Ganoderma lucidum mutant strain with high yield of polysaccharide and triterpenoid

Abstract

The invention belongs to the technical field of food microorganisms, and particularly discloses a ganoderma lucidum mutant strain capable of producing polysaccharides and triterpenoids at high yield. The strain is named as Ganoderma lucidum in Latin, the preservation number is CGMCC NO: 41680, the preservation unit is China General Microbiological Culture Collection Center, and the preservation date is December 4, 2024. The strain is named as Ganoderma lucidum, the preservation number is CGMCC NO: 41680, and the preservation number is CGMCC NO: 41680. The polysaccharide content in the ganoderma lucidum spore powder produced by the ganoderma lucidum mutant strain is more than or equal to 8%, and the total triterpenoid content is more than or equal to 12%; a culture medium formula of the ganoderma lucidum mutant strain comprises potato leachate, glucose and monopotassium phosphate, the culture temperature is 25-28 DEG C, and the pH value is 5.5-6.5. The ganoderma lucidum mutant strain is high in genetic stability, the content of active ingredients is remarkably higher than that of parent strains, and the ganoderma lucidum mutant strain is suitable for large-scale fermentation production.

Description

Technical Field

[0001] The present invention belongs to the technical field of food microbiology, and particularly relates to a new Ganoderma lucidum strain with high polysaccharide and triterpenoid production obtained by mutagenesis breeding. Background Art

[0002] Ganoderma lucidum is a fungus belonging to the genus Ganoderma in the family Polyporaceae of the class Basidiomycetes, and is known as the "immortal herb". Its medicinal history can be traced back to "Shennong's Herbal Classic". Modern pharmacological studies have shown that Ganoderma lucidum polysaccharides and triterpenoids are its main active ingredients, with effects such as immune regulation, anti-tumor, antioxidant, liver protection, and anti-inflammatory. However, the content of polysaccharides and triterpenoids in wild Ganoderma lucidum and traditional cultivated strains is relatively low, making it difficult to meet the requirements of industrial production. Therefore, the key research direction to break through the resource bottleneck is to select high-yield strains through modern biotechnology means (such as physical mutagenesis, chemical mutagenesis, and ARTP mutagenesis). Summary of the Invention

[0003] To solve the problem of the low content of polysaccharides and triterpenoids in wild Ganoderma lucidum and traditional cultivated strains, the present invention provides a Ganoderma lucidum mutagenized strain with high polysaccharide and triterpenoid production.

[0004] To solve the above technical problems, the technical solution adopted by the present invention is: a Ganoderma lucidum mutagenized strain with high polysaccharide and triterpenoid production, with the Latin name Ganoderma lucidum, deposit number CGMCC NO: 41680, deposited with the China General Microbiological Culture Collection Center on December 4, 2024; the nucleotide sequence of the Ganoderma lucidum mutagenized strain is as shown in SEQ ID NO: 1.

[0005] Furthermore, the polysaccharide content in the Ganoderma lucidum spores produced by the Ganoderma lucidum mutagenized strain is ≥8%, and the total triterpenoid content is ≥12%.

[0006] Furthermore, the culture medium formula of the Ganoderma lucidum mutagenized strain contains potato leachate, glucose, and potassium dihydrogen phosphate, and the culture temperature is 25°C to 28°C, and the pH is 5.5 to 6.5.

[0007] Compared with the prior art, the beneficial technical effects of the present invention are:

[0008] The Ganoderma lucidum mutagenized strain selected by the present invention through a composite mutagenesis method (including the synergistic effect of ultraviolet irradiation, lithium chloride chemical mutagenesis, and ARTP plasma mutagenesis) has a polysaccharide content of ≥8% and a total triterpenoid content of ≥12% in the Ganoderma lucidum spores it produces, and the polysaccharide content is increased by more than 50% compared with the parental strain; the Ganoderma lucidum mutagenized strain of the present invention has strong genetic stability, and the content of active ingredients is significantly higher than that of the parental strain, and is suitable for large-scale fermentation production. Detailed implementation manners

[0009] Example 1: A mutagenized strain of Ganoderma lucidum with high yields of polysaccharides and triterpenoids. The mutagenized strain of Ganoderma lucidum is named Taizhi 2, with the Latin name Ganoderma lucidum, the preservation number CGMCC NO: 41680, the preservation unit being the General Microbiology Center of the China Committee for Culture Collection of Microorganisms, and the preservation date being December 4, 2024; the nucleotide sequence of the mutagenized strain of Ganoderma lucidum is as shown in SEQ ID NO: 1.

