CN115927057A - Streptomyces strain YP1 and application thereof - Google Patents

Abstract

The application relates to a streptomycete strain YP1 and application thereof, belonging to the technical field of microorganisms. A durocin-producing strain, designated YP1, is isolated from soil and uses of YP1 for producing durocin are provided. The streptomycete strain YP1 is preserved in the common microorganism center of China Committee for culture Collection of microorganisms, and the preservation registration number is CGMCC No.23383, and the preservation date is 2021, 9 months and 10 days.

Description

A kind of Streptomyces strain YP1 and its application

technical field

The application relates to a Streptomyces strain YP1 and its application, belonging to the technical field of microorganisms.

Background technique

Granaticin (CAS No.: 19879-06-2) is a member of the benzoisochromanequinones (BIQs) family of antibiotics and belongs to the class of aromatic polyketide antibiotics. Its molecular formula is C22H20O10, its molecular weight is 444.38800, its boiling point is 748.3°C at a standard atmospheric pressure, its flash point is 266.8°C, its melting point is between 223-225°C, its density is 1.77g/cm3 under normal conditions, and its refractive index is 1.745. Durantin is a dark red crystal, and its solubility varies in different solutions. It is soluble in polar solutions such as water, methanol and ethanol, but has no solubility in non-polar solutions, and its solubility in water is Affected by pH, it is positively correlated with pH value, and it is red in acidic aqueous solution and blue in alkaline aqueous solution.

Durantin was first discovered in Streptomyces olivaceus (S. olivaceus). Afterwards, durianin was successively isolated from Streptomyces cantonensis GIMV4.0001 (S.vietnamensisgIMV4.0001), Streptomyces violaceoruber and Streptomyces cerevisiae (S.lateritius). Simultaneously, duracin analogues, such as duracin B and dihydrodrugin, were also isolated from some strains.

Duracin can be used as a potential antibacterial and anticancer antibiotic, but the types of strains that can produce duracin are limited, which is far from meeting the actual needs.

Contents of the invention

In order to solve the technical problem that there are few types of strains capable of producing dunicin, and to enrich the types of strains capable of producing dunicin; this application isolated a strain capable of producing dunicin from the soil, named YP1, and provided Use of YP1 to produce duracicin applications.

The strain YP1 provided in this application is isolated from the soil collected in Zhengzhou City, Henan Province, China (34°48'00.00"N, 113°39'00.00"E). Specifically, the soil collected from Zhengzhou City, Henan Province, China (34°48'00.00"N, 113°39'00.00"E) was prepared as a soil suspension, then diluted to 10-3, and spread on the Gaoshi culture cultured on basal plates at 28°C. The strains with color-producing characteristics were taken and examined under a microscope. After being proved to be a single strain, the YP1 strain was obtained through repeated isolation and repeated culture and screening of a large number of strains. The Streptomyces strain YP1 of this application has been preserved in the General Microbiology Center of the China Microbiological Culture Collection Management Committee. The preservation registration number is CGMCC No. 23383, and the preservation date is September 10, 2021.

The morphological characteristics of the strain YP1 provided by the application:

The colony of this strain is milky white or light yellow, sub-round, mainly opaque, and the surface of the colony is smooth or slightly raised. The spore filaments are straight, flexible, and spiral; the surface of the spores is smooth and powdery

In addition, the strain YP1 of the present application is an aerobic gram-positive bacterium; the culture conditions are: pH 5-8, 20-40°C; among them, the optimum pH is 7.0, and the optimum temperature is 28°C; NaCl resistant Receptivity 0-5% (w/v).

