CN114940982B - Apigenin prepared from genetically engineered bacteria and application thereof in polyester fiber manufacturing - Google Patents
Apigenin prepared from genetically engineered bacteria and application thereof in polyester fiber manufacturing Download PDFInfo
- Publication number
- CN114940982B CN114940982B CN202210643356.1A CN202210643356A CN114940982B CN 114940982 B CN114940982 B CN 114940982B CN 202210643356 A CN202210643356 A CN 202210643356A CN 114940982 B CN114940982 B CN 114940982B
- Authority
- CN
- China
- Prior art keywords
- apigenin
- fns
- prepared
- polyester fiber
- flavone synthase
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Active
Links
Images
Classifications
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N9/00—Enzymes; Proenzymes; Compositions thereof; Processes for preparing, activating, inhibiting, separating or purifying enzymes
- C12N9/0004—Oxidoreductases (1.)
- C12N9/0071—Oxidoreductases (1.) acting on paired donors with incorporation of molecular oxygen (1.14)
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N15/00—Mutation or genetic engineering; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification; Use of hosts therefor
- C12N15/09—Recombinant DNA-technology
- C12N15/63—Introduction of foreign genetic material using vectors; Vectors; Use of hosts therefor; Regulation of expression
- C12N15/70—Vectors or expression systems specially adapted for E. coli
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12P—FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE
- C12P17/00—Preparation of heterocyclic carbon compounds with only O, N, S, Se or Te as ring hetero atoms
- C12P17/02—Oxygen as only ring hetero atoms
- C12P17/06—Oxygen as only ring hetero atoms containing a six-membered hetero ring, e.g. fluorescein
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12Y—ENZYMES
- C12Y114/00—Oxidoreductases acting on paired donors, with incorporation or reduction of molecular oxygen (1.14)
- C12Y114/11—Oxidoreductases acting on paired donors, with incorporation or reduction of molecular oxygen (1.14) with 2-oxoglutarate as one donor, and incorporation of one atom each of oxygen into both donors (1.14.11)
- C12Y114/11022—Flavone synthase (1.14.11.22)
-
- D—TEXTILES; PAPER
- D01—NATURAL OR MAN-MADE THREADS OR FIBRES; SPINNING
- D01F—CHEMICAL FEATURES IN THE MANUFACTURE OF ARTIFICIAL FILAMENTS, THREADS, FIBRES, BRISTLES OR RIBBONS; APPARATUS SPECIALLY ADAPTED FOR THE MANUFACTURE OF CARBON FILAMENTS
- D01F1/00—General methods for the manufacture of artificial filaments or the like
- D01F1/02—Addition of substances to the spinning solution or to the melt
- D01F1/10—Other agents for modifying properties
- D01F1/103—Agents inhibiting growth of microorganisms
-
- D—TEXTILES; PAPER
- D01—NATURAL OR MAN-MADE THREADS OR FIBRES; SPINNING
- D01F—CHEMICAL FEATURES IN THE MANUFACTURE OF ARTIFICIAL FILAMENTS, THREADS, FIBRES, BRISTLES OR RIBBONS; APPARATUS SPECIALLY ADAPTED FOR THE MANUFACTURE OF CARBON FILAMENTS
- D01F6/00—Monocomponent artificial filaments or the like of synthetic polymers; Manufacture thereof
- D01F6/88—Monocomponent artificial filaments or the like of synthetic polymers; Manufacture thereof from mixtures of polycondensation products as major constituent with other polymers or low-molecular-weight compounds
- D01F6/92—Monocomponent artificial filaments or the like of synthetic polymers; Manufacture thereof from mixtures of polycondensation products as major constituent with other polymers or low-molecular-weight compounds of polyesters
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N2800/00—Nucleic acids vectors
- C12N2800/10—Plasmid DNA
- C12N2800/101—Plasmid DNA for bacteria
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N2800/00—Nucleic acids vectors
- C12N2800/22—Vectors comprising a coding region that has been codon optimised for expression in a respective host
Abstract
Apigenin prepared from genetically engineered bacteria and its application in polyester fiber production are provided. The invention provides an optimized flavone synthase and a coding gene thereof, the flavone synthase is modified and optimized according to an escherichia coli expression system, and a recombinant escherichia coli engineering bacterium is obtained from the flavone synthase.
