Recombination blue-green alga for producing lactic acid as well as preparation method and applications thereof

A technology of cyanobacteria and lactic acid, applied in the biological field, can solve the problems of aggravating global food, aggravating the shortage of fossil energy, and unable to achieve sustainable development of renewable energy, and achieve the effect of avoiding the use of organic matter

Inactive Publication Date: 2013-08-07
INST OF MICROBIOLOGY - CHINESE ACAD OF SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

In order to alleviate energy shortage and avoid environmental pollution, in recent years, researchers have attempted to transform bacteria through metabolic engineering to produce optically pure lactic acid by fermentation. Although this method avoids the problem of exacerbating the shortage of fossil energy, it will intensify the problem of food as raw material, such as mass production. The global food shortage problem cannot realize the sustainable development of renewable energy, so the use of solar energy directly converts CO 2 Photoautotrophic bacteria cyanobacteria converted to organic matter are an ideal pathway for the production of optically pure lactic acid

Method used

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  • Recombination blue-green alga for producing lactic acid as well as preparation method and applications thereof
  • Recombination blue-green alga for producing lactic acid as well as preparation method and applications thereof
  • Recombination blue-green alga for producing lactic acid as well as preparation method and applications thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0065] Embodiment 1, the construction of recombinant cyanobacteria S.M1

[0066] Establishment of an optically pure D-lactate synthesis pathway in the freshwater cyanobacterium Synechocystis sp. PCC 6803. Since there is pyruvate, the immediate precursor of lactic acid synthesis, in cyanobacteria cells, D-lactate dehydrogenase (D-LDH), which catalyzes the conversion of pyruvate into D-lactate, can be overexpressed in cyanobacteria cells to realize the production of D-lactate by cyanobacteria.

[0067] 1. Construction of recombinant plasmid pMD-Dldh (Synechocystis D-ldh homologous recombination integration expression vector)

[0068] 1. Acquisition of D-lactate dehydrogenase encoding gene Dldh

[0069] The D-lactate dehydrogenase coding gene Dldh was synthesized after entrusting Shanghai Bioengineering Co.

[0070] The amino acid sequence of D-lactate dehydrogenase is sequence 3 in the sequence listing, and its nucleotide sequence is 608-1788 nucleotides from the 5' end of seq...

Embodiment 2

[0103] Embodiment 2, recombinant cyanobacteria S.M1 produces optically pure D-lactic acid

[0104] 1. Preparation of optically pure D-lactic acid

[0105] method one,

[0106] Lactic acid is usually produced during fermentation. Therefore, cells are cultured under normal culture conditions. When the biomass accumulates to a certain extent, dark fermentation culture is used for induction.

[0107] 1. Illumination cultivation

[0108] The above-mentioned recombinant cyanobacteria S.M1 obtained by Example 1 was placed in a light incubator and shaken to the late logarithmic growth period (making its cell density reach OD 730 =1.5); culture time is 7 days; temperature is 30°C, light intensity is 100μm / m 2 · s, the oscillation frequency is 130r / min, the culture medium is BG-11 (using inorganic carbon as the sole carbon source), and the light culture product is obtained.

[0109] 2. Dark cultivation

[0110] Then, the light culture product was subjected to dark static culture fo...

Embodiment 3

[0130] Embodiment 3, the construction of recombinant cyanobacteria S.M2

[0131] Synthetic pathway of optically pure D-lactic acid was established in seawater cyanobacterium Synechococcus sp. PCC 7002. Since there is pyruvate, the immediate precursor of lactic acid synthesis, in cyanobacteria cells, D-lactate dehydrogenase (D-LDH), which catalyzes the conversion of pyruvate into D-lactate, can be overexpressed in cyanobacteria cells to realize the production of D-lactate by cyanobacteria. Since there is lactate dehydrogenase (LDH) coding gene ldh in Synechococcus sp. 7002, the production of optically pure lactic acid D-lactic acid by seawater cyanobacteria can be achieved by expressing D-lactate dehydrogenase with ldh as the insertion site.

[0132] 1. Construction of recombinant plasmid pMD-Dldh (Synechococcus D-ldh homologous recombination integration expression vector)

[0133] 1. Acquisition of D-lactate dehydrogenase encoding gene Dldh

[0134] The D-lactate dehydrogena...

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Abstract

The invention discloses a recombination blue-green alga for producing a lactic acid as well as a preparation method and applications thereof. The method disclosed by the invention comprises the following steps of: modifying a metabolic pathway, and importing a coding gene of D-lactic dehydrogenase into a genome of the blue-green alga to obtain the recombination blue-green alga for producing an optical pure D-lactic acid. An amino acid sequence of the D-lactic dehydrogenase is a sequence 3 in a sequence table. Proved by experiments, the recombination blue-green alga disclosed by the invention is constructed through a metabolic engineering modification method, and carbon dioxide (CO2) can be directly converted into organic matter by utilizing solar energy by utilizing the characteristic that the blue-green alga is photoautotrophic; and the recombination blue-green alga can be directly synthetized into the optical pure D-lactic acid by utilizing the solar energy and the CO2 by utilizing the characteristic that the blue-green alga can grow in seawater and sewage and the advantage that genetic manipulation can be easily performed on the blue-green alga.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to a lactic acid-producing recombinant blue-green algae and its preparation method and application. Background technique [0002] The energy problems and environmental problems facing the world have put forward the requirements for the development of renewable clean energy chemicals, and the food shortage requires the development of renewable clean energy chemicals to achieve the goal of not competing with people for food, crops, and People and crops compete for fresh water. [0003] Blue-green algae, also known as cyanobacteria, is a prokaryotic organism that can perform oxygen-evolving photosynthesis. Cyanobacteria are widely distributed in fresh water, sea water and even sewage, and can utilize CO 2 It reproduces rapidly with solar energy and is an ideal host for biosynthetic energy products. As a prokaryote, cyanobacteria have a simple cell structure and a similar genetic backgro...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N15/53C12N15/63C12N1/21C12P7/56C12R1/01
CPCY02P20/133Y02P20/59
Inventor 周杰张海峰李寅
Owner INST OF MICROBIOLOGY - CHINESE ACAD OF SCI
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