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Recombinant plasmid and genetically engineered strain for high-yield production of heme, construction method of recombinant plasmid, construction method of genetically engineered strain, and method for high-yield production of heme

A technology of genetically engineered strains and recombinant plasmids, applied in the field of metabolic engineering, can solve problems such as pollution, low yield, and time-consuming

Active Publication Date: 2021-02-02
UNIV OF SCI & TECH OF CHINA
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Problems solved by technology

[0002] At present, the production of heme mainly uses organic extraction or enzymatic hydrolysis to separate free heme from biological samples (such as plant tissue and animal blood), which is not only complicated, low yield, time-consuming, but also causes pollution problems.

Method used

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  • Recombinant plasmid and genetically engineered strain for high-yield production of heme, construction method of recombinant plasmid, construction method of genetically engineered strain, and method for high-yield production of heme
  • Recombinant plasmid and genetically engineered strain for high-yield production of heme, construction method of recombinant plasmid, construction method of genetically engineered strain, and method for high-yield production of heme
  • Recombinant plasmid and genetically engineered strain for high-yield production of heme, construction method of recombinant plasmid, construction method of genetically engineered strain, and method for high-yield production of heme

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Embodiment 1

[0035] 1. Strains, plasmids and culture methods

[0036] All strains and plasmids used in this example are listed in Table 1. Among them, Escherichia coli NEB10β was used for phenotypic verification of cloned strains and gene circuits. E. coli WM3604 was used for conjugation transfer. All E.coli and S.oneidensis were cultured in LB medium, and Escherichia coli WM3604 needed to add 2 6-diaminopimelic acid (DAP) at a final concentration of 50 μM. E.coli and S.oneidensis were cultured at 37°C and 30°C, respectively. The composition of 2YT medium is: tryptone (Tryptone) 10g / L, yeast powder (Yeast extract) 5g / L, NaCl 10g / L. Add 15g / L agar powder (Agar) to the solid medium.

[0037] Antibiotics were added to the above strains as needed: chloramphenicol: 30 μg / mL (E.coli), 10 μg / mL (S.oneidensis); tetracycline: 10 μg / mL (E.coli), 5 μg / mL (S.oneidensis). Anhydrotetracycline was used as an inducer, and was added to the medium according to the concentration indicated in the text. ...

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Abstract

The invention relates to the field of metabolic engineering, in particular to a recombinant plasmid and a genetic engineering strain for high-yield production of heme, a construction method of the recombinant plasmid, a construction method of the genetic engineering strain and a method for high-yield production of the heme. The recombinant plasmid comprises the following genes: hemA, hemL, hemB1,hemB2, hemC, hemD, hemE, hemN, hemG1, hemG2, hemG3, hemH1, hemH2 and ccmf. According to the invention, metabolic engineering and synthetic biological means are utilized to modify S.oneidensis MR-1, sothat the heme is efficiently synthesized. The application of the method to engineering modification of electroactive bacteria is helpful for the application of the bacteria to production of fine chemicals.

Description

technical field [0001] The invention relates to the field of metabolic engineering, in particular to a recombinant plasmid and a genetically engineered bacterial strain with high hemoglobin production, a construction method thereof, and a method for high hemoglobin production. Background technique [0002] At present, the production of heme mainly uses organic extraction or enzymatic hydrolysis to separate free heme from biological samples (such as plant tissue and animal blood). The process is complex, low yield, time-consuming and causes pollution problems. The development of metabolic engineering technology provides an opportunity for the production of heme by microbial cell engineering. [0003] Electroactive bacteria have evolved a unique way of extracellular electron transfer to use a variety of electron acceptors to complete the metabolic process of cellular respiration in an anaerobic environment. Such microorganisms therefore play an important role in energy recove...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/74C12N1/21C12P17/18C12R1/01
CPCC12N15/74C12N15/52C12N9/0008C12N9/88C12N9/1085C12N9/001C12P17/18C12Y102/0107C12Y205/01061C12Y402/01075C12Y401/01037C12Y103/03004C12Y499/01001C12N2830/003Y02E60/50
Inventor 俞汉青李洋汤强刘东风
Owner UNIV OF SCI & TECH OF CHINA
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