Construction method of engineering bacteria for high yield of undecyl prodigiosin

Abstract

The invention relates to a construction method of engineering bacteria for high-yield undecyl prodigiosin, and belongs to the technical field of biology. According to the invention, 16 isolated histidine kinase coding genes with unknown functions in streptomyces coelicolor are inhibited by using a dCas9 mediated CRISPRi technology, strains with significant phenotypic changes are screened out, an OhkB gene is knocked out by using a CRISPR-Cas9 technology, and it is detected that the yield of undecyl prodigiosin is increased, so that the yield of undecyl prodigiosin is increased. And further determining that the OhkB gene can negatively regulate and control the biosynthesis of undecylprodigiosin and actinopurpurin through a refilling experiment. The invention provides a novel regulatable gene locus for increasing the yield of the undecylprodigiosin and the actinopurpurin, and lays a foundation for further development and utilization of the undecylprodigiosin and the actinopurpurin.

Description

Technical Field

[0001] The invention relates to a method for constructing an engineering bacterium with high yield of undecyl prodigiosin, belonging to the field of biotechnology. Background Art

[0002] Streptomyces is a branched aerobic Gram-positive bacterium belonging to the family Streptomycetes of the order Deficientales in the kingdom Prokaryotes. It is one of the main microbial groups in the soil. The average size of the Streptomyces genome is about 8Mb, which is about twice that of the Escherichia coli genome. Its DNA has an extremely high G+C% content (73-75% on average), making it one of the biological species with the highest G+C% content known in nature to date. Streptomyces and its related species have a relatively complex life cycle and can produce a wealth of valuable secondary metabolites along with the life cycle, so they are widely used in industrial pharmaceutical production and basic research. As far as we know, about two-thirds of the natural antibiotics used in medicine and a total of more than 9,000 biologically active substances are produced by Streptomyces, which play an important role in anti-tumor, anti-fungal, anti-parasitic, and immunosuppression.

[0003] As a model strain for Streptomyces research, Streptomyces coelicolor has advantages such as clean genetic background and mature operating system, so it has extremely high research and application value. Streptomyces coelicolor can produce a variety of secondary metabolites, including the iconic actinorhodin (ACT), undecylprodigiosin (RED), calcium-dependent antibiotics (CDA) and methamicin, among which actinorhodin and undecylprodigiosin have characteristic colors, which greatly facilitates the regulation of antibiotic synthesis in Streptomyces coelicolor.

[0004] Undecylprodigiosin (UP) is a fat-soluble antibiotic with three pyrrole rings and 11 alkyl groups in the side chain. 25 H 25 N3O. Undecyl prodigiosin is obtained by the condensation of the intermediate 2-undecylpyrrole (2-Undecylpyrrole) and 4-methoxy-2,2'-bipyrrole-5-carbaldehyde (4-Methoxy-2,2'-bipyrrole-5-carbaldehyde, MBC) under the catalysis of the enzyme RedH.

[0005] Undecyl prodigiosin has many important functions. On the one hand, it has antibiotic activity, which can destroy the cell membrane structure of bacteria and cause the leakage of intracellular substances, thereby inhibiting the growth and reproduction of other microorganisms, helping Streptomyces coelicolor to gain competitive advantages in the living environment and resist the invasion of other microorganisms. At the same time, it can participate in cell signal transduction and regulate the physiological functions of cells, such as cell differentiation and spore formation. In addition, it can also regulate the balance between primary metabolism and secondary metabolism, so that cells can better adapt to environmental changes and meet their own growth and development needs. Stankovic et al. reported that undecyl prodigiosin exhibited high antibacterial properties against Micrococcus luteus ATCC 379, Candida albicans ATCC 10231 and C.albicansATCC 10259; Liu et al. found that undecyl prodigiosin could inhibit the proliferation of P388 cancer cells and induce apoptosis of the cells; Ho et al. found that undecyl prodigiosin could selectively induce apoptosis of human breast cancer cells without obvious toxicity to normal cells; in addition, Taverna reported that undecyl prodigiosin has the activity of immunosuppressants and can be developed as a potential high-efficiency immunosuppressant.