The physiological and biochemical characteristics of the strain YP1 provided by the application:

The strain has catalase, oxidase, β-galactosidase and arginine dihydrolase activities, but does not have lysine, ornithine decarboxylase and urease activities. It can hydrolyze gelatin and reduce nitrate, but can't hydrolyze Tween 40, can't produce H ₂ S and indole, VP reaction and methyl red test are negative, and can't use citric acid. Dextrin, maltose, trehalose, sucrose, stachyose, raffinose, lactose and other carbon sources can be used, but fucose, sorbitol, mannitol, arabitol, and inositol cannot be used as carbon sources. Cannot use hydroglycoside, serine, alanine, arginine, aspartic acid, but can use pectin, galacturonic acid, L-galactolide, gluconic acid, quinic acid, γ-aminobutyric acid acid, α-hydroxybutyric acid, β-hydroxy-D,L-butyric acid, D-glucose-6-phosphate, glucuronamide, D-fructose-6-phosphate, mucic acid, p-hydroxybenzene Acetic Acid, Methyl Pyruvate, D-Methyl Lactate, L-Lactic Acid, Citric Acid, Alpha-Ketoglutarate, D-Malic Acid, L-Malic Acid. Sensitive to troleandomycin, rifamycin SV, minocycline, vancomycin.

The genetic characteristics of the strain YP1 provided by the application:

The dDH values of the close-type strain Streptomyces zaomyceticus and the close-type strain Streptomyces lateritius were 28.5% and 33.9%, respectively; far below the 70% threshold of bacterial species;

The ANI values of Streptomyces zaomyceticus and Streptomyces lateritius were 83.95% and 87.56%, respectively, far below the 95-96% threshold for demarcating species; is a new Streptomyces species.

According to the analysis of morphological characteristics, physiological and biochemical characteristics and genetic characteristics, YP1 strain conforms to the characteristics of Streptomyces, belongs to Streptomyces, and is a new strain of Streptomyces (Latin Streptomyces vilmorinianum).

The application also provides the application of Streptomyces strain YP1 in producing dunicin.

Streptomyces bacterial strain YP1 of the present application produces dunicin, comprises the following steps:

(1) Plate culture: Streak the strain YP1 on the plate solid medium and culture it at a constant temperature of 36-38°C for 1-3 days;

(2) Seed liquid culture: Pick the well-grown YP1 bacteria from the plate culture medium, insert it into the high-temperature sterilized proliferation medium, and put it in a constant temperature shaker for 1-3 days, and the shaker speed is 170 -190r/min, culture temperature is 36-38°C; obtain strain YP1 seed liquid;

(3) Fermentation culture: insert the seed liquid of bacterial strain YP1 into the fermentation medium sterilized by high temperature with the inoculation amount of 4-6% v/v, put the fermentation medium into a constant temperature shaker and cultivate it for 3-5 days, The shaker speed is 170-190r/min, and the culture temperature is 36-38°C; the blue strain YP1 fermentation broth is obtained;

(4) centrifuging, rotary steaming, concentrating and purifying the fermentation broth of strain YP1 to obtain duracin;

Plate medium: 20g of agar, 10g of lactose, 5g of tryptone, 10g of yeast extract powder, 4g of NacL and 1000ml of distilled water, pH=7.0, sterilized at 121°C for 20min;

Proliferation medium: lactose 10g, tryptone 5g, yeast extract powder 10g, NacL 4g and distilled water 1000ml, pH=7.0, sterilized at 121°C for 20min;

Fermentation medium: lactose 10g, tryptone 5g, yeast extract powder 10g, NacL 4g and distilled water 1000ml, pH=7.0, sterilized at 121°C for 20min.

In some embodiments, the culture condition of the step (1) is: culture at a constant temperature of 37° C. for 2 days; in some embodiments, the culture condition of the step (2) is: culture in a constant temperature shaker for 2 days, The rotation speed of the shaker is 180r/min, and the culture temperature is 37°C;

In some embodiments, the culture conditions of the step (3) are: culture in a constant temperature shaker for 4 days, the speed of the shaker is 180r/min, and the culture temperature is 37°C.

In some embodiments, the purification includes the following operations:

(1) Purify the sample obtained by rotary evaporation and concentration with a macroporous adsorption resin, select the ethanol-water system to wash, and collect the samples washed out with 90% and 100% concentration ethanol as crude substance A;

(2) Dissolve the crude substance A in 40% methanol, filter the upper half to prepare a high performance liquid chromatograph, select the methanol-water system to elute the sample, and collect the sample washed out by 60% methanol;

Said % is a mass percentage.