Description
Technical Field
The invention belongs to the technical field of metabolic engineering and synthetic biology, and relates to a genetically engineered bacterium for producing apigenin and application thereof.
Background
Apigenin (AP) is a natural plant flavonoid with chemical structure of 4',5, 7-trihydroxyflavone, also called apigenin and apigenin, and molecular formula C 15 H 10 O 6 Is low molecular weight flavone (relative molecular mass 270.24), and melting point 347.5 deg.C; insoluble in water, moderately soluble in hot ethanol, soluble in diluted KOH and dimethyl sulfoxide (DMSO). Apigenin is mainly found in vegetables and fruits, such as celery, tea, onion, etc. The content of celery root is the highest, and is about 75mg/kg. It can be isolated and identified by spectroscopy, chromatography, mass spectrometry, etc. The apigenin structure is shown as follows:
at present, the method for obtaining apigenin mainly adopts a plant extraction method and a chemical synthesis method, wherein the plant extraction method adopts a proper solvent or means and takes plants (all or a part of the plants) as raw materials for extraction or processing to obtain the apigenin. Taking apigenin extracted from celery as an example, picking a proper amount of celery leaves, drying at low temperature for 2h, crushing, extracting with 70% ethanol under reflux for 2h according to a material-liquid ratio of 1:10, performing rotary evaporation and concentration on an extracting solution to obtain an extract, and separating and purifying the extract by sequentially passing through a macroporous resin (XDA-1), a polyamide resin and a sephadex LH-20 chromatographic column to obtain three flavone compound monomers, namely apigenin, apigenin and chrysoeriol-7-0-glucose-2-0-apioside. Yellow delphinium and the like extract apigenin from celery by using 70% ethanol, and the ratio of the feed liquid to the liquid is found to be 1:4, the extraction time is 3 hours, and when the reflux extraction is carried out twice, the content of apigenin in the obtained celery extract is 1.41 percent. Although apigenin is widely distributed in most plants, the content of apigenin in the plants is not very high, the traditional extraction method is simple to operate, but needs long time, needs a large amount of plant materials, is limited by factors such as the growing season and the growing region of the plants, is long in extraction time, high in cost, needs to improve the extraction rate and the purity, needs to consume a large amount of organic reagents, and has serious damage to human bodies and the environment. In recent years, extraction means such as the method of cracking celery by using ultrasonic waves and glucosidase combined methods are improved and optimized so as to improve the extraction efficiency of the apigenin, however, the yield is still not high, and special instruments and equipment are required. The method of plant extraction is not suitable for the mass acquisition of apigenin. The chemical synthesis method basically adopts a semi-synthesis method using natural plant monomers as raw materials. For example, shui 261073 uses naringin as raw material, adds 1, 4-dioxane solvent, adds oxydichlordicyanobenzoquinone (DDQ) as oxidant under 95 deg.c, stirs for 8h, stands for 12h at room temperature, produces apigenin glucoside crude product, then uses 4% sodium hydroxide solution to dissolve apigenin crude product, adds ethanol to adjust pH to 5-6 (temperature should be kept at 70 deg.c to 75 deg.c before adding acid), stands for separating out crystal, uses methanol/water mixed solution to recrystallize for 2 times to obtain pure apigenin, adds 6% hydrochloric acid water solution with 7 times volume to stir for 3.5h at 95 deg.c to 100 deg.c, stops stirring after reaction, adjusts pH to 5-6, stands for 4h, after precipitation, liquid is in clear liquid state, suction filters to obtain apigenin crude product, then uses 4% sodium hydroxide solution to dissolve, adds methanol, adjusts pH to 6 with hydrochloric acid, and separates out crystal. Then refluxing and purifying by ethanol to obtain the purer apigenin. The naringin used as the substrate in the method is also a dihydroflavonoids compound. Xiaojinxia et al chemically synthesize apigenin, mainly involving two steps: using p-hydroxyacetophenone as a substrate to prepare p-hydroxyacetoacetate and prepare apigenin through cyclization reaction. The steps involved in synthesizing the flavonoid compounds by the chemical method are multiple, the process is complex, the yield is low, the extremely harsh reaction conditions are also limiting factors, more toxic reagents are used in the synthesis process, and the purification difficulty of the required compounds is increased by more byproducts.