[0006] Based on the antibacterial, antitumor and immunosuppressive properties of undecyl prodigiosin, it has great application prospects in the pharmaceutical industry. However, antibiotic genes in Streptomyces are secondary metabolite genes, and most of them are in a silent state. Due to the codon swing effect, some secondary metabolite genes can be expressed, but the yield is generally not high. Therefore, how to quickly and efficiently increase the yield of antibiotics in Streptomyces is crucial for the subsequent use of antibiotics. Summary of the invention

[0007] The purpose of the present invention is to provide a method for constructing an engineering bacterium with high yield of undecyl prodigiosin and to provide a regulatory gene involved in antibiotic synthesis in Streptomyces coelicolor.

[0008] In order to achieve the above object, the technical scheme of the method for constructing an engineering bacterium with high yield of undecyl prodigiosin in the present invention is:

[0009] A method for constructing an engineered bacterium capable of producing high-yield undecyl prodigiosin, wherein the nucleotide sequence of the OhkB gene is shown in SEQ ID NO.1; and the antibiotic is undecyl prodigiosin and / or actinomycin.

[0010] The beneficial effect of the above technical solution is that the construction method of an engineered bacterium with high yield of undecyl prodigiosin of the present invention is a pioneering invention. The present invention first uses dCas9-mediated CRISPRi technology to inhibit the coding genes of 16 isolated histidine kinases of unknown functions in Streptomyces coelicolor (M145), and finds that after the gene with a nucleotide sequence as shown in SEQ ID NO.1 is inhibited, its phenotype changes significantly (i.e., the expression levels of undecyl prodigiosin and actinomycin are significantly increased), and the gene is named OhkB gene. In order to further clarify the application of OhkB gene in the synthesis of antibiotics of Streptomyces coelicolor, CRISPR-Cas9 technology is used to knock out the OhkB gene (named ΔohkB) in Streptomyces coelicolor M145. It is found through detection that the expression levels of undecyl prodigiosin and actinomycin in Streptomyces coelicolor M145 with OhkB gene knocked out are significantly increased compared with the control group (M145). At the same time, an overexpression vector for overexpressing the OhkB gene was constructed and transfected into M145 and ΔohkB for complementation experiments, respectively, and it was determined that the OhkB gene could negatively regulate the biosynthesis of undecyl prodigiosin and actinomycin in Streptomyces coelicolor. The present invention provides a new regulatory site for the biosynthesis of undecyl prodigiosin and actinomycin in Streptomyces coelicolor, further clarifies the gene network of antibiotic synthesis, and lays a foundation for increasing the yield of undecyl prodigiosin and actinomycin in Streptomyces coelicolor.

[0011] Specifically, in Streptomyces coelicolor, its genome can encode 84 histidine kinase proteins and 80 response regulatory proteins, forming 67 pairs of typical two-component systems, 17 isolated histidine kinases and 13 isolated response regulatory proteins. The coding genes of the histidine kinases and response regulatory proteins of the typical two-component system are adjacent to each other on the genome, and there is a lack of paired response regulatory proteins near the coding genes of the isolated histidine kinases. Among them, only two isolated histidine kinases, OhkA and PdtaS-c, have their functions identified among the 17 isolated histidine kinases, while only one isolated histidine kinase, PdtaS-p, has its function identified in Streptomyces virulens.

[0012] As a further improvement, the following steps are included: constructing a CRISPR / Cas9 vector of the OhkB gene, transforming it into Streptomyces coelicolor, and knocking out the OhkB gene.

[0013] As a further improvement, the CRISPR / Cas9 vector of the OhkB gene is constructed by connecting the obtained sgRNA fragment containing the OhkB gene knockout gene with the linearized starting vector to obtain the CRISPR / Cas9 vector of the OhkB gene.

[0014] As a further improvement, the nucleotide sequence of the sgRNA of the OhkB gene is shown in SEQ ID NO.3.

[0015] As a further improvement, the starting vector is pSET-dCas9. BRIEF DESCRIPTION OF THE DRAWINGS

[0016] Figure 1 The electrophoresis diagram in Example 1 of the present invention (where A is the electrophoresis diagram of the PCR product, B is the electrophoresis diagram of the sgRNA, C is the electrophoresis diagram of the linearized vector, and M is the DNA Marker)

[0017] Figure 2 The peak diagram of sequencing of 16 inhibitory plasmids in Example 1 of the present invention;

[0018] Figure 3 The phenotypic observation of 16 inhibitory strains in Example 1 of the present invention is shown;

