Method for increasing fermentation yield of mernomycin A based on enhanced chassis strain glycosyl donor

Abstract

The invention discloses a method for increasing the fermentation yield of mernomycin A based on a reinforced chassis strain glycosyl donor. Specifically, in the heterologous expression process of a sirnomycin A biosynthetic gene cluster in a chassis strain Streptomyces albus J1074, a glycosyl donor synthesis module gene and a resistance gene (MaPBP) are introduced at the same time, and on the basis of improving a glycosyl biosynthetic donor, the physiological tolerance of the chassis strain is also enhanced. Compared with a high-yield strain Streptomyces albus LX03 (CGMCC NO. 20985) of which the tolerance is only optimized, the yield of the strain Streptomyces albus LX02: pJQK610 subjected to genetic engineering optimization is improved by 285%, and the yield reaches 114 mg / L. The strain disclosed by the invention has the advantages that the yield of the strain Streptomyces albus LX02: pJQK610 is improved by 285%, and the yield of the strain Streptomyces albus LX02: pJQK610 is improved by 285%. The invention provides a high-yield technology for improving heterologous expression of the mernomycin A through collaborative optimization of precursor supply and physiological tolerance, and provides reference for efficient production of the medicine.

Description

Technical Field

[0001] This invention belongs to the field of microbial fermentation technology, and relates to a method for increasing the fermentation yield of monoxin A based on enhancing the glycosyl donor of the chassis strain; it also relates to an optimized host for heterologous expression of the monoxin A biosynthetic gene cluster. S. albus LX02::pJQK610, this strain not only improved the tolerance of the chassis strain to monoxin A, but also improved the glycosyl precursor of monoxin A biosynthesis, thus significantly increasing the yield of monoxin A. Background Technology

[0002] With the continuous emergence of new infectious diseases, the development of highly effective anti-infective drugs has always been an important research focus. Among them, monoxin A belongs to the glycophospholipid family of natural products. Compounds in this family can bind to the active site of bacterial cell wall peptidoglycan transferase (PGT), inhibiting bacterial cell wall synthesis. They exhibit good bactericidal or inhibitory effects against many Gram-positive bacteria, showing different inhibitory effects against different Gram-positive pathogens, with minimum inhibitory concentrations (MICs) ranging from 1 to 100 ng / mL. Importantly, this class of drugs is prone to drug resistance, and many derivatives also possess significant antibacterial activity and certain antitumor activity, making them important candidate compounds for new drug development. Based on the substrate composition of monoxam A's chemical structural unit, it can be decomposed into UDP-GlcUA (obtained by removing one molecule of water from the C-6 hydroxyl group) and the UDP-Glucose pathway module, UDP-GlcNAc and UDP-chinovosamine (obtained by modifying UDP-GlcNAc with MoeR5 and MoeS5) pathway modules, the 3-phosphoglycetate (3PG) module, and the MEP (synthetic farnesyl diphosphate and geranyl diphosphate) pathway module. However, commercially available monoxam is mainly obtained through fermentation of *Streptomyces ghana* ATCC14672, resulting in extremely low yields and complex compositions, posing significant challenges to the quality control and drug development of monoxam products. Summary of the Invention

[0003] The purpose of this invention is to provide a method for increasing the fermentation yield of monoxin A based on enhancing the glycosyl donor in a chassis strain. This invention provides a fermentation method that effectively increases the yield of monoxin A by jointly optimizing the physiological tolerance of monoxin A to a glycosyl synthesis donor and a *Streptomyces cepacia* chassis strain through a combined optimization strategy. By optimizing the gene circuitry for monoxin A biosynthesis, a phosphate glucose-transferred gene of the glycosyl donor synthesis is further incorporated into the site-directed mutant A penicillin-binding protein (MaPBP) gene-optimized vector pJQK557. pgm (phosphoglucomutase, PGM) catalyzes the interconversion between glucose-6-phosphate and glucose-1-phosphate (G1P), UGP ( ugp This is UDP-glucose pyrophosphorylase, which catalyzes the conversion of glucose-1-phosphate (Glc-1-P) to UDP-glucose. glmU This is a dual-domain catalytic enzyme with functions of glucosamine-1-phosphate acetyltransferase and N-acetylglucosamine-1-phosphate uridinetransferase, capable of catalyzing the conversion of N-acetylglucosamine monophosphate (GlcNAc-1-P) to UDP-N-acetylglucosamine (UDP-GlcNAc). The constructed gene circuit was transferred into the chassis strain. S. albus LX02 was obtained in the later stages S. albus LX02::pJQK610, this strain not only improved tolerance to monoxin A, but also increased the production of glycosyl precursors for monoxin A biosynthesis. Fermentation results showed that this strain significantly increased the yield of monoxin A.

