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Utilize electrical energy to fix co 2 And engineering bacteria for synthesizing isopropanol and construction method

A construction method and technology for engineering bacteria, which are applied in the field of designing and constructing engineering bacteria for high-efficiency utilization of electrical energy to fix CO2 and synthesizing isopropanol, can solve the problems of high hygroscopicity, low energy density of ethanol, and corrosiveness, and improve efficiency. Effect

Active Publication Date: 2022-07-15
深圳中科翎碳生物科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Ethanol has low energy density, high hygroscopicity, and is corrosive, which is not conducive to long-term storage and large-scale addition, so it cannot be used as an ideal fuel substitute fuel

Method used

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  • Utilize electrical energy to fix co <sub>2</sub> And engineering bacteria for synthesizing isopropanol and construction method
  • Utilize electrical energy to fix co <sub>2</sub> And engineering bacteria for synthesizing isopropanol and construction method
  • Utilize electrical energy to fix co <sub>2</sub> And engineering bacteria for synthesizing isopropanol and construction method

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Experimental program
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preparation example Construction

[0067] The invention also provides the realization of CO by using the microbial electrosynthesis method 2 A method for synthesizing isopropanol, comprising:

[0068] The first step will carry the plasmid pRK-Geo2i-ISOPROPANOL-ISODH

[0069] of G.sulfurreducens ACL was cultured in medium to exponential growth phase (OD 600nm =0.4);

[0070] In the second step, the exponentially growing bacteria were inoculated into the cathode of the H-type microbial fuel cell with an inoculum of 10%, and the mixed gas was continuously fed into the anode to provide a carbon source for the bacterial cells.

Embodiment 1G

[0074] Example 1 G.sulfurreducens ACL chassis strain construction

[0075] Construction of pUC19-aclBA plasmid

[0076] Escherichia coli containing pUC19 plasmid E. coli Beta 10 was cultured to the mid-exponential growth period, 5 mL of bacterial solution was taken, centrifuged at 6000 × g for 5 minutes to collect the bacterial cells, the medium was discarded, and plasmid DNA was extracted according to the kit instructions. Primers PUC19-F and PUC19-R were designed according to the sequence of the foreign gene insertion site, and the plasmid fragment was amplified by DNA polymerase Q5.

[0077] According to the codon-optimized isocitrate lyase sequence and the chloramphenicol gene sequence, primers ACL-F and ACL-R and Cat-F and Cat-R were designed according to the overlap between the Gibson DNA fragments, and the synthesized gene fragments were Templates were amplified by DNA polymerase Q5 to the relevant fragments, respectively.

[0078] Will G.sulfurreducens PCA (pu...

Embodiment 2

[0100] Example 2 Strain G.sulfurreducens Construction of ACL-ISODH

[0101] Construction of pRK-Geo5-ISODH plasmid

[0102] Will G.sulfurreducens The ACL original bacteria were cultured to the mid-exponential growth stage, 5 mL of bacterial liquid was taken, centrifuged at 6000 × g for 5 minutes to collect the bacterial cells, the medium was discarded, and the whole genome DNA was extracted according to the kit instructions. E. coli containing the pRK-Geo2 plasmid E. coli λS17-1 was cultured to the mid-exponential growth stage, 5 mL of bacterial liquid was taken, centrifuged at 6000 × g for 5 minutes to collect the bacterial cells, the medium was discarded, and plasmid DNA was extracted according to the kit instructions. by G.sulfurreducens ACL whole genome DNA was used as template, primers ISODH-F and ISODH-R were designed according to the sequence of isocitrate dehydrogenase, and the gene fragment was amplified by DNA polymerase Q5; Dot sequence design primers pR...

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Abstract

The present invention provides a method for designing and constructing high-efficiency use of electrical energy to fix CO by using synthetic biotechnology. 2 And the method and application of engineering bacteria for synthesizing isopropanol. The method of designing and constructing engineering bacteria is to be able to utilize electric energy and CO. 2 electroactive microorganisms Geobacter sulfurreducens ACL is the chassis. In the chassis strain, isocitrate dehydrogenase and isopropanol synthesis pathway-related enzymes are efficiently expressed by plasmids. It has been verified by experiments that the strain constructed by this method can efficiently utilize electricity and CO. 2 Synthesis of isopropanol.

Description

technical field [0001] The invention relates to the technical field of synthetic biology, in particular to a design, construction and efficient utilization of electrical energy to fix CO 2 And the construction method and application of synthetic isopropanol engineering bacteria. Background technique [0002] The microbial electrocatalysis process uses two-way electron and energy exchange between electroactive microorganisms and the external environment to achieve a wide range of applications in the field of environment and energy. Microbial electrosynthesis (MES) can directly use electrical energy to drive microorganisms to reduce and fix CO 2 , synthesizing multi-carbon compounds, providing new opportunities for renewable new energy conversion, fine chemical preparation and ecological environmental protection. However, the rate of extracellular electron transfer (EET) between the known microorganisms and electrodes is low, and it is difficult to meet the requirements of i...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N15/74C12N15/53C12N15/54C12N15/60C12N1/21C12P7/04C12R1/01
CPCC12N15/74C12N9/0006C12N9/1029C12N9/88C12N15/52C12P7/04C12Y101/01041C12Y101/01042C12Y203/01194C12Y203/01016C12Y401/01004C12Y101/01C12N2800/22Y02P20/133
Inventor 夏霖谭扬周明新王可汗
Owner 深圳中科翎碳生物科技有限公司
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