Method for increasing exogenous synthesis yield of water-soluble algal cyanine

A phycocyanin and water-soluble technology, which is applied in the field of increasing the exogenous synthetic yield of water-soluble phycocyanin, can solve problems such as low yield of water-soluble phycocyanin, and achieve the effect of high-efficiency culture conditions

Active Publication Date: 2020-02-14
ANHUI NORMAL UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] The technical problem that the present invention solves is: propose a kind of method that can improve the heterologous synthesis yield of water-soluble phycocyanin, be used for overcoming the above-mentioned water-soluble Disadvantages of low production of sex phycocyanin

Method used

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  • Method for increasing exogenous synthesis yield of water-soluble algal cyanine
  • Method for increasing exogenous synthesis yield of water-soluble algal cyanine
  • Method for increasing exogenous synthesis yield of water-soluble algal cyanine

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0036] 1 Construction of recombinant strains

[0037] According to the gene CphA encoding phycocyanin synthase according to Thermosynechococcus elongatus BP-1 in GenBank BP Sequence, artificially synthesized whole gene sequence, connected with plasmid pCOLADuet-1 through SalI and NotI restriction endonuclease sites, and the recombinant plasmid was transformed into E. coli DH5α competent cells. The recombinant plasmid was extracted and transformed into E. coli BL21 competent cells again to obtain a recombinant strain.

[0038] 2 Activated bacteria

[0039] Under aseptic conditions on the ultra-clean workbench, glycerol strains were streaked on LB solid plates supplemented with 0.1% kanamycin, placed in a constant temperature incubator at 37°C, and cultured for 12 h. A single colony of recombinant strains was picked from the plate, and cultured in 100 mL of LB liquid medium supplemented with 0.1% kanamycin for 16 h under shaking conditions at 37° C. and rotating speed of 180 r...

Embodiment 2

[0055] 1 Activated bacteria

[0056] The method is the same as example 1

[0057] 2 Fermentation culture

[0058] The method is the same as example 1

[0059] The fermentation medium formula is 2.4% yeast powder, 0.4 mL of glycerol, 10 mL of phosphate buffer (17 mM KH 2 PO 4 and 72mM K 2 HPO 4 ).

[0060] 3 Purification and extraction of phycocyanin

[0061] The method is the same as example 1

[0062] The proportion of water-soluble phycocyanin was calculated, and the result was 149.2 mg / g (phycocyanin mass / cell mass).

[0063] OD of the bacterial solution 600 Adjust to 0.5-0.6 to make SDS-PAGE samples, and perform polyacrylamide gel electrophoresis. The results are as follows figure 1 shown. Lane M is the protein molecular weight standard; lane 2 is the total protein of the culture medium with 2.4% yeast powder added.

[0064] The purified phycocyanin was subjected to SDS-PAGE electrophoresis. The result is as figure 2 . Lane M is the protein molecular weight...

Embodiment 3

[0070] 1 Activated bacteria

[0071] The method is the same as example 1

[0072] 2 Fermentation culture

[0073] The method is the same as example 1

[0074] The fermentation medium formula is 4.8% yeast powder, 0.4 mL glycerol, 10 mL phosphate buffer (17 mM KH 2 PO 4 and 72mM K 2 HPO 4 ).

[0075] 3 Purification and extraction of phycocyanin

[0076] The method is the same as example 1

[0077] The proportion of water-soluble phycocyanin was calculated, and the result was 105.3 mg / g (phycocyanin mass / cell mass).

[0078] OD of the bacterial solution 600 Adjust to 0.5-0.6 to make SDS-PAGE samples, and perform polyacrylamide gel electrophoresis. The results are as follows figure 1shown. Lane M is the protein molecular weight standard; lane 4 is the total protein of the culture medium with 4.8% yeast powder added.

[0079] The purified phycocyanin was subjected to SDS-PAGE electrophoresis. The result is as figure 2 . Lane M is the protein molecular weight standa...

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Abstract

The invention discloses a method for increasing an exogenous synthesis yield of water-soluble phycocyanin, which belongs to the technical field of biological genetic engineering and microbial fermentation. According to the method, a TB culture medium is used as a contrast; an expression vector system is induced according to escherichia coli; the phycocyanin is synthesized by using the phycocyaninsynthase; the specific culture medium components and the culture temperature are selected, so that the synthesis yield of the water-soluble algae is greatly increased, the water-soluble algae producedby the method has the characteristic that the yield is 2.2 times higher than that of the water-soluble algae produced by control culture, and more efficient culture conditions are provided for the production of the water-soluble algae.

Description

technical field [0001] The invention relates to the technical fields of biological genetic engineering and microbial fermentation, in particular to a method for improving the exogenous synthetic yield of water-soluble phycocyanin and its application. Background technique [0002] Phycocyanin (CGP) is a non-ribosomally synthesized amino acid polymer found in most cyanobacteria and some heterotrophic bacteria. It consists of a poly-L-aspartic acid (Asp) backbone with numerous arginines (Arg) linked through their amino groups to side chains formed on the carboxyl group of each aspartic acid. Arg and Asp are generally present in the polymer in equimolar amounts. Natural phycocyanin has a molecular mass ranging from 25kDa to 100kDa and has a wide range of polydispersity. Phycocyanins in the natural state can be divided into two forms according to their physical properties: insoluble and water-soluble phycocyanins. Those soluble in neutral solutions are called water-soluble phy...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12P13/02C12N1/21C12R1/19
CPCC12P13/02C12N9/93C12Y603/02029Y02A50/30
Inventor 林凌陈明明朱国萍
Owner ANHUI NORMAL UNIV
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