[0004] The objective of this invention is achieved through the following technical solutions.

[0005] In a first aspect, the present invention provides a method for increasing the yield of the antibacterial drug monoxin A in a heterologous Streptomyces host, specifically, increasing the heterologous synthesis of monoxin A by Streptomyces albopictus. Specifically, in a Streptomyces albopictus strain heterologously expressing the complete biosynthetic gene cluster of monoxin A with an optimized promoter, glycosyl donor synthesis module genes (UDP-Glu and UDP-GlcNAc) and a resistance gene are introduced. MaPBP Obtain genetically engineered strains ( S. albus LX02::pJQK610); the resulting strain was fermented to obtain monoxin A. Previous findings of this invention indicated that introducing only the glycosyl donor module resulted in insufficient tolerance to monoxin, significantly limited strain growth, and lower yield compared to introducing only the resistance gene. MaPBP The yield of strains such as LX03 is much lower.

[0006] The method of this invention significantly increases the yield of monoxin A biosynthesis. The molecular formula of monoxin A is C2. 69 H 108 N5O 34 P, with the following chemical structural formula: .

[0007] As one embodiment of the present invention, the glycosyl donor synthesis module plasmid is constructed by utilizing the universal plasmid pBluescript II SK(+) and a functional gene for efficient glycosyl product synthesis from the high-yielding strain *Streptomycin Jinggang* JG5008 (GCA_000245355.1), which produces the glycosyl product *Jinggangmycin*, to construct a glycosyl donor module synthesis gene containing the glycosyl product required for the biosynthesis of monoxin A in *E. coli*. pgm (Accession No.CP003275), ugp (Accession No. JF922303.1) and glmU (Accession No. CP003275). In the constructed glycosyl donor synthesis module, the gene... ugp and glmU The chloramphenicol resistance gene is linked between them. cat, The constructed gene cluster module contains both upstream and downstream components. Nde I. Restriction site. The constructed glycosyl donor synthesis module plasmid was named plasmid pJQK598.

[0008] As one embodiment of the present invention, the glycosyl donor module synthesizes genes. pgm , ugp and glmU All were derived from the Jinggang variant of streptomycin JG5008 (GCA_000245355.1).

[0009] As one embodiment of the present invention, the DNA sequence of the glycosyl donor synthesis module gene is shown in SEQ ID NO.1.

[0010] As one embodiment of the present invention, the glycosyl donor synthesis module genes (UDP-Glu and UDP-GlcNAc) and the resistance gene MaPBP The construction of the co-optimized overexpression plasmid for the modules is as follows: the glycosyl donor synthesis module plasmid (pJQK598) is constructed via... Nde After digestion with enzyme I, it is combined with enzymes containing the resistance gene. MaPBP plasmid (pJQK557) Nde The enzyme digestion product was ligated to construct the overexpression plasmid, denoted as plasmid pJQK610.

[0011] The above resistance genes MaPBP With strains S. albus Resistance gene in LX03 (CGMCC NO. 20985) MaPBP The sequence is consistent (see sequence SEQ ID NO. 52 in the inventor's prior patent CN 112410278 A).

