CN105985924B - A kind of streptomyces chatanoogensis and its application through gene delection transformation - Google Patents
A kind of streptomyces chatanoogensis and its application through gene delection transformation Download PDFInfo
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Abstract
The present invention relates to a kind of streptomyces chatanoogensis through gene delection transformation and its applications.The related gene of the generation of impurity generated during a kind of production natamycin with streptomyces chatanoogensis is disclosed for the first time, and obtains the streptomyces chatanoogensis that related gene knocks out.
Description
Technical field
The invention belongs to field of biotechnology, more particularly it relates to which a kind of proper tower through gene delection transformation is exerted
Add streptomycete and its application.
Background technique
Natamycin is a kind of polyene macrolide antifungal antibiotic, due to its efficiently, wide spectrum, it is safe the advantages that quilt
It is widely used in food preservative and drug.It can effectively inhibit the growth of most of yeast and mould, prevent the shape of mycotoxin
At；Since it is insoluble in water and grease, it is difficult to be absorbed by the stomach of animal or human body, thus to the toxicity of mammalian cell
It is extremely low.
As food preservative, natamycin has the taste that low dosage, high efficiency, antibacterial time are long, do not influence food
The advantages that.U.S. FDA (Food and Drug Administration) has been approved by natamycin as food preservative, and
It is recommended that natamycin is used as food additives, it is also classified as the column of GRAS product.1997, China's food additives committee
Member can evaluate natamycin and suggest that approval uses, and be included in food additives using standard, product name is
Natamycin (Natamycin TM).Natamycin is also used as high-efficient antibacterial agent to be applied in the drugs such as eye drops or ointment, for controlling
Treat the Fungal Skin Infections diseases such as fungal keratitis, foot moss.Existing more than 30 a state approvals use it as naturally at present
Biological food preservative and antibacterial additives.
The zymophyte for being usually used in producing natamycin at present both at home and abroad has brown yellow spore streptomycete, Streptomyces natalensis, Qia Ta
It exerts and adds streptomycete.In above-mentioned bacterial strains, wherein streptomyces chatanoogensis yield is higher, but in addition to receiving, he is mould in its fermentation process
Larger amount of uranidin substance is also generated except element, such yellow substance affects greatly the product quality of natamycin.
Summary of the invention
The purpose of the present invention is to provide a kind of streptomyces chatanoogensis through gene delection transformation and its applications.
In the first aspect of the present invention, a kind of streptomyces chatanoogensis genetic engineering bacterium, ORF1240 in the bacterial strain are provided
Gene and ORF1241 gene are not expressed.
In a preferred embodiment, ORF1240 has been knocked out in the genome of the streptomyces chatanoogensis genetic engineering bacterium
Gene and ORF1241 gene.
In another preferred example, in the streptomyces chatanoogensis genetic engineering bacterium, nucleosides shown in SEQ ID NO:1
Acid sequence missing.
In another preferred example, the nucleotides sequence as shown in SEQ ID NO:2 of nucleotide sequence shown in SEQ ID NO:1
Column replacement (including all replacement or partial replacement).
In another preferred example, in the streptomyces chatanoogensis genetic engineering bacterium, by the method for homologous recombination into
The row knockout；Preferably, the bacterial strain the preparation method is as follows:
(1) the pre- nucleotide sequence fragment importing for knocking out ORF1240/1241 gene is contained into ORF1240/1241 gene
Cosmid's goes out in bacterium germination, obtains intermediate bacterium, extracts the plasmid of the intermediate bacterium, obtains missing gene by homologous recombination
The cosmid of ORF1240/1241；
(2) the cosmid engagement of ORF1240/1241 gene delection is transduceed into streptomyces chatanoogensis
In (Streptomyces chattanoogensis), the gene work of ORF1240/1241 gene delection is obtained by homologous recombination
Journey recombinant bacterium.
In another preferred example, in the streptomyces chatanoogensis genetic engineering bacterium, the pre- knockout ORF1240/
The nucleotide sequence of the nucleotide sequence fragment of 1241 genes is as shown in SEQ ID NO:2；Or the streptomyces chatanoogensis is
Streptomyces chatanoogensis L10.
In another preferred example, even if the nucleotide sequence fragment of the pre- knockout ORF1240/1241 gene is being suitable for
Under conditions of expression, active albumen can not be also encoded.