Although the existing biosynthesis method for producing the apigenin through genetic engineering exists, the existing biosynthesis still remains at the level of using natural products, and the satisfactory effect cannot be achieved in both yield and substrate conversion rate, so that the production cost is high. Therefore, there is an urgent need to develop a production method capable of obtaining apigenin at an excessively high yield.
Disclosure of Invention
In order to fill the blank of the prior art, the invention provides an optimized flavone synthase (FNS-I) and a coding gene thereof, and the apigenin conversion rate of recombinant escherichia coli is remarkably improved by constructing a genetic engineering strain. In order to achieve the technical effects, the invention provides the following technical scheme:
in a first aspect of the invention, an optimized flavone synthase is provided, wherein a coding gene of the flavone synthase is shown in any one of SEQ ID NO. 2-4.
In a second aspect of the present invention, there is provided a recombinant host cell expressing the gene encoding flavone synthase described above.
In one embodiment, the recombinant host cell is E.coli.
In a third aspect of the invention, the application of the flavone synthase or the recombinant host cell in the production of apigenin and downstream products thereof is provided.
In one embodiment, the downstream product comprises a biofabric material comprising apigenin as described above.
In a fourth aspect of the present invention, there is provided a method for efficiently producing apigenin, which comprises transforming the recombinant host cell described above in a substrate containing naringenin.
Compared with the prior art, the invention achieves the following remarkable improvements:
the invention modifies and optimizes the existing flavone synthase aiming at an escherichia coli expression system, and obtains the recombinant escherichia coli, compared with the recombinant escherichia coli obtained without genetic modification, the yield and the conversion rate of apigenin are obviously improved, and the production cost is greatly reduced.
Drawings
The invention is described in further detail below with reference to the attached drawing figures:
FIG. 1 shows the synthesis route of apigenin.
Detailed Description
The technical solutions in the embodiments of the present invention will be clearly and completely described below, and it is obvious that the described embodiments are only a part of the embodiments of the present invention, and not all of the embodiments. All other embodiments, which can be derived by a person skilled in the art from the embodiments given herein without making any creative effort, shall fall within the protection scope of the present invention.
Example 1 FNS-I sequence optimization
1. Experimental Material
pETDuet-1 vector: purchased from Youbao organisms; plasmid extraction kit, gel recovery kit: purchased from OMEGA; restriction enzymes were purchased from NEB.
The standard compounds apigenin (CAS number: 520-36-5) and naringenin (CAS number: 480-41-1) are purchased from Shanghai leaf Biotech limited, and have purity of more than 98%. Other reagents are domestic analytical pure or chromatographic pure reagents and are purchased from national pharmaceutical group chemical reagent limited.
Clone competent cell DH5 α, expression competent cell Rosetta (DE 3): purchased from holo-gold.
Liquid phase detection conditions: phase A: 0.1% formic acid water, phase B: acetonitrile; separation conditions are as follows: 0-20min 20% B phase-55%, 20-22min 55-B phase-100%, 22-27min 100%, 27-35min100% B phase-20% B phase, 35-40min, 20-20% B phase; detection wavelength: 340nm, column temperature: at 30 ℃. And (3) chromatographic column: thermo syncronis C18 reverse phase column (250 mm. Times.4.6 mm,5 μm).