[0012] As one embodiment of the present invention, the universal strong promoters used for each gene include: Genes in the glycosyl donor synthesis module pgmA Subject to general strong promoters stnYp Expression control, ugp and glmU Subject to general strong promoters kasOp * controls the expression of; resistance gene MaPBP Subject to general strong promoters kasOp * indicates the control of expression.

[0013] As one embodiment of the present invention, the method includes the following steps: S1. Electroporate the overexpression plasmid (pJQK610) to... E. coli The strain obtained was ET12567 / pUZ8002, denoted as E. coli ET12567 / pUZ8002::pJQK610; S2, Utilizing strains E. coli ET12567 / pUZ8002::pJQK610 is a genera-indirect gene transfer matrix of Streptomyces. The overexpression plasmid was introduced into a strain heterologously expressing the complete biosynthetic gene cluster of monoxin A. Correct conjugates were screened to obtain a strain with optimized glycosyl donor synthesis module genes and resistance genes, denoted as strain [strain name missing]. S. albus LX02::pJQK610.

[0014] As one embodiment of the present invention, strain S. albus LX02::pJQK610 chromosome Ø31 The integration site integrates the complete biosynthetic gene cluster of monoxamycin A and the abramycin resistance gene. ØBT1 The integration site integrates the glycosyl donor synthesis module gene and the resistance gene carried on the overexpression plasmid (pJQK610). MaPBP And hygromycin resistance genes.

[0015] As one embodiment of the present invention, the fermentation culture is carried out by fermenting the genetically engineered strain (a heterologous synthetic strain of monoxin). S. albusAfter the spores of LX02::pJQK610 were activated, they were first inoculated into TSBY medium to prepare seed culture solution, and then inoculated into R5 solid medium. After culturing at 30°C for 7 days, monoxin A was extracted from the fermentation solid plate.

[0016] The method of this invention enhances the efficient biosynthesis of monoxin A in *Streptomyces whiteus* by optimizing the physiological tolerance and glycoside supply pathway of the chassis strain. In some embodiments, the method includes the following steps: S1. Construct a glycosyl donor module gene containing the gene required for the biosynthesis of monoxin A from the streptomycin Jinggang variant JG5008 (GCA_000245355.1) on the universal plasmid pBluescript II SK(+). pgm, ugp and glmU To facilitate cloning, the chloramphenicol resistance gene was linked together while assembling the three genes. cat The constructed gene cluster module contains both upstream and downstream components. Nde After confirming the correctness of the I restriction site through sequencing, plasmid pJQK598 was obtained.

[0017] S2, In genes containing monoxamine A tolerance kasOp *- Salb-PBP2 Based on plasmid pJQK557 (P238N&F200D), through Nde The I restriction site was ligated into a glycosyl donor gene module derived from plasmid pJQK598 to obtain plasmid pJQK610.

[0018] S3, Electroplasty of plasmid pJQK610 to... E. coli Among the strains ET12567 / pUZ8002, this strain is a universal strain of Escherichia coli-Streptomyces biparental conjugation transfer.

[0019] S4. Plasmid pJQK610 was introduced into a gene cluster containing the complete biosynthesis of monoxin A via an E. coli-Streptomyces biparental conjugation transfer strategy. S. albus After verification in LX02, a strain capable of efficiently synthesizing monoxin A was obtained. S. albus LX02::pJQK610.

[0020] S5, fermentation culture LX03 and strain S. albus After fermentation of two mutant strains, LX02::pJQK610, 1 mL of fermentation broth was taken from each strain and an equal amount of methanol was added. The mixture was then sonicated for 30 min in an ultrasonic bath. The two fermentation extracts were prepared by performing the same operation in parallel.

[0021] S6. The yield of monoxin A in the two strains was quantitatively analyzed using LC-QQQ-MS. The results showed that the heterologous expression yield of monoxin A in strain LX02::pJQK610 was 2.85 times higher than that in the original strain LX03. Secondly, the present invention also provides a genetically engineered strain of Streptomyces whitei prepared by the aforementioned method.

