Streptomyces albidus SY6 as well as fermentation seeds and application thereof

Abstract

The invention relates to the technical field of industrial biology, in particular to streptomyces albus SY6 and a fermentation seed and application thereof, the streptomyces albus SY6 is characterized in that the streptomyces albus SY6 is preserved in China General Microbiological Culture Collection Center on November 13, 2024, the preservation number is CGMCC No.32602, and the streptomyces albus is classified and named as streptomyces albus. The streptomyces albidus SY6 screened by the invention has a better promotion effect in the aspects of mediating a mould signal and strengthening epsilon-polylysine synthesis.

Description

Technical Field

[0001] The invention relates to the field of industrial biotechnology, and in particular to Streptomyces parvus SY6 and fermented seeds and applications thereof. Background Art

[0002] Epsilon-poly-l-lysine is a homotypic cationic polymer synthesized by Streptomyces (such as Streptomyces parviflorus, Streptomyces chromogenicus, and Streptomyces fusogenus). It is formed by the dehydration condensation of the α-carboxyl group and the ε-amino group of lysine, with a degree of polymerization ranging from 25 to 35 lysine residues. It has the effect of resisting Gram-positive / negative bacteria, molds, yeasts, and viruses. It has strong heat resistance, high water solubility, safety and non-toxicity, and environmental degradation. It is currently used as a food preservative. At present, the fermentation level of epsilon-polylysine is low, which makes the production cost high, and the high selling price limits its market application and promotion.

[0003] At present, the strains known to be able to synthesize ε-polylysine include Streptomyces, Kitasatosporium and Bacillus subtilis. However, there are differences in the ability to synthesize natamycin between different strains, and the yield of synthesized ε-polylysine is relatively low, which is difficult to meet the needs of large-scale industrial production. Therefore, it is of great significance to develop a high-yield and easy-to-cultivate ε-polylysine synthesis strain. Summary of the invention

[0004] In order to explore high-yield strains to increase the yield of synthesized ε-polylysine, the present invention provides a Streptomyces SY6 and its fermented seeds and applications. The Streptomyces SY6 provided by the present invention can strongly respond to mold signals and increase the synthesis yield of ε-polylysine.

[0005] A Streptomyces albulus SY6, characterized in that it was deposited in the General Microbiology Center of China National Microbiological Culture Collection Administration on November 13, 2024, with a deposit number of CGMCC No.32602, and was classified and named Streptomyces albulus.

[0006] The Streptomyces parvum SY6 provided by the present invention is obtained by screening Streptomyces parvum CICC 11022 through composite physical and chemical mutagenesis. The Streptomyces parvum SY6 screened by the present invention efficiently responds to mold signals and strengthens the synthesis of ε-polylysine, and can be effectively applied to direct addition of mold signals in fermentation or mixed fermentation of Streptomyces parvum and mold to improve the synthesis yield of ε-polylysine.

[0007] The invention provides a fermentation seed of Streptomyces parvum SY6, wherein the fermentation seed of Streptomyces parvum SY6 is obtained by inoculating spores of Streptomyces parvum SY6 into an ε-polylysine fermentation seed culture medium and culturing for 1d to 3d under the conditions of 25°C to 35°C and 180rpm to 220rpm.

[0008] The present invention also provides an application of the Streptomyces parvus SY6 in synthesizing ε-polylysine.

[0009] Furthermore, the Streptomyces parvus SY6 is used to increase the yield of ε-polylysine.

[0010] Furthermore, the Streptomyces parvus SY6 is used to increase the concentration of ε-polylysine.

[0011] The present invention provides a method for synthesizing ε-polylysine by Streptomyces parvum SY6, comprising the following steps:

[0012] Prepare ε-polylysine fermentation medium, sterilize, add glucose solution, inoculate the fermentation seeds of Streptomyces parviflorus SY6, ferment at an initial pH of 6.0-7.5 and a temperature of 25°C-35°C, and use sterilized ammonia water to control the pH to 3.0-5.0 until the fermentation is completed; when the glucose concentration in the fermentation broth is less than 10 g / L, maintain the glucose concentration in the fermentation broth at 10 g / L-20 g / L, and when the NH4 + When the -N concentration is lower than 0.5g / L, the NH4 + The -N concentration is controlled in the range of 0.5 g / L to 1.0 g / L; the fermentation is terminated when the ε-polylysine concentration in the fermentation broth no longer increases.