Beneficial effect

The Streptomyces strain YP1 provided by this application is a new strain under the genus Streptomyces, which enriches the species of Streptomyces;

The Streptomyces strain YP1 provided in this application can produce duracin, which provides a new way for the acquisition of natural duracin;

The Streptomyces strain YP1 provided in the present application can produce dunicin with a yield of 180 U/mL.

deposit information

Preservation unit: General Microbiology Center of China Committee for Microorganism Culture Collection

Address: Institute of Microbiology, Chinese Academy of Sciences, No. 3, Yard 1, Beichen West Road, Chaoyang District, Beijing

Deposit date: September 10, 2021

Deposit number: CGMCC No.23383

Taxonomic designation: Streptomyces vilmorinianum.

Description of drawings

Fig. 1 is the colony morphology photo of Streptomyces strain YP1 provided by the application on the ISP2 medium;

Fig. 2 is the colony morphology photo of Streptomyces strain YP1 provided by the application on the ISP3 medium;

Fig. 3 is the colony morphology photo of Streptomyces strain YP1 provided by the application on the ISP4 medium;

Fig. 4 is the colony morphology photo of Streptomyces strain YP1 provided by the application on the ISP5 medium;

Figure 5 is a photo of the colony morphology of Streptomyces strain YP1 provided by the application on the ISP6 medium;

Figure 6 is a photo of the colony morphology of Streptomyces strain YP1 provided by the application on the ISP7 medium;

Fig. 7 is the sequence alignment of the gene sequence of the strain YP1 of the present application through Genebank, and the establishment of a phylogenetic tree using MEGA6.0.

Detailed ways

In the specific examples described below, unless otherwise specified, all are commonly used methods in the art. If no specific technique or condition is indicated in the examples, it shall be carried out according to the technique or condition described in the literature in this field or according to the product specification. The reagents or instruments used were not indicated by the manufacturer, and they were all commercially available conventional products. In the present application, unless otherwise specified, the stated % is a mass percentage.

Explanation of abbreviations used in this application:

d: day;

dDDH value: Genome data hybridization value (digital DNA–DNA hybridization)

ANI value: average nucleotide identity (average nucleotide identity)

MEGA6: Software - Molecular Evolutionary Genetics Analysis Version 6.0 (Molecular Evolutionarygenetics Analysis)

MK-n(Hm): Menaquinone, n represents the number of isoprene units, m represents the number of hydrogen atoms replacing isoprene

anteiso-c18:0: 14-methyl-hexadecanoic acid

anteiso-C16:0: 12-methyl-tetradecanoic acid

iso-C17:0: 14-methyl-pentadecanoic acid

The various culture media used in this application and its composition, pH are as follows:

ISP2 medium: yeast extract powder 4g, malt extract 10g, glucose 4g, deionized water 1L and solid plus agar powder 20g, pH=7.3;

ISP3 medium: oat flour 20g, agar 18g, ferrous sulfate heptahydrate 0.001g, manganese chloride 0.001g, zinc sulfate 0.001g and deionized water 1L, pH=7.3±0.2;

ISP4 medium: soluble starch 10g, potassium dihydrogen phosphate 1g, magnesium sulfate 1g, sodium chloride 1g, ammonium sulfate 2g, calcium sulfate 2g, ferrous sulfate heptahydrate 0.001g, manganese chloride 0.001g, zinc sulfate 0.001g, Agar 15g and deionized water 1L, pH=7.2±0.2;

ISP5 medium: aspartic acid 1g, potassium dihydrogen phosphate 1g, ferrous sulfate heptahydrate 0.001g, manganese chloride 0.001g, zinc sulfate 0.001g, agar 15g and deionized water 1L, pH=7.4±0.2;

胨5g、酵母浸粉1g、柠檬酸铁铵0.5g、磷酸氢二钾1g、硫代硫酸钠0.08g、琼脂15g和去离子水1L，pH＝6.7±0.2；ISP6 medium: peptone 15g,

5g of peptone, 1g of yeast extract powder, 0.5g of ferric ammonium citrate, 1g of dipotassium hydrogen phosphate, 0.08g of sodium thiosulfate, 15g of agar and 1L of deionized water, pH=6.7±0.2;