[0022] Thirdly, the present invention also provides a strain of white Streptomyces. S. albus LX02::pJQK610, with accession number CGMCC No.35966.

[0023] The white Streptomyces of the present invention Streptomycess albus LX02::pJQK610 was deposited on September 18, 2025, at the China General Microbiological Culture Collection Center (No. 3, Courtyard 1, Beichen West Road, Chaoyang District, Beijing, Institute of Microbiology, Chinese Academy of Sciences), with accession number CGMCC No. 35966.

[0024] Streptomyces white in this invention Streptomycess albus LX03 was deposited on November 2, 2020, at the China General Microbiological Culture Collection Center (No. 3, Courtyard 1, Beichen West Road, Chaoyang District, Beijing, Institute of Microbiology, Chinese Academy of Sciences), with accession number CGMCC No. 20985.

[0025] Compared with existing monoxin A production technology, the present invention has the following advantages: This invention utilizes a strategy that combines optimization of the substrate supply pathway in the biosynthesis of monoxin A with enhancement of the tolerance of chassis strains. The constructed gene pathway is integrated into the chromosome of *Streptomyces albopictus* chassis strains, resulting in optimized heterologous expression strains. S. albus LX02::pJQK610, this strain not only contains a highly efficient glycosyl donor in the biosynthesis of monoxin A, but also a resistance gene. MaPBP Overexpression of monoxin A further enhanced the host's tolerance to monoxin A, and the constructed heterologous expression strain S. albus The yield of monoxin A in LX02::pJQK610 was 2.85 times higher than that of its originating strain LX02, which laid a good foundation for the creation of efficient biosynthesis of monoxin A. Attached Figure Description

[0026] Other features, objects, and advantages of the invention will become more apparent from the following detailed description of non-limiting embodiments with reference to the accompanying drawings: Figure 1 A schematic diagram illustrating the construction of the glycosyl module donor integrative plasmid pJQK610; Figure 2 Establish the standard curve for the quantitative analysis of meroxamycin A; Figure 3 strain S. albus LX03 and S. albus Comparison results of fermentation yield of LX02::pJQK610. Detailed Implementation

[0027] The following examples, in conjunction with the accompanying drawings, further illustrate the present invention. These embodiments are implemented based on the technical solution of the present invention and provide detailed implementation methods and processes. These embodiments will help those skilled in the art to further understand the present invention, but the scope of protection of the present invention is not limited to the following embodiments. For those skilled in the art, appropriate adjustments and improvements can be made without departing from the spirit of the present invention, and these all fall within the scope of protection of the present invention. Experimental methods in the following embodiments that do not specify specific conditions are performed under conventional conditions or the manufacturer's recommended conditions.

[0028] Example Constructing high-yield strains of monoxin A containing gene pathways S. albus Construction of LX02::pJQK610 Step 1: Identification and overexpression of candidate genes for the glycosyl donor gene module pgmA and ugp Gene Fragment Construction: Since UDP-N-acetylglucosamine and UDP-glucose are two important glycosyl donors required in the biosynthesis of monoxin A, insufficient glycosyl donors lead to low synthesis efficiency. This invention's research found that in the primary metabolism of *Streptomyces*, the protein GlmU is a dual-domain catalytic enzyme with functions of glucosamine-1-phosphate acetyltransferase and N-acetylglucosamine-1-phosphate uridine transferase, catalyzing the conversion of N-acetylglucosamine 1-phosphate (GlcNAc-1-P) to UDP-GlcNAc. UGP is a UDP-glucose pyrophosphorylase, catalyzing the conversion of glucose 1-phosphate (Glc-1-P) to UDP-Glc. *Streptomyces hygroscopicus* var. *jinggangmycin* JG5008 can efficiently synthesize jinggangmycin, and the functional genes of the glycosyl donors have strong catalytic abilities. Therefore, relevant genes from this strain were selected for assembling the glycosyl module gene circuit.