[0013] Furthermore, the fermentation seeds of Streptomyces parvum SY6 are inoculated at an inoculation amount of 5 g / 100 mL to 10 g / 100 mL.

[0014] Furthermore, the fermentation liquid also includes live fungal cell liquid.

[0015] Furthermore, the live fungus liquid is one of a live Aspergillus niger liquid, a live Alternaria alternata liquid and a live Botrytis cinerea liquid.

[0016] Furthermore, the live fungal cell liquid is inoculated at an inoculation rate of 5 g / L to 20 g / L.

[0017] The culture medium formula used in the present invention is:

[0018] (1) 1 L Betana medium: 10 g glucose, 1 g yeast powder, 2 g peptone, 20 g agar, and the balance is distilled water, pH 7.5.

[0019] (2) 1 L ε-polylysine fermentation seed medium: glucose 20 g, yeast powder 5 g, (NH4)2SO4 10 g, MgSO4·7H2O 0.g5, K2HPO4·3H2O 0.8 g, KH2PO4 1.36 g, FeSO4·7H2O 0.03 g, ZnSO4·7H2O 0.04 g, the balance is distilled water, pH 6.8.

[0020] (3) 1L ε-polylysine fermentation medium (g / L): glucose 60 g, yeast powder 5 g, (NH4)2SO4 10 g, MgSO4·7H2O 0.5 g, K2HPO4·3H2O 0.8 g, KH2PO4 1.36 g, FeSO4·7H2O 0.03 g, ZnSO4·7H2O 0.04 g, the remainder is distilled water, pH 6.8.

[0021] (4) 1LYPD liquid medium: 20 g glucose, 10 g yeast extract, 20 g peptone, and the balance distilled water, pH 6.5.

[0022] For all the above cases involving glucose, the sugar-containing substances were prepared separately from other ingredients, sterilized at 121°C for 20 min, and then combined into the same system culture medium.

[0023] The molds described in the present invention include Alternaria alternate CICC 40292, Aspergillus niger CICC 40102, Penicillium chrysogenum CICC 40702, and Rhodotorulamucilaginosa CICC 32140 purchased from China Industrial Microbiological Culture Collection Administration Center; Botrytis cinerea CGMCC 3.3790 purchased from China General Microbiological Culture Collection Administration Center; Streptomyces albulus CICC 11022 purchased from China Industrial Microbiological Culture Collection Administration Center. Description of biological material preservation:

[0024] Streptomyces albulus SY6, referred to as Streptomyces albulus SY6 in the present invention, has been deposited in the General Microbiology Center of China Microorganism Culture Collection Administration on November 13, 2024, with a deposit number of CGMCC No.32602. The address of the deposit unit is No. 3, Yard No. 1, Beichen West Road, Chaoyang District, Beijing, Postal Code: 100101, and is classified and named Streptomyces albulus.

[0025] Compared with the prior art, the present invention has the following beneficial effects:

[0026] The Streptomyces parvuligerus SY6 provided by the present invention is obtained by composite physical and chemical mutagenesis of Streptomyces parvuligerus CICC 11022. The Streptomyces parvuligerus SY6 provided by the present invention can directly add fermentation to synthesize ε-polylysine, and the fermentation yield is 2.25 times that of Streptomyces parvuligerus CICC 11022; in addition, it can also efficiently respond to mold signals and promote the increase of ε-polylysine production, among which the mixed fermentation yield of Streptomyces parvuligerus SY6 and live Alternaria alternata is the highest, which is 3.31 times that of Streptomyces parvuligerus CICC 11022. BRIEF DESCRIPTION OF THE DRAWINGS

[0027] In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the drawings required for use in the embodiments or the description of the prior art will be briefly introduced below. Obviously, the drawings described below are only some embodiments of the present invention. For ordinary technicians in this field, other drawings can be obtained based on these drawings without paying creative work.

[0028] Figure 1 The figure shows the screening of high-producing bacteria of ε-polylysine based on the size of the plate inhibition zone. In the figure, red yeast is used as an indicator bacteria to screen high-producing bacteria. A is the front side of the plate; B is the back side of the plate. The numbers and the circles inside them are high-producing bacteria that produce larger inhibition zones.

