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A method for increasing the biomass and carotenoid content of Chlamydomonas reinhardtii by using DNA methyltransferase inhibitors

A carotene and transferase technology, which is applied in the field of food science, can solve the problems of affecting the yield of recombinant protein, reducing the yield of the final target product, and low biomass of Chlamydomonas reinhardtii, and achieves the advantages of pigment accumulation, convenient operation, and increased biomass. and carotenoid content

Active Publication Date: 2021-03-30
SOUTH CHINA UNIV OF TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0003] Although Chlamydomonas reinhardtii has been paid attention to as a bioreactor and has certain applications, some aspects still need to be further developed and improved. One of the aspects is that the biomass of Chlamydomonas reinhardtii that is purely photosynthetic and autotrophic is low, which affects the recombination Protein production, or reduce the production of final target products (high value-added products including carotenoids)

Method used

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  • A method for increasing the biomass and carotenoid content of Chlamydomonas reinhardtii by using DNA methyltransferase inhibitors
  • A method for increasing the biomass and carotenoid content of Chlamydomonas reinhardtii by using DNA methyltransferase inhibitors
  • A method for increasing the biomass and carotenoid content of Chlamydomonas reinhardtii by using DNA methyltransferase inhibitors

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0041] (1) Cultivation of Chlamydomonas reinhardtii

[0042] First prepare Chlamydomonas reinhardtii TAP liquid culture medium, its composition is: Tris 2.42g / L, TAP-salts (containing NH 4 Cl 15g / L, MgSO 4 ·7H 2 O 4g / L, CaCl 2 2H 2 O 2g / L) 25mL / L, phosphate solution (containing K 2 HPO 4 28.2g / 100mL, KH 2 PO 4 14.4g / 100mL) 1mL / L, trace element solution (containing Na 2 EDTA·2H 2 O 5g / 100mL, ZnSO 4 ·7H 2 O 2.2g / 100mL, H 3 BO 3 1.14g / 100mL, MnCl 2 4H 2 O 0.5g / 100mL, FeSO 4 ·7H 2 O0.5g / 100mL, CoCl 2 ·6H 2 O 0.16g / 100mL, CuSO 4 ·5H 2 O 0.16g / 100mL, (NH 4 ) 6 Mo 7 o 24 4H 2 (00.11g / 100mL) 1mL / L, acetic acid 1mL / L, adjust the pH to 6.95-7.0.

[0043] Prepare 40mM 5-azacytidine (5AzaC) mother solution: weigh 0.09768g 5AzaC powder and dissolve it in 10mL deionized water.

[0044] Then Chlamydomonas reinhardtii cells were inoculated in 100 mL of the above-mentioned liquid medium, and 5AzaC was added to the final concentrations of 0 (control), 100 μM and 800...

Embodiment 2

[0056] (1) Cultivation of Chlamydomonas reinhardtii

[0057] First prepare Chlamydomonas reinhardtii TAP liquid culture medium, with embodiment 1.

[0058] Prepare 40mM 5-aza-2'-deoxycytidine (5AzadC) mother solution: weigh 0.09128g 5AzadC powder and dissolve it in 10mL deionized water.

[0059] Then Chlamydomonas reinhardtii cells were inoculated in 100 mL of the above-mentioned liquid medium, and 5AzadC was added to a final concentration of 0 (control), 100 μM and 800 μM (that is, 250 μL and 2000 μL of 40 mM 5AzadC mother solution were added respectively), and the light intensity was 8000 Lx, and the light was dark. The cycle is 14h:10h (light for 14h, no light for 10h) (alternate light and dark culture), the temperature is 25°C, the rotation speed is 50r / min, and the culture is 5-7 days.

[0060] (2) Determination of biomass

[0061] Take 3-4 mL of algae liquid cultured for 5 to 7 days, and measure the absorbance value (OD 7s0 ), found that the biomass of the treatment g...

Embodiment 3

[0065] (1) Cultivation of Chlamydomonas reinhardtii

[0066] With embodiment 1, but culture time is 14 days.

[0067] (2) Determination of biomass

[0068] After 90 mL of algal cells were collected by centrifugation, they were dried in an oven at 55°C for 48 hours to constant weight, and then weighed directly. It was found that the dry weight of algal cells in the 100 μM and 800 μM 5AzaC treatment groups was 15.2% and 17.6% higher than that of the untreated control group, respectively.

[0069] (3) Extraction and determination of pigment

[0070] The method of extracting and measuring pigments from Chlamydomonas reinhardtii was the same as that in Example 1. After calculation, it was found that adding 100 μM and 800 μM 5AzaC to treat Chlamydomonas reinhardtii after 14 days of culture, the carotenoid content was 11.4% and 10.5% higher than that of the control group, respectively.

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Abstract

The invention discloses a method for improving the biomass and carotenoid content of Chlamydomonas reinhardtii by using a DNA methyltransferase inhibitor, and belongs to the field of food science andtechnology. According to the method, after the Chlamydomonas reinhardtii cells are cultured in a liquid medium containing a DNA methyltransferase inhibitor for a certain period of time, the biomass ismonitored, the algae cells are collected and the pigments are extracted every day. The method of the invention is simple and feasible, has a good effect, is convenient to operate, and can help quickly and effectively improve the biomass and carotenoid content of Chlamydomonas reinhardtii after adding of the DNA methyltransferase inhibitor. In the invention, the dosage of the DNA methyltransferaseinhibitor is very small, the effect is remarkable, and the operation is convenient.

Description

technical field [0001] The invention belongs to the technical field of food science, and in particular relates to a method for increasing the biomass and carotenoid content of Chlamydomonas reinhardtii by using a DNA methyltransferase inhibitor. Background technique [0002] Chlamydomonas reinhardtii has simple culture conditions, a short growth cycle, complete genome sequencing, clear genetic background, easy genetic transformation, and accurate post-translational modification and folding of eukaryotic proteins; it belongs to the safety level (generally recognized as safe, GRAS) ) microorganisms, the expression product does not contain toxins, which is easy to purify and reduce the cost of protein production; in addition, the cultivation of Chlamydomonas reinhardtii does not need to occupy valuable arable land resources, and does not need to consume a lot of fertilizer and fresh water resources. It has been considered to have great development potential At present, Chlamydo...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12P23/00C12N1/12C12R1/89
CPCC12N1/12C12P23/00
Inventor 姜建国梁明华宋德幸梁志聪
Owner SOUTH CHINA UNIV OF TECH
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