ISP7 medium: glycerol 15g, tyrosine 0.5g, aspartic acid 1g, dipotassium hydrogen phosphate 0.5g, magnesium sulfate 0.5g, sodium chloride 0.5g, ferrous sulfate 0.5g, ferrous sulfate heptahydrate 0.001g , manganese chloride 0.001g, zinc sulfate 0.001g, agar 20g and deionized water 1L, pH=7.3±0.1;

Plate (slant) medium: agar 20g, lactose 10g, tryptone 5g, yeast extract powder 10g, NaCL 4g and distilled water 1000ml, pH=7.0, sterilized at 121°C for 20min;

Proliferation/fermentation medium: lactose 10g, tryptone 5g, yeast extract powder 10g, NacL 4g and distilled water 1000ml, pH=7.0, sterilized at 121°C for 20min;

LB medium: Trypsin 10g, yeast extract powder 5g, NacL 10g and distilled water 1000ml, natural pH, sterilized at 115°C for 30min;

YPD medium: glucose 20, tryptone 20, yeast extract powder 10 and distilled water 1000ml, natural pH, sterilized at 115°C for 30min;

The identification of embodiment 1 bacterial strain YP1

1.1 Morphological features

The obtained pure culture of strain YP1 was inoculated into ISP2, ISP3, ISP4, ISP5, ISP6 and ISP7 culture medium respectively, and the morphological characteristics were observed: the results are as follows:

ISP2 medium, 28°C, cultured for 2 days; the colony is milky white, almost round, opaque, the surface of the colony is slightly raised upwards, and no pigment is secreted; the morphology of the spore filaments and spores is shown in Figure 1 when observed under a microscope;

ISP3 medium, 28°C, cultured for 2 days, the colony is milky white, almost round, opaque, the surface of the colony is slightly depressed, and initially secretes brown pigment, and finally secretes blue-purple pigment; observed under a microscope, the morphology of spore filaments and spores is shown in Figure 2 shown;

ISP4 culture medium, 28°C, cultured for 2 days; the colony is light yellow, round, opaque, the surface of the colony is smooth, and secretes pink pigment; under the microscope, the morphology of sporocystosis and spores is shown in Figure 3;

ISP5 medium, 28°C, cultured for 2 days; the colony is white, round, opaque, the surface of the colony is smooth, and no pigment is secreted; under the microscope, the morphology of sporocystosis and spores is shown in Figure 4;

ISP6 medium, 28°C, cultured for 2 days; the colony is milky white, round, slightly transparent, the surface of the colony protrudes upwards, and secretes black pigment; under the microscope, the morphology of spore filaments and spores is shown in Figure 5;

ISP7 culture medium, 28°C, cultured for 2 days; the colony is light milky white, almost round, transparent, the surface of the colony is slightly raised upwards, and secretes gray-blue-purple pigment; observed under a microscope, the morphology of spore filaments and spores is shown in Figure 6;

The culture characteristics of bacterial strain YP1 are shown in Table 1;

Table 1

The morphological characteristics of the strain YP1 indicated that the strain YP1 had typical characteristics of the genus Streptomyces and was a type of Streptomyces.

In addition, the strain YP1 is Gram-positive by Gram staining, which is a Gram-positive bacterium; under the condition of constant culture medium, the shaker is cultivated at a low speed (less than 100rpm/min) (low dissolved oxygen) strain The growth of YP1 was poor, indicating that the strain YP1 is an aerobic bacteria.

The culture conditions of strain YP1 were optimized. The experimental results showed that the culture conditions of the strain YP1 were: pH5-8, 20-40°C; among them, the optimum pH was 7.0, and the optimum temperature was 28°C. NaCl tolerance 0-5% (w/v).

1.2 Physiological and biochemical characteristics

Detection method: The detection method of the physiological and biochemical characteristics of the strain is carried out according to "Berger's Bacterial Identification Manual" and "Common Bacterial System Identification Manual".

The test results are shown in Table 2.

Explanation of test results: "-" means negative (not detected), "+" means positive (detected), "W" means undetectable.