[0029] Figure 2 It is the Streptomyces parvus SY6 that produces high ε-polylysine. DETAILED DESCRIPTION

[0030] The specific embodiments of the present invention are described in detail below, but it should be understood that the protection scope of the present invention is not limited by the specific embodiments. Based on the embodiments in the present invention, all other embodiments obtained by ordinary technicians in this field without creative work belong to the scope of protection of the present invention. The experimental methods described in the embodiments of the present invention are conventional methods unless otherwise specified, and the materials, reagents, etc. used in the following embodiments can be obtained from commercial channels unless otherwise specified.

[0031] Example 1: Breeding method of Streptomyces parvum SY6

[0032] S1, preparation of a single spore suspension of Streptomyces molitor CICC 11022: spores of Streptomyces molitor CICC 11022 were spread on a Betana solid medium, cultured at 30°C for 8 days, washed and collected with pre-sterilized 20% glycerol, and after adding glass beads, placed in an oscillating incubator at 200 rpm for 5 hours, and then oscillated in an ultrasonic cleaner for 20 minutes to fully disperse the spores. Finally, the single spore suspension of Streptomyces molitor CICC 11022 was filtered 5 times with pre-sterilized cotton wool, and the filtered single spore suspension of Streptomyces molitor CICC 11022 was aliquoted and stored in a -80°C refrigerator for later use.

[0033] Preparation of fungal signal extract: scrape 5 rings (about 1×10 3 The spores of the mold (Botrytis cinerea) were inoculated into a 500 mL Erlenmeyer flask containing 80 mL of YPD medium and cultured in a constant temperature shaking incubator at 200 rpm at 30°C for 4 days.

[0034] Scrape 5 rings (about 1×10 3 ) Botrytis cinerea spores were inoculated into a 500 mL Erlenmeyer flask containing 80 mL YPD medium and cultured in a constant temperature shaking incubator at 200 rpm at 23° C. for 4 days.

[0035] The cultured fungus was washed with deionized water for 3 times, resuspended with 65% ethanol, and the mixture was placed in a mortar and ground with quartz sand for 10 minutes. The supernatant was then centrifuged and dried in a forced air drying oven and then reconstituted with water. The signal extract of Streptomyces parvum was obtained after high temperature sterilization at 121°C for 15 minutes and stored at -20°C. The volume ratio of the wet weight of the Streptomyces parvum cells to the reconstituted water was 1g:3mL.

[0036] S2, 1L strain selection medium: Beta medium containing 0.05g LiCl and 120mL mold signal extract.

[0037] Prepare the mixed solution: 2.5 μL of 0.5 mg / mL NTG (nitrosoguanidine), 0.5 mL of 5 g / L (pH = 6) citric acid buffer and 0.5 mL of about 5.0 × 10 7 The mixture was mixed with a single spore suspension of Streptomyces microalbuminus CICC 11022 and incubated at 30°C in a constant temperature shaking incubator at 100 rpm for 60 min. 5 times.

[0038] Spreading culture: dilute to 10 -5 Take 1 mL of the mixed solution and spread it on the strain selection medium prepared in S2, and culture it in a constant temperature incubator at 30°C for 8 days until colonies grow out one after another.

[0039] S3, pre-cultured red yeast: obtained by culturing in YPD liquid medium at 30°C and 180 rpm for 1 day.

[0040] In the clean bench, lift one side of the strain breeding medium and slide it in a circular motion to release the adhesion between the strain breeding medium and the culture dish, lift the strain breeding medium, inject 1 mL of pre-cultured red yeast into the gap between the strain breeding medium and the culture dish, rotate the strain breeding medium so that the red yeast is evenly distributed in the gap between the strain breeding medium and the culture dish, and place the system in a 30°C incubator for 1 day to obtain the first round of mutagenesis strains.

[0041] Observe the size of the red yeast inhibition zone of the first round of mutagenesis strains on the colony of the strain selection medium below the agar block, and compare it with the inhibition zone corresponding to the first round starting strain Streptomyces parvum CICC 11022. Pick out the strains with significantly larger inhibition zones for streaking purification and expansion culture to obtain the first round of high-yield strain spores, such as Figure 1 shown.