[0029] Using the genome of *Streptomyces hygroscopicus* var. *JG5008* (GCA_000245355.1) as a template, clones were performed... pgmA Genes (PgmA-F / R) and ugp (upg-F / R) gene, using plasmid pDR3- containing a strong promoter stnYp and pDR3- kasOp* (Microbial Cell Factories Using 2023, 22, 127 as templates, the promoter was cloned using primers. stnYp (StnYp-F / R) and KasOp * (Kas-upg-F / R), and then use overlap PCR to... ​ and ​ (PgmA-R / StnYp-F), ​ and ​ * Assembly is performed using primers (Kas-upg-F / upg-R). Finally, the two fragments are assembled again using primers PgmA-F and upg-R to obtain the final product. ​ Genes and ​ The gene fragment linked by the gene, the DNA fragment length is 2.6kb, in ​ Downstream with ​ The I restriction site and 20nt homologous arm are used for assembly with the pBlueScriptⅡSK(+) plasmid.

[0030]

[0031] Step Two: Overexpression ​ Construction of the chloramphenicol-coupled gene fragment: Using the genome of Streptomyces hygroscopicus var. Jinggangense JG5008 as a template, the fragments were cloned separately. ​ Gene (glmU-F / R), ​ The (Kas-glmU-F / R) promoter and chloramphenicol resistance gene (Cat-F / R) were used to cross-link the resistance selection marker gene with the resistance selection marker gene via overlap PCR (Cat-F / glmU-R). ​ The integrated gene fragment is approximately 2.3 kb in size. ​ Downstream with ​ The I restriction site and 20 nt homologous arm were used for assembly with the pBlueScriptⅡSK(+) plasmid. Finally, ​ - ​ With cat- ​ Two gene fragments and ​ The pBlueScriptⅡSK(+) plasmid digested by RV was assembled and ligated into a three-point segment, and the plasmid pJQK598 was obtained after sequencing verification.

[0032]

[0033] Step 3: Overexpression of genes supplying complete glycosyl groups ( ​ , ​ Construction of the chloramphenicol-coupled fragment module: The plasmid pJQK598 obtained in step two was used... ​The fragment containing the monoxamine A resistance gene was digested with enzyme I and recovered (approximately 5 kb). ​ and φBT1 ​ The integrative plasmid pJQK557 (its construction can be found in CN112410278A) also utilizes ​ I single enzyme digestion was performed, and the vector fragment of approximately 9.2 kb was recovered. The two were then ligated to obtain the correct plasmid, named pJQK610. ​ ).

[0034] Step 4: The plasmid pJQK610 obtained in Step 3 was introduced into the recipient strain LX02 via indirect co-transfer between Escherichia coli and Streptomyces LX02.

[0035] The plasmid pJQK610 successfully constructed in step three was converted to [a specific plasmid] via electroporation. ​ In ET12567 / pUZ8002, select the correct clones and place them in 3 mL of liquid LB medium containing 1 / 1000 volume of chloramphenicol (25 mg / mL), 3 μL of kanamycin (50 mg / mL), and hygromycin (50 mg / mL). Incubate overnight at 37°C with a shaker at 220 rpm. Then, transfer 10% of the overnight culture to 5 mL of fresh LB medium (containing 1 / 1000 volume of chloramphenicol (25 mg / mL), 3 μL of kanamycin (50 mg / mL), and hygromycin (50 mg / mL)) and incubate for 3 hours. Then, wash the bacterial cells twice with 5 mL of fresh LB solution to thoroughly remove the antibiotics from the culture. Collect 3 mL of fresh spore solution (containing approximately 1 × 10⁻⁶ spores) of the recipient strain LX02 (see CN112410278A). 8 The spores were washed twice with TES solution and then suspended in 0.5 mL of TES solution. They were then heat-shocked in a 50°C water bath for 10 min, quickly removed, and cooled to room temperature under tap water. An equal volume of 2× spore pre-germination culture medium and 0.5 μM calcium chloride solution were added, and the mixture was incubated at 37°C in a shaker for 2.5 hours to pre-germinate the spores, preparing for conjugation transfer. The spores were collected by centrifugation at 5000 rpm / min, washed once with LB medium, and then resuspended in 1 mL of LB medium. The spores were thoroughly shaken and dispersed on a mixer. They were then mixed with a donor-acceptor medium containing plasmid pJQK610 at a 1:10 donor-acceptor ratio. ​After mixing ET12567 / pUZ8002 cells, the mixture was evenly spread onto antibiotic-free MS plates. After the plates were dried, they were transferred to a 30°C incubator and incubated for approximately 16 hours. The plates were then removed and covered with 1.5 mL of sterile water containing hygromycin (1 mg) and nalidixic acid (1 mg). The plates were dried again and transferred to a 30°C incubator. Single colony conjugates were observed to grow on the plates after 5-6 days. Finally, the cells were... ​ -F / R and hyg-F / R were used as PCR primers to verify the correctness of the binding sites, and to obtain the heterologous synthetic strain of monoxin A. ​ LX02:: pJQK610.