Table 2

1.3 Gene characteristics

Carry out DNA sequencing to the bacterial strain YP1 of the present application, adopt its 16SrRNA gene sequence to amplify its 16SrRNA gene sequence as shown in SEQ ID NO.2 and SEQ ID NO.3, then carry out sequencing, its 16S rRNA gene sequence is as SEQ ID NO.1 shown. The gene sequence has been added to GenBank/EMBL with accession number MK801227.1. The genome sequence data of the Streptomyces strain YP1 of the present application shows that the G+C content in its DNA is 71.5% (mass content). The dDH values of the Streptomyces bacterial strain YP1 of the present application and the close-type strain Streptomyces zaomyceticus (Streptomyces zaomyceticus, NBRC 13348), the close-type strain Streptomyces lateritius (Streptomyces lateritius, LMG 19282) are respectively 28.5% and 33.9%; Far lower A threshold of 70% of the bacterial species recommended by Mayr-Koltov et al. The ANI values of the Streptomyces bacterial strain YP1 of the application and the near-type strain Streptomyces zaomyceticus (Streptomyces zaomyceticus, NBRC 13348) and the near-type strain Streptomyces erythromyces (Streptomyces lateritius, LMG 19282) are respectively 83.95% and 87.56%, far lower than A threshold of 95-96% of species is suggested for delimitation. The gene sequence of the strain YP1 of the present application was aligned through Genebank, and a phylogenetic tree was established using MEGA6.0. The results are shown in FIG. 7 . Through phylogenetic analysis of 16S rDNA sequence, strain YP1 was identified as Streptomyces vilmorinianum group.

SEQ ID NO.2:27F (5'-AGAGTTTGATCCTGGCTCAG-3');

SEQ ID NO.3:1492R (5'-TACGACTTAACCCCAATCGC-3').

1.4 Chemical characteristics

Cell wall hydrolysis: Put 15mg of freeze-dried cells of strain YP1 into a vial with a screw cap, add 1ml of 1mol/L H2SO4 and mix well, hydrolyze at 100°C for 2h, add saturated barium hydroxide to adjust the pH to 5.2-5.5, spin Evaporate to dryness, add 0.3ml water to redissolve; obtain the cell wall hydrolyzate of YP1 strain.

The cell wall hydrolyzate of the YP1 strain was analyzed and contained diaminopimelic acid (DAP), glucose and ribose. Cell quinones were detected, and the detected quinones were MK-9 (H6) (56%), MK-9 (H8) (38%), MK-8 (H6) (3%), MK-9 (H4) (1%) and MK-9(H2) (1%). Determination of polar lipids and fatty acids, the detected polar lipids are: phosphatidylethanolamine (PE), phosphatidylinositol (PI), phosphatidylinositol mannoside (PIM), an unknown phospholipid; the main Fatty acids (>10%) were iso-c16:0 and C18:0.

Cell Wall Hydrolysates: Cellular respiratory quinones, cellular sugar types, polar lipids, and fatty acids can signal the presence of a certain class of microorganisms. Cytoquinones refer to the common respiratory quinones of aerobic bacteria (menaquinone, ubiquinone). Streptomyces should only be able to detect menaquinones, but not ubiquinone (coenzyme Q). Cell sugars can divide actinomycetes into four categories, as shown in Table 3. Polar lipids can divide actinomycetes into five classes, as shown in Table 4. According to the fatty acid composition of the cells, bacteria can be divided into two large groups. One group contains branched and/or inverse branched chain fatty acids as main constituents. Another group consists of straight chain fatty acids including monounsaturated acids. The major constituents of Gram-positive fatty acids contain only cis-branched or trans-branched fatty acids.

The cell wall hydrolyzate contains DAP, indicating that it is a Gram-positive bacterium, and its cell sugars are glucose and ribose. It can be known that this bacterial strain belongs to type C actinomycetes. Respiratory quinones of aerobic bacteria contain two types of compounds, menaquinone and ubiquinone, among which gram-positive bacteria only contain menaquinone compounds, while gram-negative bacteria and facultative anaerobic bacteria contain ubiquinone Compounds, so the strain belongs to Gram-positive aerobic bacteria. The marked polar lipid determined by this strain is PE, which belongs to PⅡ type actinomycetes.