[0042] S4, use an inoculation loop to scrape two rings of the first round of high-yield bacterial strain spores, inoculate them into a 250mL conical flask containing 30mL ε-polylysine fermentation seed medium, and culture them in a constant temperature shaking incubator at 30°C and 200rpm for 2d; inoculate the high-yield bacterial spores into a 250mL conical flask containing 30mL ε-polylysine fermentation medium at an inoculation amount of 8g / 100mL, and culture them in a constant temperature shaking incubator at 30°C and 200rpm for 2d. Among them, when the fermentation pH naturally drops to 4.0, add pre-sterilized sodium citrate buffer (pH 4.0) to the triangular flask in the clean bench so that the final concentration of citric acid in the triangular flask is 10g / L to maintain a constant pH. After the fermentation is completed, the ε-polylysine concentration in the fermentation broth is measured to obtain the optimal strain of Streptomyces SY1 for the first round of mutagenesis.

[0043] S5, using Streptomyces parvum SY1 as the second round starting strain, a single spore suspension of Streptomyces parvum SY1 was prepared according to the method described in S1.

[0044] S6, the concentration is about 1.0×10 7 Spores of Streptomyces parvum SY1 at 100 / mL were spread on the strain selection medium, and the above plates were irradiated at 30 cm under a 15 W ultraviolet lamp in a clean bench for 1 min, and then placed in a constant temperature incubator at 30°C for 8 days, until colonies grew out one after another.

[0045] S7, according to S3, screen and obtain a strain with an inhibition zone larger than that of the second round starting strain Streptomyces parvum SY1, and according to the steps in S4, screen and obtain the Streptomyces parvum SY2 strain with the highest yield.

[0046] S8, the Streptomyces SY2 was repeatedly subjected to nitrosoguanidine and UV mutagenesis according to the above method, and the plate with mold signal added was used as the screening condition and the yeast inhibition zone was used as the screening method to select strains that efficiently responded to Streptomyces signals and promoted the synthesis of ε-polylysine. After repeated 3 rounds of mutagenesis, the Streptomyces SY6 strain with high ε-polylysine production was obtained, such as Figure 2 shown.

[0047] S. parvum SY6 was serially subcultured and its ε-polylysine synthesis ability was identified in the following manner: spores of S. parvum CICC 11022 and SY6 (approximately 1×10 7 ) were inoculated into a 500mL Erlenmeyer flask containing 60mL of liquid seed culture medium, and cultured in a constant temperature shaking incubator at 220rpm at 30°C for 2 days until a large number of granular mycelial balls were formed; the mature fermentation seed liquid was added to a 250mL Erlenmeyer flask containing 30mL of ε-polylysine fermentation medium at an inoculum amount of 8g / 100mL, and then the flask was placed in a constant temperature shaking incubator and cultured at 30°C and 200rpm for 1 day, and a pre-sterilized sodium citrate buffer (pH=4.0) with a final concentration of 10g / L was added to the flask in an ultra-clean bench to maintain the pH, and the flask was returned to the incubator and continued to be cultured in the original manner for 1 day. After the end, the polylysine concentration in the fermentation broth was detected.

[0048] Detection of ε-polylysine concentration in fermentation broth: The fermentation broth was appropriately diluted with 0.2 mM sodium phosphate buffer (pH = 7.0), 2 mL was taken out and mixed with 2 mL 1 mM methyl orange aqueous solution, placed at 30 ° C for 30 min, centrifuged at 4000 rpm for 15 min, and the supernatant was diluted 20 times with 0.2 mM sodium phosphate buffer (pH = 7.0) and the absorbance was measured at 465 nm in a spectrophotometer. The absorbance was substituted into the ε-polylysine concentration standard curve to calculate the actual ε-polylysine concentration in the fermentation broth sample.

[0049] The ε-polylysine shake flask fermentation levels of Streptomyces parviflorus SY6 after 10 consecutive passages are as follows:

[0050] Table 1 ε-polylysine shake flask fermentation level (g / L) of Streptomyces SY6 strain after 10 consecutive passages

[0051]

[0052]

[0053] NOTE: The "added signal" for S6 will be approximately 1.0 × 10 7 Spores of Streptomyces parvum were spread on the strain selection medium; “no signal added” means: about 1.0×10 7Spores of Streptomyces parvum strain were spread on the strain breeding medium.