In another preferred example, the streptomyces chatanoogensis genetic engineering bacterium is in Chinese microorganism strain preservation management
The deposit number of committee's common micro-organisms center is CGMCC No.9479.
In another aspect of this invention, a kind of method preparing the streptomyces chatanoogensis genetic engineering bacterium is provided,
The described method includes: knocking out ORF1240/1241 gene from streptomyces chatanoogensis genome by the method for homologous recombination.
In a preferred embodiment, the method is as follows:
(1) the pre- nucleotide sequence fragment importing for knocking out ORF1240/1241 gene is contained into ORF1240/1241 gene
Cosmid's goes out in bacterium germination, obtains intermediate bacterium, extracts the plasmid of the intermediate bacterium, obtains missing gene by homologous recombination
The cosmid of ORF1240/1241；
(2) the cosmid engagement of ORF1240/1241 gene delection is transduceed into streptomyces chatanoogensis
In (Streptomyces chattanoogensis), the gene work of ORF1240/1241 gene delection is obtained by homologous recombination
Journey recombinant bacterium.
In another preferred example, the nucleotide sequence of the pre- nucleotide sequence fragment for knocking out ORF1240/1241 gene
As shown in SEQ ID NO:2；Or the streptomyces chatanoogensis is streptomyces chatanoogensis L10.
In another aspect of this invention, the purposes of any streptomyces chatanoogensis genetic engineering bacterium in front is provided,
For producing natamycin.
In another aspect of this invention, a kind of method producing natamycin is provided, which comprises
(1) any streptomyces chatanoogensis genetic engineering bacterium in culture front, obtains cultured products；
(2) natamycin is obtained from cultured products.
Other aspects of the invention are apparent to those skilled in the art due to this disclosure
's.
Detailed description of the invention
The electrophoresis proof diagram of Fig. 1, ORF1240/1241 gene knockout engineering bacteria pcr amplification product, wild type accordingly expand
Segment is about 1850bp, knocks out the corresponding amplified fragments of strain and is about 1200bp.
Wherein, 1 is negative control, and 2 knock out the verifying of strain external primers for ORF1240/1241, and 3 be wild type external primers,
4 be DL2000Marker.
Fig. 2, ORF1240/1241 gene knockout engineering bacterium fermentation HPLC figure.Wild type and knockout strain 120h fermentation liquid yellow
Element detection.
Wherein 1#, 2#, 3#, 4# are uranidin, can be seen from the chart that fermentation 120h knocks out strain and generates without uranidin.
Fig. 3, wild type and ORF1240/1241 gene knockout engineering bacterium fermentation 120h fermentation liquid HPLC detection, can from figure
ORF1240/1241 gene pairs natamycin yield is knocked out substantially without influence to find out.
Specific embodiment
The present inventor is dedicated to finding for the first time using streptomyces chatanoogensis production natamycin by in-depth study
The related gene of the generation of impurity of generation during a kind of production natamycin with streptomyces chatanoogensis, by the clpp gene
Except the natamycin product that impurity reduction, Quality advance later, can be obtained.The present invention is completed on this basis.
As used herein, " knockout " or " deletion " refers to the technology for deleting target gene from genome.
As used herein, " uranidin " or " yellow substance " refers to that streptomyces chatanoogensis produces natamycin mistake
Yellow is presented in the compound impurities generated in journey, and this impurity and natamycin, which coexist, leads to natamycin product quality not
Ideal, reduces product quality, and downstream purification is difficult.
As used herein, " ORF1240/ORF1241 " refers to ORF1240 gene and ORF1241 gene.
As described herein, unless otherwise stated, " target gene " is ORF1240 gene and ORF1241 gene.
The present invention provides a kind of streptomyces chatanoogensis by genetic modification, in the bacterial strain ORF1240 gene and
ORF1241 gene is not expressed substantially." not the expressing substantially " refers to that streptomyces chatanoogensis gene is not expressed or low
Expression.Wherein, ORF1240 gene and ORF1241 gene low expression refer to ORF1240 gene and ORF1241 gene in the bacterial strain
Expression quantity be lower than wild type streptomyces chatanoogensis 20%；Preferably less than wild type streptomyces chatanoogensis
10%；It is optimal to be lower than 2% more preferably lower than the 5% or lower of wild type streptomyces chatanoogensis.