Table 3 The main glycoforms of the whole cell wall of actinomycetes

Galactose Arabic candy Xylose maduro pond Type A + + - - Type B - - - + Type C - - - - D type - + + -

+ means detected; - means not detected.

Table 4 Polar lipid types of actinomycetes

+ means detected; - means not detected; PE: phosphatidylethanolamine, PC: phosphatidylcholine, PME: phosphatidylmethylethanolamine, PG: phosphatidylglycerol, GluNus: phospholipid containing glucosamine with unknown structure;

The antibacterial performance of embodiment 2 bacterial strain YP1 fermented liquid

Plate culture: Take out the glycerol tube of the preserved strain YP1 from the refrigerator, thaw it naturally at room temperature, use an inoculation loop to streak the glycerol tube of the strain YP1 on the solidified plate solid medium, and cultivate it in a constant temperature incubator at 37°C for 2 days . The plate-activated strain produces a beautiful blue substance on the plate.

Seed liquid culture: Pick the well-grown YP1 bacteria from the plate culture medium that produces blue pigment, insert it into the high-temperature sterilized proliferation medium in the ultra-clean bench, and put the Erlenmeyer flask containing 50ml of the proliferation medium (with a capacity of 250ml) was placed in a constant temperature shaker for 2 days, the rotation speed of the shaker was 180r/min, and the culture temperature was 37°C. Obtain the seed solution of strain YP1.

Fermentation culture: In the ultra-clean bench, the bacterial strain YP1 seed liquid in the middle and late logarithmic growth period is inserted into the high-temperature sterilized fermentation medium with an inoculum size of 5% (v/v), and the 50ml fermentation medium The Erlenmeyer flask was cultured in a constant temperature shaker for 4 days, the speed of the shaker was 180r/min, and the culture temperature was 37°C. Obtain the blue strain YP1 fermentation liquid. The fermentation broth is centrifuged, concentrated by rotary evaporation, and the excess liquid is evaporated to obtain a purple-red concentrated sample.

Adopt the Oxford cup method to measure the in vitro inhibitory effect of the fermented liquid of bacterial strain YP1 to 4 kinds of test bacteria (bacteria concentration is 107CFU/mL), the result shows, bacterial strain YP1 fermented liquid is to Gram-positive bacterium namely Staphylococcus aureus and Bacillus subtilis It has inhibitory effect, but has no inhibitory effect on Gram-negative bacteria, namely Escherichia coli and fungus Candida albicans. The antibacterial results are basically consistent with the antibacterial properties of duracin. It is preliminarily judged that there are duracin and its analogues in the fermentation broth of strain YP1 . The specific results are shown in Table 5:

Table 5 Oxford Cup Determination of the Antibacterial Effect of Streptomyces Strain YP1 Fermented Liquid

strain Antibacterial zone diameter/mm Staphylococcus aureus 18.98±0.18 Bacillus subtilis 18.55±0.24 Escherichia coli - Candida albicans -

Note: "-" means no antibacterial diameter; antibacterial zone diameter>20 is extremely sensitive, 15-20 is highly sensitive, 10-14 is moderately sensitive, <10 is lowly sensitive, and 0 is not sensitive.

The identification of embodiment 3 bacterial strain YP1 fermented liquid products

The purple-red concentrated sample obtained in Example 2 was subjected to the following separation and purification. At the same time, the determination of the molecular weight and the identification of the structure of the isolated and purified pure substance are carried out, and the specific operations are as follows:

(1) Purify and concentrate samples with macroporous adsorption resin, select ethanol-water system for experiment, wash samples with deionized water, 10%, 30%, 47%, 75%, 90%, 100% ethanol gradient, select 90% The sample washed out with 100% high-concentration ethanol is regarded as crude substance A, and the sample washed out with water is selected as crude substance B;