2, FNS-I sequence modification:
based on codon preference of escherichia coli, GC content, codon balance and the like are comprehensively considered, an original FNS-I sequence (SEQ ID NO.1 is optimized to obtain 3 optimized sequences FNS-IM1, FNS-IM2 and FNS-IM3, nucleotide sequences of the sequences are respectively shown in SEQ ID NO.2-4, and the three sequences are synthesized by a whole gene sequence of Suzhou Jinzhi biological science and technology limited company.
Example 2 construction of engineering bacteria
1. Respectively adding NcoI and BamHI at two ends of the three optimized FNS-I gene sequences and the original FNS-I gene sequence, synthesizing the optimized FNS-I gene by using a gene synthesis technology, and constructing cloning vectors pUC57FNS-IM1, pUC57FNS-IM2, pUC57FNS-IM3 and pUC57FNS-I;
2. constructing a recombinant expression vector, double digesting pETDuet-1 plasmid and obtained pUC57FNS-IM1, pUC57FNS-IM2, pUC57FNS-IM3 and pUC57FNS-I plasmids respectively by NcoI and BamHI, obtaining pETDuet-1 and FNS-IM1, FNS-IM2, FNS-IM3 and pUC57FNS-I fragments by glue recovery, then connecting pETDuet-1 with FNS-IM1, FNS-IM2, FNS-IM3 and pUC57FNS-I respectively by using T4DNA ligase, transforming DH5 alpha competent cells by the connecting products, coating ampicillin resistant plates, culturing overnight at constant temperature of 37 ℃, screening positive recombinants, obtaining pETDue-FNS-IM1 plasmids, pETDue-pES-IM 2 plasmids, TDue-pES-3 plasmids and FNS-IM3 plasmids by extracting the plasmids;
3. respectively transforming the pETDue-FNS-IM1 plasmid, pETDue-FNS-IM2 plasmid, pETDue-FNS-IM3 plasmid and pETDue-FNS-I plasmid obtained in the above steps into escherichia coli Rosetta (DE 3), coating an ampicillin resistance plate, inverting overnight at the constant temperature of 37 ℃ for culturing, screening to obtain positive recombinants, namely obtaining FNS-IM1, FNS-IM2, FNS-IM3 and FNS-I engineering bacteria respectively, and preserving glycerin tubes.
Example 3 production of apigenin by fermentation of engineering bacteria
FNS-IM1, FNS-IM2, FNS-IM3, FNS-I engineering bacteria strains are respectively subjected to amplification culture according to the following methods:
(1) Under the aseptic condition, taking a ring strain from a glycerol tube by using an inoculating ring, transplanting the ring strain into a primary seed culture medium, culturing at 37 ℃ and 240rpm until the OD600 is between 3.0 and 4.0 to obtain primary seed bacterial liquid;
(2) Transferring the primary seed bacterial liquid into a secondary seed culture medium according to the inoculation amount of 6-10%, and culturing at 37 ℃ and 240rpm for 3-5 h to obtain secondary seed bacterial liquid;
(3) Inoculating the cultured secondary seed bacterial liquid into a fermentation tank according to the inoculation amount of 6-10%, wherein the initial fermentation parameters are as follows: the temperature is 37 ℃, the rotating speed is 200rpm, the pH value is 7.0, the ventilation volume is 50L/h, and the tank pressure is 0.05MPa; along with the prolonging of the fermentation time, the bacteria concentration is gradually increased, and the dissolved oxygen is controlled to be 20-40% by adjusting the rotating speed and the ventilation quantity; when the dissolved oxygen and the pH value are simultaneously and rapidly increased, feeding materials in a flowing manner is started; when OD600 reaches 20-25 ℃, the temperature is reduced to 30 ℃, then 0.2mM inducer is added, the fermentation is continued for 8-12 h, then the mixture is put into a tank, and thalli are collected by low-temperature centrifugation and refrigerated.