The bacterial strain that ORF1240 gene and ORF1241 gene are not expressed substantially can be inhibited by various genes, gene is heavy
The technologies such as silent, gene knockout construct.For example, can by based on the gene Knockout of homologous recombination come by ORF1240 base
Cause and ORF1241 gene are knocked out from chromosome, so that ORF1240 gene and ORF1241 gene delection；It can be directed to
ORF1240 gene and ORF1241 gene design interferential RNA or GEM 132 to make ORF1240 gene and ORF1241 gene
Expression inhibiting or silencing.
It is a kind of that the method for ORF1240 gene and ORF1241 gene delection is made to be gene as preferred embodiment of the invention
Knockout technology, in a preferred embodiment of the invention, external structure are used to knock out the plasmid of target gene, pass through homologous recombination
Method, the ORF1240 gene and ORF1241 gene knockout on streptomyces chatanoogensis chromosome.Containing aburamycin
Culture medium on, what can be grown is that the bacterial strain of homologous recombination has occurred in genome, carries out screening homologous double cross wherein
The bacterial strain changed.
When the correlation between ORF1240 gene and ORF1241 gene and the generation of uranidin is disclosed it by the present inventor
Afterwards, those skilled in the art can lower to ORF1240 gene and ORF1241 gene the transformation of its expression in a variety of ways, this
The bacterial strain obtained after a little transformations and transformation should be included in the present invention.For example, when taking the method for gene knockout to implement
When transformation, other than gene knockout method detailed above, the target gene that those skilled in the art can provide according to the present invention,
Other regions that knock out (are included in these regions insertion exogenous sequences, or make on selection ORF1240 gene and ORF1241 gene
These regions missing is deleted), it can also implement to knock out by other segments other than insertion aburamycin.In addition, can also
Critical mutation is introduced so that gene expression product loss of function, gene low expression or is not expressed or expression product in short, allowing
Inactive method belongs to the knowledge that those skilled in the art are grasped, these methods can be all introduced in the present invention.
It will be containing the pre- construction (such as plasmid) for knocking out segment it should be understood that various transformation technologies well known in the art can be used
It is transferred in host strain.In a preferred embodiment, plasmid electricity is transferred in host strain.Method identification well known in the art can be used
Whether the host strain converted is required host strain.It is identified for example, PCR method can be used.
In one particular embodiment of the present invention, the ORF1240/ in above-mentioned knockout streptomyces chatanoogensis genome
1241 genes are realized by the method for homologous recombination.More specifically, the method for above-mentioned homologous recombination includes the following steps:
(1) cosmid containing ORF1240/1241 gene will be imported containing the pre- segment for knocking out ORF1240/1241 gene
Go out in bacterium germination, obtain intermediate bacterium, extract the plasmid of the intermediate bacterium, obtain the cosmid of missing gene ORF1240/1241；
(2) cosmid of ORF1240/1241 gene delection is engaged in streptomyces chatanoogensis of transduceing, is obtained
The gene engineering recombinant bacterium of ORF1240/1241 gene delection.
Above-mentioned bacterium germination out is prepared according to the method included the following steps: the cosmid containing ORF1240/1241 gene is led
Enter in E. coli BW25113/PIJ790, obtains out bacterium germination；The cosmid of above-mentioned ORF1240/1241 gene delection
It is engaged in streptomyces chatanoogensis of transduceing by E.coli ET12567/PUZ8002.
In a particular embodiment, streptomyces chatanoogensis described in SEQ ID NO:1 is streptomyces chatanoogensis L10.
Streptomyces chatanoogensis by genetic modification of the invention can be applied to production natamycin.It is demonstrated experimentally that
For the bacterial strain compared with wild type streptomyces chatanoogensis, tunning does not generate yellow substance, facilitates the separation of natamycin
Purifying, and improve natamycin product quality.
Present invention will be further explained below with reference to specific examples.It should be understood that these embodiments are merely to illustrate the present invention
Rather than it limits the scope of the invention.In the following examples, the experimental methods for specific conditions are not specified, usually according to conventional strip
Part such as J. Pehanorm Brooker etc. is write, Molecular Cloning:A Laboratory guide, the third edition, Science Press, condition described in 2002, or
According to the normal condition proposed by manufacturer.
The culture medium prescription applied in following embodiments is as follows:
LB culture medium culture:
Peptone 10g, yeast extract 5g, NaCl 10g, adds water to be settled to 1000mL, then adds if solid medium
1.5% agar powder.