[0036]

[0037] Step 5: Treatment of the strain ​ LX03 (see CN112410278A) and ​ A comparison of the fermentation yields of two strains, LX02:: pJQK610 and monoxin A, was conducted. This fermentation process involves two steps: activation with seed culture medium and solid-state fermentation on R5 medium. The primary seed culture medium is TSBY, which consists of 30 g of Oxoid tryptophan soup powder, 5 g of Difco yeast extract, and 103 g of sucrose, diluted to 1 L with distilled water, and has a pH of 7.2. It is then autoclaved at 115°C for 25 min. The fermentation medium R5 consists of 60 g / L sucrose, 10 g / L glucose, 5 g / L yeast extract, 0.25 g / L K₂SO₄, 2 g / L MgCl₂, 0.1 g / L casein hydrolysate, 2 mL / L trace elements, 5.73 g / L TES buffer, and 15 g / L agar powder, diluted to 1 L with distilled water, and has a pH of 7.2. CoCl2 1 mg / L (added after sterilization), KH2PO4 (0.5%) 10 mL / L (added after sterilization), CaCl2 (5M) 4 mL / L (added after sterilization), L-proline (20%) 15 mL / L (added after sterilization). The autoclaved R5 solid fermentation medium was poured into 9cm diameter petri dishes in a laminar flow hood, approximately 30 mL per dish. The bacterial strain was then added... ​ LX03 and ​ LX02::pJQK610 were transferred to seed culture medium and cultured in a shaker at 220 rpm and 30°C for 24 h to obtain fermentation seed liquid. 200 μL of the fermentation seed liquid was inoculated into each culture dish, spread evenly, and then placed in a constant temperature incubator at 30°C for 7 days. The solid fermentation culture was then collected, and the production of monoxin A was analyzed according to the following method.

[0038] The solid-state fermentation product was cut into small pieces and placed in 250 mL Erlenmeyer flasks. An equal volume (30 mL) of analytical grade methanol solution was added to each flask. After thorough mixing, ultrasonic extraction was performed for 15 min to obtain the fermentation extract of the strain. Then, 1 mL of the fermentation extract was added to a 1.5 mL centrifuge tube and centrifuged at 12000 rpm for 10 min. The supernatant was then filtered through a 0.22 μm microporous membrane and placed into a liner tube for LC-QQQ-ESI-MS analysis. A standard curve for meroxam A was constructed using the MRM mode of mass spectrometry. ​ Then, the yield of monoxin A was calculated based on the regression equation. The results showed that the strain... ​ The yield of monoxin A in LX02::pJQK610 reached 114.5 ± 8 mg / L, compared to... ​ The heterologous expression yield of LX03 (39.8 ± 6 mg / L) increased by approximately 2.85-fold. ​ ).