2. Identifying the production capacity of the strain on apigenin:
(1) After FNS-IM1, FNS-IM2, FNS-IM3 and FNS-I engineering bacteria are washed by a potassium phosphate buffer solution with PH =6, cells are respectively placed in the following transformation systems for apigenin transformation (three groups are repeated): the transformation system comprises:
5% of wet bacteria of the engineering bacteria, 0.1M potassium phosphate buffer, 2.0M naringenin (apigenin synthesis substrate, figure 1), 5mM ascorbate and 10 μ MFeSO 4 The pH was adjusted to 6 with 2mol/L NaOH solution and the conversion was carried out at a temperature of 35 ℃ and 150rpm for 10 hours.
The substrate conversion rate is calculated by the substrate content existing in the conversion system, the naringenin content is calculated by preparing different concentration gradients (0, 0.05,0.1,0.5,1,2.5,5,7.5, 10, 15 and 20 g/L) of naringenin standard substances, measuring an A240 value and drawing a standard curve.
(2) And (3) conversion result:
TABLE 1
| Apigenin product concentration (mg/L) | Conversion (%) | |
| FNS-IM1 | 108.55±2.67 | 87 |
| FNS-IM2 | 121.34±1.89 | 98 |
| FNS-IM3 | 101.46±2.21 | 82 |
| FNS-I (control group) | 88.67±1.87 | 72 |
As can be seen from the results in Table 1, the optimized flavone synthases FNS-IM1, FNS-IM2 and FNS-IM3 have conversion rates superior to those of the original sequence FNS-I, wherein the FNS-IM2 has the optimal conversion capacity, and the conversion rate can reach 98%.
EXAMPLE 4 preparation of Terylene macrobio fiber containing apigenin component
Large biological fiber: bioactive molecules are added into fibers (cotton, hemp, wool, silk, viscose, polyester, nitrile and brocade) for modification, so as to produce the active fiber with biological functions. The polyester macrobio fiber containing the apigenin component prepared by the invention is an active fiber with antibacterial and antiviral functions and the like, which is prepared by adding the apigenin component in the production process of the polyester fiber.
1. Preparation of SiO by sol-gel method 2 Nano-microspheres:
weighing 0.5g of CTAB, dissolving in 1000mL of deionized water, stirring at 40 ℃, dropwise adding a NaOH solution after the solution is clarified, and adjusting the pH to 11.8; the temperature was then raised to 80 ℃; after 30min, 2.5mL TEOS is added, and the mixture is continuously stirred for 2h at the temperature of 80 ℃; standing the obtained white suspension, cooling to room temperature, centrifuging, washing with deionized water and absolute ethyl alcohol repeatedly for 3 times at a rotating speed of 9000r/min, drying the obtained white gel in a vacuum drying oven for 12 hours, placing the dried sample in a muffle furnace, and calcining at 550 ℃ for 6 hours; cooling to obtain white solid powder which is mesoporous SiO 2 And (3) nanoparticles. Grinding and crushing for later use to prepare the mesoporous SiO 2 The granularity of the nano particles is 100 +/-10 nm, and the aperture is 2.5 +/-0.5 nm; 2. concentrating and purifying the apigenin produced in the example 3 according to a conventional method in the field to obtain a pure apigenin product (the purity is more than or equal to 95%), ultrasonically dissolving the pure apigenin product by