2 × YT culture medium prescription:
Peptone 16g, yeast extract 10g, NaCl 5g, adds water to be settled to 1000mL, pH7.0.
MS culture medium:
Mannitol 20g, soybean powder 20g, agar 15g add water to be settled to 1000mL.
YEME fluid nutrient medium:
Yeast extract 3g, peptone 5g, yeast extract 3g, glucose 10g.
In following embodiments, the nucleotide sequence of ORF1240/1241 gene such as SEQ ID NO:1.
Embodiment 1, PCR-Targeting technology construct ORF1240/1241 gene knockout engineering bacteria
1, the segment for knocking out ORF1240/1241 gene in advance obtains
According to streptomyces chatanoogensis ORF1240/1241 gene order and the aburamycin resistance of plasmid pHY773
The primer (primer 1 and primer 2) of a pair of long 59nt and 58nt of sequence design, wherein primer 1 includes that the aburamycin of 20nt is anti-
Property the FRT sequence of upstream region of gene and one section of 39nt sequence of ORF1240/1241 upstream region of gene, primer 2 includes the Ah of 19nt
The complementary series of the 39nt of the FRT sequence and ORF1240/1241 downstream of gene in Bradley mycin resistant gene downstream.
Two primer sequences are as follows:
Primer 1:
5’gcggttcaccccccagggccgccgcgaggacgtccagcaATTCCGGGGATCCGTCGAC C 3’(SEQ
ID NO:3)
Primer 2:
5’cgaccggtgtcttggcgagcttctcggtttcctgcgcggTGTAGGCTGGAGCTGCTTC 3’(SEQ
ID NO:4)
Using the plasmid pHY773 containing A Bula resistance as template, PCR amplification is carried out using primer 1 and primer 2, is obtained
PCR product is the segment for knocking out ORF1240/1241 gene in advance, and nucleotides sequence is classified as the SEQ ID NO in sequence table:
2。
2, the cosmid containing ORF1240/1241 gene rotates into E.coli BW25113/PIJ790
1%E.coli BW25113/PIJ790 is inoculated in LB liquid medium of the 5 μ L containing 25 μ g/mL chloramphenicol, 30
DEG C, 200rpm overnight incubation.The bacterium solution for taking 100 μ L to shake up is inoculated in 10mL MgSO containing 20mM4And 25 μ g/L chloramphenicol
In SOB culture medium, 30 DEG C, 200rpm cultivates 3~4h to OD600 about 0.4.Thallus is collected in centrifugation, the rear sterilizing 10% with pre-cooling
Glycerol purging makes thallus suspend and is centrifuged to collect thallus again, and 10% glycerol of sterilizing of 100 μ L pre-cooling is added in repetition after washing 3 times, mixes
It takes 50 μ L to be added in electric shock cup after closing uniformly, while 100ng cosmid::ORF1240/1241 mixing, electric converter 200 is added
Ω, 25 μm of F, 2.5KV electricity turn 4ms.1mL pre-cooling is added in electricity immediately 30 DEG C of concussions of LB liquid medium after turning are incubated for 1h.Bacterium solution
It is uniformly coated on containing 50 μ g/mL ammonia benzyl mycins, on the LB solid medium of 25 μ g/mL chloramphenicol, 30 DEG C of overnight incubations are obtained
Bacterium colony be the E.coli BW25113/pIJ790 for being transferred to ORF1240/1241 cosmid, be named as E.coli BW25113/
pIJ790/1240。
3, the acquisition of the cosmid of ORF1240/1241 gene delection
The 100 μ L of E.coli BW25113/pIJ790/1240 that take shake culture overnight above-mentioned 2 obtain in contain 50 μ
G/mL ammonia benzyl mycin, in the SOB culture medium of 25 μ g/mL chloramphenicol, the concentration that 100 μ L are added is the L-arabinose solution of 1M,
30 DEG C, 200rpm cultivates 3~4h to OD600 about 0.4, and thallus is centrifuged after simultaneously 10% glycerol washes 3 times with 2 the methods adds 50 μ L extremely
In electric shock cup, and SEQ ID NO:2 segment (the pre- piece for knocking out ORF1240/1241 gene prepared in 100ng step 1 is added
Section), electric converter 200 Ω, 25 μm of F, 2.5KV electricity turn 4ms.1mL pre-cooling is added in electricity 0 DEG C of concussion of LB liquid medium after turning is incubated
Educate 1h, bacterium solution is uniformly coated on containing 50 μ g/mL ammonia benzyl mycins, on the LB solid medium of 50 μ g/mL aburamycins, 37 DEG C
Overnight incubation.