[0010] The mutagenized strain of Ganoderma lucidum was obtained by mutagenesis and cultivation by Shandong Taishan Health Technology Co., Ltd. The breeding method of the mutagenized strain of Ganoderma lucidum (Taizhi 2) includes the following steps:

[0011] (1) Preparation of protoplasts.

[0012] The original strain Taizhi 1 was inoculated into a PDB liquid medium and cultured with shaking at 27°C and 150 rpm for 7 days, and the mycelia were collected. The mycelia were treated with 2% lysing enzyme at 31°C and 40 rpm for 3 hours to prepare protoplasts.

[0013] (2) Composite mutagenesis treatment.

[0014] First, ultraviolet-chemical mutagenesis was carried out: The protoplasts were suspended in a 0.6 mol / L mannitol solution, irradiated with ultraviolet light at 254 nm for 30 seconds, and then treated with a lithium chloride solution at room temperature for 20 minutes.

[0015] Then, ARTP mutagenesis was carried out: The bacterial solution after ultraviolet-chemical mutagenesis was spread on a metal slide and placed in the treatment chamber of the ARTP system. The power was set at 120 W, the helium gas flow rate was 10 L / min, and the treatment time was 60 seconds to induce secondary mutations.

[0016] (3) Regeneration culture and screening.

[0017] The mutagenized protoplasts were spread on a PDA regeneration medium and cultured in the dark at 27°C for 7 days to screen for mutant strains with fast growth rates and stable colony morphologies.

[0018] (4) Verification of genetic stability.

[0019] The screened strains were cultured by continuous 5-generation shake flask subculture, and the fluctuation range of the contents of polysaccharides and triterpenoids was detected (the coefficient of variation ≤ 5%).

[0020] (5) Determination of active ingredients.

[0021] The contents of polysaccharides and triterpenoids were quantitatively analyzed by high performance liquid chromatography (HPLC) to screen for high-yield strains.

[0022] (6) Optimization of liquid fermentation.

[0023] The selected strain was cultured by liquid fermentation, and the temperature, pH and dissolved oxygen conditions were optimized to verify the potential for industrial production. The culture medium formula of the new strain of Ganoderma lucidum No. 2 in this example contains potato leachate, glucose and potassium dihydrogen phosphate, and the culture temperature was controlled at 25°C to 28°C and the pH was 5.5 to 6.5.

[0024] In this example, the new strain of Ganoderma lucidum No. 2 with high yields of polysaccharides and triterpenoids obtained through mutagenesis breeding has a polysaccharide content of ≥8% and a total triterpenoid content of ≥12% in the produced Ganoderma lucidum spore powder.

[0025] Three strains of Ganoderma lucidum No. 1 (Ganoderma lucidum parent) and 41 strains of Ganoderma lucidum No. 2 were cultured by liquid fermentation, and their polysaccharide content and triterpenoid content were measured respectively for comparative analysis. The results showed that the polysaccharide content of the Ganoderma lucidum spore powder collected from Ganoderma lucidum No. 1 was about 3% - 3.5% after wall-breaking; the polysaccharide content of the Ganoderma lucidum spore powder collected from Ganoderma lucidum No. 2 was about 8% - 9.3% after wall-breaking; the polysaccharide content in the spore powder of Ganoderma lucidum No. 2 was increased by more than 50% compared with Ganoderma lucidum No. 1 (parent strain).

[0026] Of course, the above description is not a limitation of the present invention, and the present invention is not limited to the above examples. Changes, modifications, additions or substitutions made by those skilled in the art within the scope of the essence of the present invention should also fall within the protection scope of the present invention.

Claims

1. A Ganoderma lucidum mutant strain with high yields of polysaccharides and triterpenoids, characterized in that, The Latin name is Ganoderma lucidum, the preservation number is CGMCC NO: 41680, the preservation unit is the General Microbiology Center of the China Committee for Culture Collection of Microorganisms, and the preservation date is December 4, 2024; the nucleotide sequence of the mutagenized Ganoderma lucidum strain is as shown in SEQ ID NO:

1.

2. The Ganoderma lucidum mutagenic strain with high yields of polysaccharides and triterpenoids according to claim 1, characterized in that, The polysaccharide content in the Ganoderma lucidum spore powder produced by the mutagenized Ganoderma lucidum strain is ≥8%, and the total triterpene compound content is ≥12%.

3. The Ganoderma lucidum mutagenic strain with high yields of polysaccharides and triterpenoids according to claim 2, characterized in that, The culture medium formula of the mutagenized Ganoderma lucidum strain contains potato leaching solution, glucose and potassium dihydrogen phosphate, the culture temperature is 25°C - 28°C, and the pH is 5.5 - 6.5.