[0019] Figure 4 The ohkB sgRNA, homology arm fragment, linearized vector electrophoresis diagram and knockout plasmid pKCcas9-ohkB verification diagram in Example 2 of the present invention (wherein A is the homology arm fragment electrophoresis diagram, B is the connection fragment, C is the linearized vector electrophoresis diagram, and D is the knockout plasmid pKCcas9-ohkB verification diagram)

[0020] Figure 5 The construction of the ΔohkB deletion mutant in Example 2 of the present invention (wherein A is a knockout pattern diagram of the OhkB gene, and B is a double exchange identification of the ΔohkB mutant);

[0021] Figure 6 This is the phenotypic observation of M145 and ΔohkB in Example 3 of the present invention;

[0022] Figure 7 The effect of the deletion of the ohkB gene in Example 3 of the present invention on the synthesis of undecylprodigiosin and actinomycin (wherein, * represents P<0.05, ** represents P<0.01, *** represents P<0.001);

[0023] Figure 8 The verification results of the complemented plasmid pIB139-ohkB in Example 4 of the present invention (where A is the enzyme cutting verification graph and B is the sequencing peak graph);

[0024] Fig. 9 The effects of complementation and overexpression of the ohkB gene on the strain phenotype in Example 4 of the present invention;

[0025] Fig.10The effect of complementing the ohkB gene in Example 5 of the present invention on undecylprodigiosin and actinomycin (wherein ** represents P<0.01, *** represents P<0.001, and NS represents no statistical difference (P>0.5)). DETAILED DESCRIPTION

[0026] The prior art found that microorganisms produce pigments mainly to resist unfavorable growth environments and improve survival ability. Streptomyces coelicolor has a strong adaptability and can survive in extreme environments such as the North and South Poles, which is related to the actinomycin and undecyl prodigiosin produced by it. Undecyl prodigiosin has a variety of important biological activities such as antibacterial, antitumor and immunosuppression, and has great application prospects in the pharmaceutical industry, but currently has a problem of low yield. Based on this, the present invention provides a method for constructing an engineered bacterium with high yield of undecyl prodigiosin.

[0027] The present invention is further described below in conjunction with specific embodiments, but the protection scope of the present invention is not limited thereto; however, these embodiments are only examples and do not constitute any limitation on the scope of the present invention. The details and forms of the present invention may be modified or replaced without departing from the spirit and scope of the present invention, but these modifications and replacements fall within the protection scope of the present invention. The test methods in the following embodiments are conventional methods unless otherwise specified. The test materials used in the following embodiments are purchased from conventional biochemical reagent manufacturers unless otherwise specified.

[0028] The components and preparation process of the culture medium used in the following embodiments of the present invention are as follows:

[0029] MS medium: 5g mannitol, 5g soybean cake powder, 5g agar powder, 250mL distilled water. Sterilize at 121℃ for 20min after mixing.

[0030] MM solid culture medium: L-asparagine 0.1 g, K2HPO4 0.05 g, MgSO4·7H2O 0.02 g, FeSO4·7H2O0.001 g, D-mannitol 1 g, agar powder 1 g, distilled water 100 mL, adjust pH to 7.10, sterilize at 115°C for 30 min.

[0031] Note: MM medium is based on MM solid medium without agar powder.

[0032] The primer sequences used in the following examples of the present invention are shown in Table 1.

[0033] Table 1 Specific nucleotide sequences of primers

[0034]

[0035]

[0036] A specific embodiment of the method for constructing an engineering bacterium with high yield of undecyl prodigiosin of the present invention:

[0037] The present invention firstly uses dCas9-mediated CRISPRi technology to inhibit the coding genes of 16 isolated histidine kinases of unknown functions in Streptomyces coelicolor, and screens out strains with significant phenotypic changes; then, by constructing a ΔohkB deletion mutant, the growth conditions that are conducive to the synthesis of undecyl prodigiosin and actinomycin are explored, strains with phenotypic changes are screened out, and complementation experiments are performed for verification, and the changes in the production of undecyl prodigiosin and actinomycin are measured. The specific implementation operations are as follows:

[0038] Example 1 Screening of genes encoding isolated histidine kinases

[0039] In this example, dCas9-mediated CRISPRi technology was used to inhibit the coding genes of 16 isolated histidine kinases of unknown functions in Streptomyces coelicolor to screen out strains with significant phenotypic changes. The specific implementation operations are as follows:

[0040] CRISPRi plasmids were constructed to inhibit 16 isolated histidine kinase genes, namely SCO1217, SCO1220, SCO1654, SCO2152, SCO2359, SCO3119, SCO3134, SCO3144, SCO4009, SCO4120, SCO5006, SCO5454, SCO6163, SCO6364, SCO7009, and SCO7230. Taking the construction of SCO3119 inhibition plasmid as an example, the plasmid pSET-dCas9 was used as a template, and the primers gRNA-SCO3119-S1 and pSET-dCas9-R were used for the first round of PCR reaction. The reaction system is shown in Table 2, and the reaction procedure is shown in Table 3. The size of the amplified product SCO3119-F1 is 202bp, and the product size is in line with expectations (such as Figure 1 (shown in A).

[0041] Table 2 PCR reaction system

[0042]

[0043]

[0044] Table 3 PCR reaction program

[0045]

[0046] The second round of PCR reaction was performed using the first round PCR product as a template and primers gRNA-Gibson-R and SCO3119-Gib-F (the system and conditions of the second round of PCR were the same as those of the first round, except that the template was replaced with the first round PCR reaction product and the primers were replaced with gRNA-Gibson-R and SCO3119-Gib-F). The sgRNA targeting SCO3119 was amplified to a size of 242 bp. The product size was in line with expectations (e.g. Figure 1 The gel was recovered and purified for later use.

[0047] Using pSET-dCas9 as the starting plasmid, the starting plasmid pSET-dCas9 was double-digested with restriction endonucleases EcoRI and BcuI (using FastDigest restriction endonucleases from Thermo Fisher Scientific, the system and reaction conditions refer to the FastDigest enzyme manual) to linearize the vector. The size of the digestion product was in line with expectations (such as Figure 1 (as shown in C).

[0048] The recombinant product was transformed into E. coli Top10 competent cells to obtain the recombinant plasmid pSET-dCas9-SCO3119, which was sent to Shanghai Sangon Biotech Co., Ltd. for sequencing (such as Figure 2 shown).

[0049] Taking the construction of the inhibitory strain M145 / pSET-dCas9-SCO3119 as an example, the correctly sequenced integration plasmid pSET-dCas9-SCO3119 was transformed into ET12567 / pUZ8002 competent cells. Through conjugation transfer, the plasmid DNA enters the recipient bacterial cells and integrates into the Streptomyces genome. Culture at 30°C for 4-5 days. Use a toothpick to pick up the grown conjugates and draw a 1cm 2 The cells were collected from the small cubes and cultured at 30°C for 4-5 days. The bacteria were streaked densely with a toothpick and cultured at 30°C. After 4-5 days, the spores were collected and stored in a -80°C refrigerator to obtain the inhibitory strain M145 / pSET-dCas9-SCO3119.

[0050] The spores of 16 inhibitory strains were taken out from the cryopreserved tubes, pre-germinated and adjusted to OD 450 =1.0, take 6 μL and drop it on MM medium, draw a line with a toothpick, and culture at 30°C for 4-5 days. Using M145 / pSET-dCas as a control, observe the changes in growth, morphological differentiation and antibiotic production of each inhibitory strain, and screen the inhibitory strain experiment with significant phenotypic changes.

[0051] The results are as follows Figure 3 As shown, compared with the control strain M145 / pSET-dCas9, inhibition of SCO3119 significantly increased RED production, suggesting that SCO3119 negatively regulates the synthesis of ACT and RED in Streptomyces coelicolor. In subsequent experiments, the isolated histidine kinase SCO3119 was named OhkB.

[0052] Example 2 Construction of ΔohkB deletion mutant

[0053] This example uses the gene with significant phenotypic changes screened out in Example 1 as the research object, and uses CRISPR-Cas9 technology to knock it out to construct a ΔohkB deletion mutant. The specific implementation operations are as follows:

[0054] Using pKCcas9dO as a template, primers gRNA-SCO3119-F / R were used to PCR amplify (the reaction system and reaction conditions are as described in Example 1) the sgRNA targeting ohkB, with a size of 117 bp. Using M145 spores as a template, primers SCO3119-up-F / R and SCO3119-down-F / R were used to PCR amplify (the reaction system and reaction conditions are as described in Example 1) the upstream and downstream homology arm fragments of ohkB, with sizes of 1088 bp and 1214 bp, respectively.