The single colonie of picking growth, is inoculated with LB overnight incubation, and extract plasmid, and what is obtained is ORF1240/1241 gene
The cosmid of missing.
4, the building of the streptomyces chatanoogensis of ORF1240/1241 gene delection
Transduction is combined to enter by E.coli ET12567/PUZ8002 the cosmid of ORF1240/1241 gene delection
The streptomyces coelicolor of ORF1240/1241 gene delection is obtained in streptomyces chatanoogensis L10, specific building is as follows:
E.coli12567/PUZ8002 competent cell is added in the cosmid of 3 μ L ORF1240/1241 gene delections
In, 30min is placed on ice, places 2min on ice after 42 DEG C of heat shock 90s immediately, and 37 DEG C of oscillation incubations of 1mL LB culture medium are added
1h, centrifugation are fallen supernatant and are coated on containing 50 μ g/mL aburamycins, 25 μ g/mL chloramphenicol to about 100 μ L, that is mould for 50 μ g/mL cards
On the LB solid medium of element, 37 DEG C of overnight incubations, picking single bacterium, which is fallen within, to be added on the 10mL LB liquid medium for stating antibiotic,
37 DEG C, 200rpm cultivates 3~4h to OD600 about 0.4.Thalline were collected by centrifugation, and isometric antibiotic-free LB is washed 2~3 times, centrifugation
Thallus is collected, with 2 × YT suspension thalline of 0.5mL.
By the spore suspension of streptomyces chatanoogensis L10 in 2 × YT of 0.5mL, 45 DEG C of water-bath 10min take out cooling
To room temperature.
The E.coli12567/PUZ8002 suspension and streptomycete spore of cosmid containing ORF1240/1241 gene delection
Thallus is received in sub- suspension mixing, 4000rpm centrifugation, is coated on MS plate, and 30 DEG C of 16~20h of culture are rear to be coated with 30 μ L naphthyridones
Sour (30mg/mL) and 30 μ L aburamycins (50mg/mL), 30 DEG C are cultivated 3 days, the external primers of the single colonie design grown
(primer 4 and primer 5) carries out PCR screening, and verification result is shown in Fig. 1.Therefore, final ORF1240/1241 gene knockout work is obtained
Journey bacterium, is named as L10-YPKO.
Primer 4:
GCGAGCTTCTCGGTTTCCTG(SEQ ID NO:5)；
Primer 5:
CCACACGATGGAGGAGCGCTA(SEQ ID NO:6)。
Embodiment 2 utilizes the production of ORF1240/1241 gene knockout engineering bacteria
The ORF1240/1241 gene knockout engineering bacteria and wild-type strain that previous embodiment 1 obtains, are such as issued
Ferment:
YEME culture medium inoculated (50mL triangular flask adds 10mL culture medium, and 5 beades are added), 30 DEG C, 200rpm,
It is transferred afterwards for 24 hours into fresh YEME culture medium (50mL triangular flask adds 10mL culture medium, and 5 beades are added), inoculum concentration
1%, 30 DEG C, 200rpm, ferment 96h, and sampling plus 5 times of volumes methanol concussion 30min crack thallus, and centrifuging and taking supernatant carries out
HPLC detection.
HPLC detection method: A phase (+0.1% trifluoroacetic acid of water), B phase (+0.1% trifluoroacetic acid of methanol), B phase is from 10%
(0min) rises to 100% (30min).Detector setting: uranidin detects 400nm, natamycin 303nm.Chromatographic column:
Agilent, XDB-C18 column (5 μm, 4.6*150mm).
Fig. 2 is shown in ORF1240/1241 gene knockout engineering bacteria and wild type fermentation liquid uranidin HPLC detection, shows
ORF1240/1241 gene knockout engineering bacteria does not produce uranidin really, and yellow is clearly present in the tunning of wild-type strain
Element.Therefore, ORF1240/1241 gene knockout engineering bacteria of the invention can significantly reduce impurity uranidin in tunning
Content, so that tunning quality significantly improves.