(2) Dissolve the crude substance A in 40% methanol, and filter the upper half to prepare a high-performance liquid chromatograph; select methanol-water system to elute the sample, and use 40%, 60%, 70%, 80%, 96% Gradient flushing with methanol, re-dissolving the sample washed out with 60% methanol in 60% methanol, continuing the first half of preparative high-performance liquid chromatography for separation and purification, and selecting the methanol-water system to elute the sample. Use 60% methanol and high-concentration methanol (94%) to wash the samples respectively, select the samples washed out with 60% methanol and combine them, evaporate to dryness and weigh them as X1; choose the samples washed out with 94% methanol and have a single component The samples were combined, evaporated to dryness and weighed as X2. The purity and molecular weight of X1D were determined by liquid chromatography-mass spectrometer. The NMR sample solvent of pure substances is CD3OD, and the Varian 500MHz spectrometer is used for experiments. The identification results are: X1 is duracin, and X2 is the analogue of duracin: 4,8,13-trihydroxy-6,11-dione-trihydrodicacin A (4,8,13-trihydroxy-6 ,11-dione-trihydrogranaticins A), namely TDTA. (3) The crude substance B is scanned by infrared spectrum, and the infrared spectrum has characteristic absorption peaks at 3271.55cm-1, 1635.99cm-1, and 1589.87cm-1, which have typical characteristic absorption peaks of melanin; it is melanin.

Claims (10)

1. In a Streptomyces strain YP1, its preservation number is CGMCC.23383. 2. Streptomyces bacterial strain YP1 according to claim 1, its 16S rRNA gene sequence is as shown in SEQ ID NO .1. 3. Streptomyces bacterial strain YP1 according to claim 2, the PCR primer of amplification 16S rRNA gene sequence is as shown in SEQ ID NO .2 and SEQ ID NO .3. 4. Streptomyces strain YP1 according to claim 1, 2 or 3, tolerates the NaCl of 0-5% w/v. 5. The application of Streptomyces strain YP1 described in any one of claims 1-4 in producing dunicin. 6. The application according to claim 5, comprising the steps of: (1) Plate culture: Streak the strain YP1 on the plate solid medium, and culture it at a constant temperature of 36-38°C for 1-3 days; (2) Seed liquid culture: Pick the well-grown YP1 bacteria from the plate medium, insert it into the high-temperature sterilized proliferation medium, and put it in a constant temperature shaker for 1-3 days, and the shaker speed is 170 -190r/min, culture temperature is 36-38°C; obtain strain YP1 seed liquid; (3) Fermentation culture: Inoculate the seed liquid of strain YP1 into the high-temperature sterilized fermentation medium with an inoculation amount of 4-6% v/v, and place the fermentation medium in a constant temperature shaker for 3-5 days. The shaker speed is 170-190r/min, and the culture temperature is 36-38°C; the blue strain YP1 fermentation broth is obtained; (4) Centrifuge the fermentation broth of strain YP1, concentrate it by rotary steaming, and purify it to obtain duracin; Plate medium: 20g of agar, 10g of lactose, 5g of tryptone, 10g of yeast extract powder, 4g of NacL and 1000ml of distilled water, pH=7.0, sterilized at 121°C for 20 minutes; Proliferation medium: lactose 10g, tryptone 5g, yeast extract powder 10g, NacL 4g and distilled water 1000ml, pH=7.0, sterilized at 121 ℃ for 20 min; Fermentation medium: lactose 10g, tryptone 5g, yeast extract powder 10g, NacL 4g and distilled water 1000ml, pH=7.0, sterilized at 121°C for 20 min. 7. The application according to claim 6, wherein the culture condition of the step (1) is: culture at a constant temperature of 37° C. for 2 days. 8. The application according to claim 6, the culture condition of the step (2) is: culture in a constant temperature shaker for 2 days, the speed of the shaker is 180r/min, and the culture temperature is 37°C. 9. The application according to claim 6, the culture condition of the step (3) is: culture in a constant temperature shaking table for 4 days, the speed of the shaking table is 180r/min, and the culture temperature is 37°C. 10. The application according to claim 6, 7, 8 or 9, said purification comprising the following operations: (1) Purify the sample obtained by rotary evaporation and concentration with a macroporous adsorption resin, select the ethanol-water system for washing, and collect the samples washed out with 90% and 100% concentration of ethanol as crude substance A; (2) Dissolve the crude material A in 40% methanol, filter the upper half to prepare a high performance liquid chromatograph, select the methanol-water system to elute the sample, and collect the sample washed out with 60% methanol; Said % is a mass percentage.

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