[0054] Example 2: Synthesis of ε-polylysine in a 5L fermenter based on Streptomyces parvum SY6

[0055] S. albus SY6 fermentation seeds: scrape 3 rings of S. albus SY6 spores (about 1×10 7 ) were inoculated into a 500-mL Erlenmeyer flask filled with 60 mL of liquid seed culture medium, and cultured in a constant temperature shaking incubator at 30° C. and 200 rpm for 2 days until a large number of granular mycelial balls were formed to obtain fermentation seeds of Streptomyces parvus SY6.

[0056] Prepare 3L ε-polylysine fermentation medium, add it to a 5L liquid fermentation tank, sterilize it at 121℃ for 20min, and use a peristaltic pump to add all the 500mL solution of 240g glucose that was sterilized separately (sterilized at 121℃ for 20min), set the temperature to 30℃, the stirring paddle speed to 200rmp and the ventilation volume to 1vvm as the initial parameters. Inoculate the obtained Streptomyces SY6 fermentation seeds with an inoculum of 8g / 100mL, adjust the pH to 6.8 with 12.5% ​​ammonia water to start fermentation. When the pH naturally drops to 4.0, dynamically add 12.5% ​​ammonia water to maintain pH = 4.0, and use the speed and dissolved oxygen coupling setting to control the dissolved oxygen at 33% until the fermentation ends at 192h. When the glucose concentration in the fermentation broth is lower than 10g / L, add 600g / L of pre-sterilized glucose solution through a peristaltic pump exogenous flow to maintain the fermentation broth concentration range at 15g / L. When NH4 + When the -N concentration is lower than 0.5g / L, 200g / L (NH4)2SO4 solution is added through a peristaltic pump to remove NH4 + -N concentration was controlled within the range of 0.75 g / L. During the fermentation process, the fermentation broth was sampled every 12 h, and the ε-polylysine concentration, bacterial dry weight, residual glucose, and NH4 in the fermentation broth were determined according to the process described in the determination method. + -N concentration.

[0057] Detection of ε-polylysine concentration in fermentation broth: The fermentation broth was appropriately diluted with 0.2 mM sodium phosphate buffer (pH = 7.0), 2 mL was taken out and mixed with 2 mL 1 mM methyl orange aqueous solution, placed at 30 ° C for 30 min, centrifuged at 4000 rpm for 15 min, and the supernatant was diluted 20 times with 0.2 mM sodium phosphate buffer (pH = 7.0) and the absorbance was measured at 465 nm in a spectrophotometer. The absorbance was substituted into the ε-polylysine concentration standard curve to calculate the actual ε-polylysine concentration in the fermentation broth sample.

[0058] Glucose concentration in fermentation broth: 300 μL of fermentation broth was mixed with 700 μL of anhydrous ethanol, and the mixture was centrifuged after standing for 1 h to obtain the supernatant, and then filtered through a 0.45 μm filter membrane for HPLC detection. An organic acid test column (Aminex HPX-87H, 300×7.8 mm; Hercules, USA) was used, the mobile phase was set to 5 mM sulfuric acid, the column temperature was controlled at 60°C, and the injection volume was 10 μL.

[0059] The dry cell weight (DCW) was determined by the filter paper differential gravimetric method. 10 mL of fermentation broth was taken out from a 5 L fermenter, centrifuged at 4,000 × g for 10 min, the precipitate was washed twice with distilled water, and filtered with filter paper (Φ7 cm, medium speed, SCRC) that had been previously dried at 105 ° C and weighed. The filter paper was then placed at 105 ° C and dried to constant weight and weighed. The weight difference before and after was calculated, and the dry cell weight in the fermentation broth was calculated based on the volume of 10 mL of the original fermentation broth.

[0060] After the final fermentation, the ε-polylysine concentration of Streptomyces SY6 was 20.9 g / L, and the dry weight of the bacteria was 28.2 g / L. The starting strain Streptomyces CICC 11022 was cultured with the same fermentation process, and its final ε-polylysine concentration was 9.3 g / L, and the dry weight of the bacteria was 24.6 g / L. The yield of ε-polylysine synthesized by the high-yield strain Streptomyces SY6 was 2.25 times that of the starting strain Streptomyces CICC 11022.