using a 95% ethanol solution to obtain a 30% apigenin suspension, and weighing 10g of mesoporous SiO prepared in the step 1 2 Sequentially adding nano powder and 0.02g of dispersant XD-5040 into 500mL of deionized water, and shearing and dispersing by using a magnetic stirrer at 250r/min; obtaining SiO containing apigenin component 2 Nano composite dispersion liquid, volatilizing the solvent of the dispersion liquid to obtain mesoporous SiO containing apigenin component 2 Grinding and crushing a molecular nest (the particle size is 9000 meshes) for later use;
firstly, the mesoporous SiO containing apigenin component prepared in step 2 2 Vacuum drying the molecular nest and the polyester chip at 180 ℃ for 24h; then, according to the following steps of 1:10, adding 0.1 percent of dispersant XD-5040, 0.5 percent of antioxidant 1098 and 0.2 percent of coupling agent KH-550 into a double-screw extruder for mixing and granulation, controlling the melting temperature at 260-280 ℃ and the screw rotating speed at 260r/min; obtaining a master batch containing apigenin; 3. mixing the mother particles containing apigenin with common polymerEster slices were as per 1:20, 0.2 percent of dispersant PT-200E, 0.2 percent of antioxidant KB-6 and 0.15 percent of coupling agent Z-6020 are added into a double-screw extruder for mixing and spinning, the spinning speed is 1700m/min, the stretching temperature is 80 ℃, the side-blown cooling air temperature is 30 ℃, the side-blown air speed is 0.5m/s, the total stretching ratio is 5 times, and the winding speed is 1050m/min, thus preparing the polyester macrobio-fiber containing apigenin.
The polyester macrobio fiber containing the apigenin component is made into fiber fabric through a spinning production line, and the bacteriostasis rate of the fiber fabric containing the apigenin component and the bacteriostasis rate of the fiber fabric after 100 times of standard washing (GB/T8629-2017) are tested according to GB/T20944.2-2007. The results are shown in table 2 below.
TABLE 2
Through detection, the fabric obtained by spinning the polyester macrobio-fiber containing the apigenin component has obvious inhibition effects on staphylococcus aureus, escherichia coli and candida albicans on the surface of human skin, and still has excellent antibacterial property after being washed for many times.
Unless otherwise stated, the proportions described in the present examples are mass proportions, and the percentages described are mass percentages.
Although the present invention has been described in detail with reference to the foregoing embodiments, it will be apparent to those skilled in the art that modifications may be made to the embodiments described above, or equivalents may be substituted for elements thereof. Any modification, equivalent replacement, or improvement made within the spirit and principle of the present invention should be included in the protection scope of the present invention.
Sequence listing
<110> Baicao Bingbio technology (Qingdao) Co., ltd
<120> apigenin prepared by genetically engineered bacteria and application thereof in polyester fiber manufacturing
<130> 5
<141> 2022-06-08
<160> 4
<170> SIPOSequenceListing 1.0
<210> 1
<211> 1074
<212> DNA
<213> Artificial Sequence
<400> 1
