[0036] Plasmid: pET series plasmid pET28a(+) produced by Novagen;

[0037] Escherichia coli BL21 (DE3) genotype: (fhuA2[lon]ompT gal(λDE3)[dcm]ΔhsdS, λDE3=λsBamHIoΔEcoRI-B int::(lacI::PlacUV5::T7genel)i21Δnin5);

[0038] Restriction enzymes: FastDigest series enzymes NcoI, NdeI, BamHI, EcoRI, SacI, XhoI produced by Fermentas. Use its recommended reaction system for the reaction;

[0039] High-fidelity PCR enzyme: KOD-plus-Neo produced by TOYOBO, for PCR reaction to amplify fragments. Use the recommended reaction system and reaction method for the reaction.

[0040] LB liquid medium: 10g sodium chloride, 10g peptone, 5g yeast powder, add distilled water to 1L;

[0041] LB solid medium: Add agar at a final concentration of 1.5% to the LB liquid medium.

[0042] Resistance screening medium: add ampicillin and / or kanamycin antibiotic mother solution to LB medium (liquid / solid), so that the final concentration ...

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Abstract

The invention discloses a method for optimizing a synthetic route of fatty acid in escherichia coli. In a step of constructing key proteins of FabG, FabZ, FabI and TesA' of metabolic pathways of the fatty acid in the escherichia coli, escherichia coli rare codons AGG (arginine) of different quantities are respectively inserted into the key proteins of FabG, FabZ, FabI and TesA' of the metabolic pathways of the fatty acid in the escherichia coli. According to the method disclosed by the invention, the gene expression can be accurately controlled in vivo, and the corresponding metabolic pathways are optimized; and a system for producing the fatty acid in the escherichia coli, constructed by adopting the method, and can be used for efficiently producing extracellular fatty acid the yield of which is about two times that of a rare codon AGG-free non-control system.

Description

technical field [0001] The invention relates to a method for optimizing the fatty acid synthesis pathway of Escherichia coli. Background technique [0002] With the shortage of petroleum resources and the deteriorating environment, people's demand for environmentally friendly and renewable energy is becoming more and more urgent, and bioenergy should be born accordingly. Currently, available commercial biofuels include short-chain alcohols from corn and sucrose, and fatty acid methyl ester derivatives from rapeseed and tallow, but these biofuels have raw materials that are comparable to human needs. Contradictory question. To overcome these shortcomings, the best approach is to use fermentation to produce biofuels, which mainly include short-chain alcohols, hydrocarbons in the isoprenoid synthesis pathway, and microbial fatty acid derivatives. So far, microorganisms that can produce biofuels mainly include bacteria, fungi, yeast, and microalgae. However, since natural stra...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N15/53C12N15/60C12N15/55C12N15/10C12N15/70C12P7/62
Inventor 马钢贺林王右王毓舒
Owner SHANGHAI JIAOTONG UNIV
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