[0061] Example 3: Synthesis of ε-polylysine in a 5L fermentation tank based on Streptomyces SY6 and various fungal signals

[0062] Prepare 3Lε-polylysine fermentation medium, add it to a 5L liquid fermentation tank, sterilize it at 121℃ for 20min, and add 500mL of a solution of 240g glucose sterilized separately (sterilized at 121℃ for 20min) with a peristaltic pump, set the temperature to 30℃, the stirring paddle speed to 200rmp and the ventilation volume to 1vvm as initial parameters. The fermentation seed liquid of Streptomyces SY6 obtained in Example 2 was inoculated with an inoculum of 8g / 100mL, and the pH was adjusted to 6.8 with 12.5% ​​ammonia water to start fermentation. When the fermentation was 24h, 500mL of the mold signal extract described in S1 in Example 1 was added to the fermentation tank from the feed bottle at one time. When the pH dropped to 4.0 naturally, 12.5% ​​ammonia water was added dynamically to maintain pH 4.0, and the dissolved oxygen was regulated at 30-35% by the speed and dissolved oxygen coupling setting until the fermentation was completed at 192h. When the glucose concentration in the fermentation broth was lower than 10 g / L, 600 g / L of pre-sterilized glucose solution was added through a peristaltic pump to maintain the fermentation broth concentration at 15 g / L. +When the -N concentration is lower than 0.5g / L, 200g / L (NH4)2SO4 solution is added through a peristaltic pump to remove NH4 + -N concentration was controlled within the range of 0.75 g / L. During the fermentation process, the fermentation broth was sampled every 12 h, and the ε-polylysine concentration, bacterial dry weight, residual glucose, and NH4 in the fermentation broth were determined according to the process described in the determination method. + -N concentration.

[0063] The detection was performed according to the detection method in Example 2. Finally, Streptomyces SY6 was used: (1) Aspergillus niger signal was added to the ε-polylysine fed-batch fermentation. After 192 hours of fermentation, the ε-polylysine concentration was 48.7 g / L and the dry weight of the bacteria was 43.6 g / L. The starting strain Streptomyces CICC 11022 was cultured using the same fermentation process, and its final ε-polylysine concentration was 16.2 g / L and the dry weight of the bacteria was 36.2 g / L. Under the condition of exogenous addition of Aspergillus niger signal, the ε-polylysine fed-batch fermentation yield of Streptomyces SY6 was 2.85 times that of the starting strain Streptomyces CICC 11022.

[0064] (2) Alternaria signal was added to the ε-polylysine fed-batch fermentation. After 192 h of fermentation, the ε-polylysine concentration was 51.2 g / L and the dry weight of the bacteria was 46.8 g / L. The starting strain Streptomyces parthenogeneticus CICC11022 was cultured using the same fermentation process, and its final ε-polylysine concentration was 17.9 g / L and the dry weight of the bacteria was 37.4 g / L. Under the condition of exogenous addition of Alternaria signal, the ε-polylysine fed-batch fermentation yield of Streptomyces parthenogeneticus SY6 was 2.86 times that of the starting strain Streptomyces parthenogeneticus CICC11022.

[0065] (3) In the fed-batch fermentation of ε-polylysine, the Botrytis cinerea signal was added. After 192 h of fermentation, the ε-polylysine concentration was 43.2 g / L and the dry weight of the bacteria was 40.1 g / L. The starting strain Streptomyces parthenolides CICC11022 was cultured using the same fermentation process, and its final ε-polylysine concentration was 13.8 g / L and the dry weight of the bacteria was 32.2 g / L. Under the condition of exogenous addition of Botrytis cinerea signal, the ε-polylysine fed-batch fermentation yield of Streptomyces parthenolides SY6 was 3.13 times that of the starting strain Streptomyces parthenolides CICC11022.

[0066] (4) When the Penicillium chrysogenum signal was added to the ε-polylysine fed-batch fermentation, the ε-polylysine concentration was 49.2 g / L and the dry weight of the bacteria was 44.3 g / L after 192 h of fermentation. The starting strain Streptomyces parthenolides CICC11022 was cultured with the same fermentation process, and its final ε-polylysine concentration was 18.8 g / L and the dry weight of the bacteria was 39.1 g / L. Under the condition of exogenous addition of the Penicillium chrysogenum signal, the ε-polylysine fed-batch fermentation yield of Streptomyces parthenolides SY6 was 2.62 times that of the starting strain. The above data show that Streptomyces parthenolides SY6 can respond to these four different fungal signals together and efficiently, and greatly enhance the biosynthesis of ε-polylysine under signal activation.

