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A kind of method for improving astaxanthin production of Phaffia rhodozyma

A technology of Phaffia rhodozyme and Phaffia rhodozyma, which is applied in the biological field, can solve the problems of not fully revealing the way of astaxanthin production, and achieve the effects of reducing feedback inhibition, reducing metabolic competition, and improving metabolic flux and production

Active Publication Date: 2022-07-26
QINGDAO INST OF BIOENERGY & BIOPROCESS TECH CHINESE ACADEMY OF SCI
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Problems solved by technology

However, the above-mentioned single gene knockout did not fully reveal the way to increase astaxanthin production by regulating sterol synthesis

Method used

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  • A kind of method for improving astaxanthin production of Phaffia rhodozyma
  • A kind of method for improving astaxanthin production of Phaffia rhodozyma
  • A kind of method for improving astaxanthin production of Phaffia rhodozyma

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Embodiment

[0043] 1. Materials and methods

[0044] 1.1 Bacteria and culture conditions

[0045] Phaffia rhodozyma strain CBS 6938 (purchased from the Netherlands CBS Culture Collection) was used as the material. For routine culture, YPD liquid medium (glucose 20 g / L, peptone 20 g / L, yeast extract 10 g / L) was used, and the culture conditions were 22° C., 250 rpm.

[0046] 1.2 Construction of gene knockout vector

[0047] Homologous recombination was used to knock out sterol synthesis genes, and the knocked-out sterol synthesis genes have been annotated with corresponding enzymes in the GenBank genome database. In this example, in addition to CYP51 and CYP61, other 10 gene knockout vectors as shown in Table 1 were constructed.

[0048] Table 1. GenBank ID of sterol synthesis genes and their Scaffold

[0049]

[0050] The specific vector was constructed by Gibson assembly method, the DNA fragment was obtained by PCR method, and the primers used are shown in Table 2. All PCR amplifi...

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Abstract

The invention relates to the field of biotechnology, in particular to a method for knocking out a sterol synthesis gene to improve astaxanthin production of Phaffia rhodozyma. The recombinant strain obtained by knocking out one or several genes in the sterol synthase encoding gene of Ph. Knockout of several genes in combination with CYP61 improves the fermentation yield of Phaffia rhodozyma astaxanthin. By knocking out the sterol synthesis gene of Phaffia rhodozyma, the invention reduces the metabolic competition of sterol generation and astaxanthin synthesis or the feedback inhibition of sterol to mevalonate pathway, so as to improve the metabolic flow and yield of astaxanthin synthesis. The astaxanthin content of the single-gene knockout strain can reach 2.2 times that of the control strain; the astaxanthin production of the double-gene knockout strain can be increased by 38% on the basis of the single-gene knockout strain. The method can be applied to construct high-producing strains of astaxanthin.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to a method for knocking out a sterol synthesis gene to improve astaxanthin production of Phaffia rhodozyma. Background technique [0002] Astaxanthin is a terpene compound belonging to the class of carotenoids. Astaxanthin has strong antioxidant, coloring, anti-inflammatory, and anti-cancer effects (Park J, Chyun J, Kim Y, et al. Nutr Metab, 2010, 7:18). [0003] Phaffia rhodozyma can synthesize astaxanthin in a proportion of 50% to 85% of the cellular carotenoids, and has outstanding advantages in growth rate, cell density, anti-pollution, etc. It is considered to be an important cell factory for astaxanthin production (Bellora N, Moliné M, David-Palma M, et al. BMC Genomics, 2016, 17:901). Phaffia rhodozyma has a low yield of astaxanthin, and the astaxanthin content of the usual strain is less than 0.1 mg / g dry cell weight, which does not have the economy required for industrial ap...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N1/19C12P23/00C12N15/52C12N15/81C12N15/90C12R1/645
CPCC12P23/00C12N9/00C12N15/815C12N15/905
Inventor 王士安张宁李福利
Owner QINGDAO INST OF BIOENERGY & BIOPROCESS TECH CHINESE ACADEMY OF SCI
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