Fig. 3 is shown in the HPLC detection of wild type fermentation liquid natamycin, it is seen then that knocks out ORF1240/1241 gene pairs and receives that he is mould
Plain yield is substantially without influence.
Biomaterial preservation
Streptomyces chatanoogensis (Streptomyces chattanoogensis) in the present invention, by China Microbiological
Culture presevation administration committee common micro-organisms center (China, Beijing) preservation, preservation date: on July 18th, 2014, preservation
Number: CGMCC No.9479.
All references mentioned in the present invention is incorporated herein by reference, independent just as each document
It is incorporated as with reference to such.In addition, it should also be understood that, after reading the above teachings of the present invention, those skilled in the art can
To make various changes or modifications to the present invention, such equivalent forms equally fall within model defined by the application the appended claims
It encloses.
Claims (10)
1. the gene that one kind is prepared using streptomyces chatanoogensis (Streptomyces chattanoogensis) L10 as starter bacteria
Engineering bacteria, which is characterized in that ORF1240 gene and ORF1241 gene are not expressed in the genetic engineering bacterium, the gene work
In the genome of journey bacterium, sequential nucleotide deletion shown in SEQ ID NO:1.
2. the genetic engineering bacterium prepared as described in claim 1 using streptomyces chatanoogensis L10 as starter bacteria, feature exist
In having knocked out ORF1240 gene and ORF1241 gene in the genome of the genetic engineering bacterium.
3. the genetic engineering bacterium prepared as claimed in claim 2 using streptomyces chatanoogensis L10 as starter bacteria, feature exist
In carrying out the knockout by the method for homologous recombination.
4. the genetic engineering bacterium prepared as claimed in claim 3 using streptomyces chatanoogensis L10 as starter bacteria, feature exist
In, the genetic engineering bacterium the preparation method is as follows:
(1) the pre- nucleotide sequence fragment importing for knocking out ORF1240/1241 gene is contained into ORF1240/1241 gene
Cosmid's goes out in bacterium germination, obtains intermediate bacterium, extracts the plasmid of the intermediate bacterium, obtains missing gene by homologous recombination
The cosmid of ORF1240/1241；
(2) cosmid of ORF1240/1241 gene delection is engaged in the streptomyces chatanoogensis L10 that transduces, by homologous heavy
Group obtains the gene engineering recombinant bacterium of ORF1240/1241 gene delection.
5. the genetic engineering bacterium prepared as claimed in claim 4 using streptomyces chatanoogensis L10 as starter bacteria, feature exist
In the nucleotide sequence of the pre- nucleotide sequence fragment for knocking out ORF1240/1241 gene is as shown in SEQ ID NO:2.
6. the genetic engineering bacterium prepared as described in claim 1 using streptomyces chatanoogensis L10 as starter bacteria, feature exist
In the genetic engineering bacterium is CGMCC in the deposit number of China Committee for Culture Collection of Microorganisms's common micro-organisms center
No.9479。
7. a kind of side for preparing the genetic engineering bacterium described in claim 1 prepared using streptomyces chatanoogensis L10 as starter bacteria
Method, which is characterized in that the described method includes: being knocked out from streptomyces chatanoogensis L10 genome by the method for homologous recombination
ORF1240/1241 gene, so that sequential nucleotide deletion shown in SEQ ID NO:1.
8. the method for claim 7, which is characterized in that the method is as follows:
(1) the pre- nucleotide sequence fragment importing for knocking out ORF1240/1241 gene is contained into ORF1240/1241 gene
Cosmid's goes out in bacterium germination, obtains intermediate bacterium, extracts the plasmid of the intermediate bacterium, obtains missing gene by homologous recombination
The cosmid of ORF1240/1241；
(2) cosmid of ORF1240/1241 gene delection is engaged in the streptomyces chatanoogensis L10 that transduces, by homologous heavy
Group obtains the gene engineering recombinant bacterium of ORF1240/1241 gene delection.
9. the purposes of any genetic engineering bacterium prepared using streptomyces chatanoogensis L10 as starter bacteria of claim 1-6,
It is characterized in that, for producing natamycin.
10. a kind of method for producing natamycin, which is characterized in that the described method includes:
(1) any genetic engineering bacterium prepared using streptomyces chatanoogensis L10 as starter bacteria of culture claim 1-6,
Obtain cultured products；
(2) natamycin is obtained from cultured products.
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