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Endophytic strains with high camptothecin yield and application thereof

A technology of camptothecin and bacterial strains, which is applied in the field of medicinal plant endophytic bacteria research, can solve the problems of poor genetic stability and limitations, and achieve the effect of good genetic stability

Active Publication Date: 2021-08-20
ZHEJIANG CHINESE MEDICAL UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] In the process of producing medicinal ingredients by endophytes, poor genetic stability and other characteristics limit their industrial application. Therefore, it is of great significance to obtain endophyte production strains that can stably inherit camptothecin

Method used

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  • Endophytic strains with high camptothecin yield and application thereof
  • Endophytic strains with high camptothecin yield and application thereof
  • Endophytic strains with high camptothecin yield and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0024] Embodiment 1, the separation of endophytic fungus of camphor tree seed

[0025] First, rinse the magma seeds in tap water overnight. The seeds were then washed with 75% (v / v) ethanol for 3 minutes, rinsed with sterile water for 5 times, disinfected with 50% (v / v) sodium hypochlorite for 5 minutes, rinsed again with sterile water for 5 times for surface disinfection, and finally placed on sterile filter paper. Dry on top. Subsequently, cut the surface-sterilized camphor tree seeds to a size of 2mm, place them on PDA medium, culture them at 25°C for 5-10 days, and periodically purify to obtain three single colonies of S1, S2, and S3.

Embodiment 2

[0026] Colony morphology analysis of embodiment 2, S1, S2, S3 single bacterium colony

[0027] Identification of endophytic fungi by morphological methods.

[0028] like figure 1 As shown, the morphology of the S1 colony is as follows: the primary hyphae are white, and the aerial hyphae are more developed. As the culture time prolongs, the center of the colony becomes dark green, the opposite side of the colony is brownish yellow, the texture of the colony is fluffy, the edges of the colony are irregular, and no infiltration out of fluid.

[0029] The colony morphology of S2 is as follows: the primary hyphae are white, the colony texture is fluffy, and the center of the colony is black.

[0030] The shape of S3 colony is as follows: at the initial stage of culture, it is white and round, with smooth edges, dry surface, and grows rapidly. The colony is radially grooved, surrounded by white hyphae in the middle, and is gray-green in color. After 4-5 days, the back of the col...

Embodiment 3

[0031] Embodiment 3, endophytic fungus S1, S2, S3 microscopic examination and identification of producing camptothecin

[0032] The isolated strains were stained with lactophenol cotton blue, and their morphology was observed under a microscope. microscopic analysis such as figure 2 It shows that the vegetative mycelium of strain S1 is composed of branched hyphae with transverse septa, and the spores are relatively thin. Conidiophores grow from the mycelium, which are erect and short, and conidia are formed at the top. With obvious diaphragm and mediastinum. The hyphae of the S2 strain were filamentous and transparent. Conidia are few and mostly solitary. The conidia of S3 strain had fewer branches, the ends were radial, the top was nearly spherical, and green spores were produced. The spores were ovoid. The walls of the conidia are smooth and the hyphae are transparent.

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Abstract

The invention provides endophytic strains with high camptothecin yield. The endophytic strains with high camptothecin yield are Alternaria temuissima ZCMUKL-S1 with the preservation number of CCTCC (China Center for Type Culture Collection) NO: M2021189, or Alternaria citricancri ZCMUKL-S2 with the preservation number of CCTCC NO: M2021190, or Aspergillus versicolor ZCMUKL-S3 with the preservation number of CCTCC NO: M2021191. The endophytic fungi have high camptothecin yield, the camptothecin yield of unit dry mycelium powder of the Aspergillus versicolor ZCMUKL-S3 is up to 116 [mu]g / g, which is the highest yield of camptothecin produced by the endophytic fungi reported at present, and the endophytic fungi have potential application value for industrial production of camptothecin. In addition, the endophytic fungi with high camptothecin yield provided by the invention have better genetic stability.

Description

technical field [0001] The invention relates to an endophytic fungus capable of high camptothecin production, which belongs to the research field of medicinal plant endophytes. [0002] technical background [0003] Camptothecin is a monoterpene indole alkaloid with unique topoisomerase I inhibition and thus unique inhibitory activity against cancer. A variety of new camptothecin anti-tumor drugs with good drug efficacy, low toxicity and side effects and good solubility have been screened out and used in clinical practice. It is the third largest plant anti-cancer drug. [0004] Medicinal plants such as camptotheca and snakeroot can biosynthesize camptothecin. Among them, camptothecin is the main source of camptothecin production, but its content is still low, and it is very difficult to directly extract from camptothecin, which can no longer meet the requirements. Great demand in the medical market. Therefore, it is of great significance to obtain alternative sources for t...

Claims

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Application Information

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IPC IPC(8): C12N1/14C12P17/18C12R1/645C12R1/66
CPCC12P17/18C12N1/145C12P17/188C12R2001/66C12R2001/645
Inventor 开国银冯岳黄超
Owner ZHEJIANG CHINESE MEDICAL UNIVERSITY
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