[0055] The results are shown in the figure Figure 4 As shown in A and B, the size of the PCR product is consistent with the expectation. The recovered and purified sgRNA and upstream and downstream homology arm fragments were connected by Overlap PCR, and after double digestion with HindIII and BcuI, they were cleaned and recovered for use.

[0056] The starting plasmid pKCcas9dO was double-digested with restriction endonucleases HindIII and BcuI to linearize the vector. The size of the digestion product was in line with expectations (e.g. Figure 4 The vector is recovered and purified for later use.

[0057] The ohkB sgRNA and upstream and downstream homology arm fragments were connected to the linearized pKCcas8dO through the sticky ends, and the recombinant plasmids with correct enzyme digestion were sent to Shanghai Sangon Biotech Co., Ltd. for sequencing. If the sequencing was correct, the knockout plasmid pKCcas9-ohkB (such as Figure 4 (shown in D).

[0058] The knockout plasmid pKCcas9-ohkB with correct sequencing was transformed into ET12567 / pUZ8002 competent cells, and the knockout plasmid pKCcas9-ohkB was introduced into the cells of Streptomyces coelicolor M145 by conjugation transfer, and the CRISPR / Cas9 gene editing technology was used to construct the in-frame deletion mutant of ohkB. The conjugates were picked and cultured on MS medium (containing Apra) and cultured at 30°C for 3-5 days. After the colonies grew, single colonies were selected and subcultured three times on MS medium to loop out the plasmid. The third-generation colonies were identified for resistance, and single colonies that only grew on MS medium without antibiotics but could not grow on MS medium containing Apra were selected. PCR identification was performed using primers sco3119-in-ID-F / R to obtain the ΔohkB mutant strain without plasmid. The results are shown as follows. Figure 5 shown.

[0059] Example 3 Exploration of growth conditions and phenotypic observation

[0060] In this example, the ΔohkB deletion mutant constructed in Example 2 is used as a strain for producing undecyl prodigiosin and actinomycin, and the conditions for producing antibiotics are explored. The specific implementation operations are as follows:

[0061] Nitrogen source and carbon source are necessary nutrient sources for microbial growth and reproduction. By changing the type and amount of carbon source and nitrogen source in the culture medium, the phenotypic changes and antibiotic synthesis of the strain can be greatly affected. On the basis of the basic MM medium, the basic medium (Minimal medium, MM) L-asparagine 0.1g, K2HPO4 0.05g, MgSO4·7H2O0.02g, FeSO4·7H2O 0.001g, glucose 1g, agar 1g, water 100mL, on this basis, L-asparagine was replaced with different nitrogen sources (Leu, Ile, Asp, Asn, Glu, GluNa+) and glucose was replaced with different carbon sources (D-mannitol, glucose, arabinose, fructose, sucrose), and the phenotype of the ΔohkB mutant strain under different carbon source or nitrogen source conditions was observed. At the same time, within the pH requirement range of the basic MM medium, a pH gradient (7.00, 7.05, 7.10, 7.15 and 7.20) was set. The carbon source suitable for the growth of the ΔohkB mutant strain was preliminarily explored, and the carbon source, nitrogen source and pH conditions that were conducive to antibiotic synthesis and yield determination were screened.

[0062] Through phenotypic observation of the ΔohkB mutant, it was found that the optimal growth condition for the ΔohkB deletion mutant was MM medium with D-mannitol as the carbon source, asparagine as the nitrogen source, and a pH of 7.10. In order to explore the effect of isolated histidine kinase OhkB on the production of undecylprodigiosin, the ΔohkB deletion mutant was cultured in an optimized medium. Starting from 48 hours, samples from the shake flask were taken every 24 hours for antibiotic production determination. The supernatant was aspirated and the absorbance was measured at a wavelength of 640nm (for measuring actinomycin) and 530nm (for measuring undecylprodigiosin) to estimate the production of undecylprodigiosin and actinomycin. The results are shown in Figure 2. Figure 6 and Figure 7 As shown, under the culture conditions, the RED production in the ΔohkB mutant was always higher than that in the starting strain M145. By measuring the OD value at a wavelength of 530nm at the 144th hour, the RED production in the ΔohkB mutant was 2.8 times higher than that in the control strain M145. Under the culture conditions, the RED production in the ΔohkB mutant was always higher than that in the starting strain M145. By measuring the OD value at a wavelength of 640nm at the 144th hour, the ACT production in the ΔohkB mutant was 3.9 times higher than that in the control strain M145.