[0067] Example 4: Synthesis of ε-polylysine in a 5L fermentation tank based on a mixed bacterial system of mold and Streptomyces parvum SY6

[0068] Preparation of live fungal cell suspension: scrape 5 rings (about 1×10 3 The spores of the fungus (Botrytis cinerea) were inoculated into a 500 mL Erlenmeyer flask containing 80 mL of YPD medium and cultured in a constant temperature shaking incubator at 200 rpm at 30°C for 4 days.

[0069] Scrape 5 rings (about 1×10 3 ) Botrytis cinerea spores were inoculated into a 500 mL Erlenmeyer flask containing 80 mL YPD medium and cultured in a constant temperature shaking incubator at 200 rpm at 23° C. for 4 days.

[0070] Prepare 3Lε-polylysine fermentation medium, add it to a 5L liquid fermentation tank, sterilize it at 121℃ for 20min, and add 500mL of a solution of 240g glucose sterilized separately (sterilized at 121℃ for 20min) with a peristaltic pump, set the temperature to 30℃, the stirring paddle speed to 200rmp and the ventilation volume to 1vvm as initial parameters. The fermentation seeds of Streptomyces SY6 obtained in Example 2 were inoculated with an inoculum of 8g / 100mL, and the pH was adjusted to 6.8 with 12.5% ​​ammonia water to start fermentation. At 24h of fermentation, under the protection of the fire ring, 500mL of a live bacterial culture of one of the four molds obtained was added to the fermentation tank from the inoculation port. When the pH naturally dropped to 4.0, 12.5% ​​ammonia water was added dynamically to maintain pH 4.0, and the dissolved oxygen was regulated at 33% by the speed and dissolved oxygen coupling setting until the fermentation ended at 192h. When the glucose concentration in the fermentation broth was lower than 10 g / L, 600 g / L of pre-sterilized glucose solution was added through a peristaltic pump to maintain the fermentation broth concentration at 15 g / L. + When the -N concentration is lower than 0.5g / L, 200g / L (NH4)2SO4 solution is added through a peristaltic pump to remove NH4 +-N concentration was controlled at 0.75 g / L. During the fermentation process, the fermentation broth was sampled every 12 h, and the ε-polylysine, dry weight of bacteria, residual glucose, and NH4 in the fermentation broth were determined according to the process described in the determination method. + -N concentration.

[0071] The detection was performed according to the detection method in Example 2. Finally, Streptomyces parvum SY6 was used: (1) A culture solution containing live Aspergillus niger was added to the ε-polylysine fed-batch fermentation. After 192 hours of fermentation, the ε-polylysine concentration was 60.2 g / L and the dry weight of the bacteria was 45.1 g / L. The starting strain Streptomyces parvum CICC 11022 was cultured using the same fermentation process, and its final ε-polylysine concentration was 20.5 g / L and the dry weight of the bacteria was 38.5 g / L. Under the condition of exogenous addition of live Aspergillus niger culture solution, the ε-polylysine fed-batch fermentation yield of Streptomyces parvum SY6 was 2.93 times that of the starting strain Streptomyces parvum CICC 11022.

[0072] (2) When the culture medium containing live Alternaria alternata was added to the ε-polylysine fed-batch fermentation, the ε-polylysine concentration was 65.9 g / L and the dry weight of the bacteria was 44.3 g / L after 192 h of fermentation. The starting strain Streptomyces parthenogeneticus CICC 11022 was cultured using the same fermentation process, and its final ε-polylysine concentration was 19.9 g / L and the dry weight of the bacteria was 39.4 g / L. Under the condition of exogenous addition of Alternaria alternata signal, the ε-polylysine fed-batch fermentation yield of Streptomyces parthenogeneticus SY6 was 3.31 times that of the starting strain Streptomyces parthenogeneticus CICC 11022.

[0073] (3) In the fed-batch fermentation of ε-polylysine, the Botrytis cinerea signal was added. After 192 h of fermentation, the ε-polylysine concentration was 49.2 g / L and the dry weight of the bacteria was 44.6 g / L. The starting strain Streptomyces parthenolides CICC11022 was cultured using the same fermentation process, and its final ε-polylysine concentration was 16.3 g / L and the dry weight of the bacteria was 37.2 g / L. Under the condition of exogenous addition of Botrytis cinerea signal, the ε-polylysine fed-batch fermentation yield of Streptomyces parthenolides SY6 was 3.02 times that of the starting strain Streptomyces parthenolides CICC11022.