[0039] In summary, this invention provides a high-yield technology for heterologous expression of monoxin A through synergistic optimization of precursor supply and physiological tolerance, offering a reference for the efficient production of this drug. Specifically, it involves the optimization of the monoxin A biosynthetic gene cluster in chassis strains. ​ During the heterologous expression of J1074, a glycosyl donor synthesis module gene and a resistance gene (Mutant class Apenicillin-binding proteins P238N & F200D point mutation genes, MaPBP) were simultaneously introduced. This not only enhanced the availability of glycosyl biosynthetic donors but also improved the physiological tolerance of the chassis strain. The strain was optimized through genetic engineering. ​ LX02::pJQK610, compared to high-yield strains that only optimize tolerance. ​ The yield of LX03 (CGMCC NO. 20985) increased by 285%, reaching 114 mg / L.

[0040] The foregoing has described in detail the specific embodiments of the present invention. It should be further noted that the present invention is not limited to the specific embodiments described above. Those skilled in the art can make various adjustments or modifications within the scope of the claims, and such adjustments or modifications do not affect the substantive content of the present invention.

Claims

1. A method for improving the heterologous synthesis of monoxin A by *Streptomyces albopictus*, characterized in that, In a *Streptomyces* strain with optimized promoter and heterologous expression of the complete biosynthetic gene cluster of monoxin A, a glycosyl donor synthesis module gene and a resistance gene were introduced. MaPBP Genetically engineered strains were obtained; the resulting strains were fermented to obtain monoxin A.

2. The method according to claim 1, characterized in that, The glycosyl donor synthesis module plasmid was constructed in *E. coli* using the universal plasmid pBluescript II SK(+), containing a glycosyl donor module synthesis gene required for the biosynthesis of monoxin A. pgm , ugp and glmU .

3. The method according to claim 1, characterized in that, The DNA sequence of the glycosyl donor synthesis module gene is shown in SEQ ID NO.

1.

4. The method according to claim 1, 2 or 3, characterized in that, Glycosyl donor synthesis module gene and resistance gene MaPBP The construction of the co-optimized overexpression plasmid for the modules is as follows: the glycosyl donor synthesis module plasmid is constructed by... Nde After digestion with enzyme I, it is combined with enzymes containing the resistance gene. MaPBP plasmid Nde The enzyme digestion product was ligated to construct the overexpression plasmid.

5. The method according to claim 4, characterized in that, The universal strong promoters used for each gene include: Genes in the glycosyl donor synthesis module pgmA Subject to general strong promoters stnYp Expression control, ugp and glmU Subject to general strong promoters kasOp * controls the expression of; resistance gene MaPBP Subject to general strong promoters kasOp * indicates the control of expression.

6. The method according to claim 4, characterized in that, The method includes the following steps: S1. Electroporate the overexpression plasmid to... E. coli The strain obtained was ET12567 / pUZ8002, denoted as E. coli ET12567 / pUZ8002::pJQK610; S2, Utilizing strains E. coli ET12567 / pUZ8002::pJQK610 is a genera-indirect gene transfer matrix of Streptomyces. The overexpression plasmid was introduced into a strain heterologously expressing the complete biosynthetic gene cluster of monoxin A. Correct conjugates were screened to obtain a strain with optimized glycosyl donor synthesis module genes and resistance genes, denoted as strain [strain name missing]. S. albus LX02::pJQK610.

7. The method according to claim 6, characterized in that, strains S. albus LX02::pJQK610 chromosome Ø 31 The integration site integrates the complete biosynthetic gene cluster of monoxamycin A and the abramycin resistance gene. ØBT1 The integration site integrates the glycosyl donor synthesis module gene and the resistance gene carried on the overexpression plasmid. MaPBP And hygromycin resistance genes.

8. The method according to claim 4, characterized in that, The fermentation culture involves activating the spores of the genetically engineered strain, first inoculating them into TSBY medium to prepare a seed culture solution, then inoculating them into R5 solid medium, culturing at 30°C for 7 days, and then extracting monoxin A from the fermentation solid plate.

9. A genetically engineered strain of Streptomyces albopictus prepared by the method according to any one of claims 1-8.

10. A white Streptomyces strain Streptomycess albus LX02::pJQK610, with accession number CGMCCNo.35966.

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