[0063] Example 4 Backfill Experiment Verification

[0064] In this example, in order to further determine the role of the ohkB gene in the antibiotic production of Streptomyces coelicolor, an ohkB gene overexpression vector was constructed and transformed into 145 and ΔohkB deletion mutants for phenotypic observation. The specific implementation operations are as follows:

[0065] The ohkB gene fragment was amplified using the spore suspension of Streptomyces coelicolor M145 as a template and recovered after double digestion with Ndel and EcoRI; pIB139 was digested with Ndel and EcoRI and recovered. The fragment was connected to the linearized pIB139 vector through the sticky ends to obtain the plasmid pIB139-ohkB, which was sent to the company for sequencing and identification. The results are as follows Figure 8 shown.

[0066] pIB139 and pIB139-ohkB were transformed into ET12567 / pUZ8002 competent cells and integrated into the genome of M145 and ΔohkB strains by conjugation transfer to obtain M145 / pIB139, ΔohkB / pIB139, overexpression strain M145 / pIB139-ohkB and complementation strain ΔohkB / pIB139-ohkB strains. The results are shown in Fig. 9 As shown, from the observation of the phenotype, it can be seen that the phenotype of the complemented strain returned to the wild type, while the overexpression strain showed no obvious phenotype.

[0067] Example 5 Determination of antibiotic production

[0068] In this example, the four engineered bacteria constructed in Example 4 were cultured, and the yields of undecyl prodigiosin and actinomycin were measured. The specific implementation operations are as follows:

[0069] Take 25 μL of spores and inoculate them into a 250 mL shake flask containing 25 mL of MM medium (MM medium consists of 0.1 g of L-asparagine, 0.05 g of K2HPO4, 0.02 g of MgSO4·7H2O, 0.001 g of FeSO4·7H2O, 1 g of D-mannitol, 100 mL of water, pH 7.10), and culture at 30°C and 230 rpm for 5-6 days. Starting from 48 h, collect 1 mL of samples every 24 h, let them stand to allow the bacteria to precipitate, and take the supernatant to measure OD 640 OD 530 , to estimate the amount of RED synthesized. Fig.10 As shown, compared with M145 / pIB139, the RED production in ΔohkB / pIB139 increased by 3.3 times, the RED production in ΔohkB / pIB139 increased by 3.4 times, and the production of RED and ACT in the complemented strain ΔohkB / pIB139-ohkB was restored, demonstrating that the OhkB gene negatively regulates the biosynthesis of RED and ACT.

[0070] Finally, it is noted that the above embodiments are only used to illustrate the technical solutions of the present invention, rather than to limit it. Although the present invention has been described in detail with reference to the aforementioned embodiments, those skilled in the art should understand that they can still modify the technical solutions described in the aforementioned embodiments, or replace some or all of the technical features therein with equivalents. However, these modifications or replacements do not cause the essence of the corresponding technical solutions to deviate from the scope of the technical solutions of the embodiments of the present invention.

Claims

1. A method for constructing an engineering bacterium with high yield of undecyl prodigiosin, characterized in that: Streptomyces coelicolor was used as the starting bacterium to inhibit the expression of OhkB gene; the nucleotide sequence of the OhkB gene is shown in SEQ ID NO.

1.

2. The method for constructing an engineered bacterium with high yield of undecyl prodigiosin according to claim 1, characterized in that: The method comprises the following steps: constructing a CRISPR / Cas9 vector of the OhkB gene, transforming the vector into Streptomyces coelicolor, and knocking out the OhkB gene.

3. The method for constructing an engineered bacterium with high yield of undecyl prodigiosin according to claim 2, characterized in that: The construction of the CRISPR / Cas9 vector of the OhkB gene is as follows: the obtained sgRNA fragment containing the OhkB gene knockout is connected to the linearized starting vector to obtain the CRISPR / Cas9 vector of the OhkB gene.

4. The method for constructing an engineered bacterium with high yield of undecyl prodigiosin according to claim 3, characterized in that: The nucleotide sequence of the sgRNA of the OhkB gene is shown in SEQ ID NO.

3.

5. The method for constructing an engineered bacterium with high yield of undecyl prodigiosin according to claim 3 or 4, characterized in that: The starting vector is pSET-dCas9.

Images