[0074] (4) When the Penicillium chrysogenum signal was added to the ε-polylysine fed-batch fermentation, the ε-polylysine concentration was 62.9 g / L and the dry weight of the bacteria was 47.2 g / L after 192 h of fermentation. The starting strain Streptomyces parthenolides CICC11022 was cultured with the same fermentation process, and its final ε-polylysine concentration was 21.2 g / L and the dry weight of the bacteria was 43.4 g / L. Under the condition of exogenous addition of the Penicillium chrysogenum signal, the ε-polylysine fed-batch fermentation yield of Streptomyces parthenolides SY6 was 2.97 times that of the starting strain Streptomyces parthenolides CICC11022. The above data show that Streptomyces parthenolides SY6 can respond to these four different fungal signals together and efficiently, and can greatly enhance the level of ε-polylysine biosynthesis under the effect of coexistence with fungi.

[0075] Although preferred embodiments of the present invention have been described, additional changes and modifications may occur to these embodiments once those skilled in the art are aware of the basic inventive concepts.

[0076] Obviously, those skilled in the art may make various changes and modifications to the present invention without departing from the spirit and scope of the present invention. Thus, if these modifications and variations of the present invention fall within the scope of the claims of the present invention and their equivalents, the present invention is also intended to include these modifications and variations.

Claims

1. A Streptomyces albulus SY6, characterized in that It was deposited in the General Microbiology Center of China Microorganism Culture Collection Administration on November 13, 2024, with the deposit number CGMCC No.32602 and the classification name Streptomyces albulus.

2. A fermented seed of Streptomyces parvum SY6 according to claim 1, characterized in that: The fermentation seeds of Streptomyces parvum SY6 are obtained by inoculating spores of Streptomyces parvum SY6 into an ε-polylysine fermentation seed culture medium and culturing for 1 to 3 days at 25° C. to 35° C. and 180 rpm to 220 rpm.

3. Use of the Streptomyces parvum SY6 according to claim 1 in the synthesis of ε-polylysine.

4. The use according to claim 3, characterized in that: The Streptomyces parvus SY6 is used for increasing the yield of ε-polylysine.

5. The use according to claim 3, characterized in that: The Streptomyces parvus SY6 is used to increase the concentration of ε-polylysine.

6. A method for synthesizing ε-polylysine by Streptomyces parvum SY6 according to claim 3, characterized in that: The steps include: Prepare ε-polylysine fermentation medium, sterilize, add glucose solution, inoculate the fermentation seeds of Streptomyces parviflorus SY6, ferment at an initial pH of 6.0-7.5 and a temperature of 25°C-35°C, and use sterilized ammonia water to control the pH to 3.0-5.0 until the fermentation is completed; when the glucose concentration in the fermentation broth is less than 10 g / L, maintain the glucose concentration in the fermentation broth at 10 g / L-20 g / L, and when the NH4 + When the -N concentration is lower than 0.5g / L, the NH4 + The -N concentration is controlled in the range of 0.5 g / L to 1.0 g / L; the fermentation is terminated when the ε-polylysine concentration in the fermentation broth no longer increases.

7. The method for synthesizing ε-polylysine by Streptomyces parvum SY6 according to claim 6, characterized in that: The fermentation seeds of Streptomyces parvum SY6 are inoculated at an inoculation rate of 5 g / 100 mL to 10 g / 100 mL.

8. The method for synthesizing ε-polylysine by Streptomyces parvum SY6 according to claim 6, characterized in that: The fermentation liquid also includes live fungal cell liquid.

9. The method for synthesizing ε-polylysine by Streptomyces parvum SY6 according to claim 8, characterized in that: The live fungus liquid is one of the live fungus liquid of Aspergillus niger, the live fungus liquid of Alternaria alternata and the live fungus liquid of Botrytis cinerea.

10. The method for synthesizing ε-polylysine by Streptomyces parvum SY6 according to claim 8, characterized in that: The live fungal cell liquid is inoculated at an inoculation rate of 5 g / L to 20 g / L.

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