atggctccaa caactattac tgcattggcc aaggaaaaaa cacttaactc tgattttgtc 60
cgggatgagg atgagcgtcc caaagttgcc tacaatcaat tcagcactga aattcccatt 120
atttctttag ctggtatcga tgatgattcc aatggcagga gacctgaggt gtgtcgtaaa 180
atagtggagg ccttcgaaga ctgggggatt ttccaggtag ttgatcacgg tattgacagc 240
ggtttgatcg cggaaatgtc tcgtctgtct cgtgaattct ttgctttgcc tgccgaggag 300
aaacttcggt atgatactac tggtggaaag agaggcggct tcactatctc cactcatctt 360
cagggtgacg atgtgaagga ttggcgtgag tttgttgttt atttttcgta cccagtcgat 420
gctcgggact actcgagatg gcctgataag ccagagggat ggaggtctgt tacggaggtt 480
tatagtgaga agttgatggc gctaggtgcc aagttactgg aagtgctatc agaggccatg 540
gggcttgaaa aagaggctct tacagaggct tgtgtgaaca tggaacagaa agtgttgatt 600
aattactatc ctacatgtcc ccaaccggac ttgacacttg gagtcagaag gcacacggat 660
ccgggtacga ttaccatttt gcttcaggac atggttgggg ggttacaggc taccagggat 720
ggcggcaaaa cttggattac tgttcagcct gtcgagggag cttttgtcgt caatttgggt 780
gatcatggtc attatttgag caatggaagg ttcaagaatg ccgatcacca agcagtagtg 840
aattcaactt ctagcagatt gtctatcgca actttccaga acccggctca gaatgctata 900
gtgtatccat taaagatcag ggagggcgag aagccaattc ttgaggaggc catgacatac 960
gccgagatgt ataagaaaaa catgactaaa catattgagg tggctaccca gaagaaattg 1020
gccaaggaga aaagattgca gaacgagaag gccaagctgg agacgaaatt ttag 1074
<210> 2
<211> 1074
<212> DNA
<213> Artificial Sequence
<400> 2
atggctccag cagctattac tgcattggcc aaagaaaaaa cacttaactc tgattttgtc 60
cgggatgaag atgaacgtcc cagaattgcc tacagtcagt tcagcactga aattcccatt 120
atttctttag ctggtatcga tgatgattcc aatggcagga gacctgaagt gtgtcgtaaa 180
atagtggaag ccttcgaaaa ctgggggatt ttccaggtag ttgatcacgg tattgacagc 240
ggtttgatcg cggaaatgtc tcgtctgtct cgtgaattct ttgctttgcc tgccgaagaa 300
aaacttcggt atgatactac tggtggaaaa agaagcggct tcactatctc cactcatctt 360
cagggtgacg atgtgaaaga ttggcgtgaa tttgttgttt atttttcgta cccagtcgat 420
gctcgggact actcgaaatg gcctgataaa ccagaaggat ggaagtctgt tacggaagtt 480
tatagtgaaa agttgatggc gctaggtgcc aaattactgg aaatgctatc agaagccatg 540
gggcttgaaa aaaaggctct tacagaagct tgtgtgaaca tggaacagaa agtgttgatt 600
aattactatc ctacatgtcc ccagccggac ttgacacttg gaatcagaaa gcacacggat 660
ccgggtacga ttaccatttt gcttcaggac atggttgggg ggttacaggc taccagggat 720
ggcggcagaa cttggattac tgttcagcct gtcgaaggaa cttttgtcgt cagtttgggt 780
gatcatggtc attatttgaa cagtggaaag ttcagaaatg ccgatcacca agcagtagtg 840
aattcagctt ctagcagatt gtctatcgca actttccaga acccggctca gaatgctata 900
gtgtatccat taaaaatcag ggaaggcgaa aaaccagttc ttgaagaagc catgacatac 960
gccgaaatgt ataaaaaaaa catgactaaa catattgaag tggctaccca gaaaaaattg 1020
gccagaaaga aaaaattgca gaacgaaaaa gccagactgg agacgaaatt ttag 1074
<210> 3
<211> 1074
<212> DNA
<213> Artificial Sequence
<400> 3
atggctccag cagctattat tgcattggcc aaagaaaaaa catttaactc tgattttgtc 60
cgggatgaag atgaacgtcc cagaattgcc tatagtcagt tcagcattga aattcccatt 120
atttctttag ctggtatcga tgatgattcc aatggcagga gacctgaagt gtgtcgtaaa 180
atagtggaag ccttcgaaaa ctgggggatt ttccaggtag ttgatcatgg tattgacagc 240
ggtttgatcg cggaaatgtc tcgtctgtct cgtgaattct ttgctttgcc tgccgaagaa 300
aaacttcggt atgatattat tggtggaaaa agaagcggct tcattatctc cattcatctt 360
cagggtgacg atgtgaaaga ttggcgtgaa tttgttgttt atttttcgta cccagtcgat 420
gctcgggact actcgaaatg gcctgataaa ccagaaggat ggaagtctgt tatggaagtt 480
tatagtgaaa agttgatggc gctaggtgcc aaattattgg aaatgctatc agaagccatg 540
gggcttgaaa aaaaggctct tatagaagct tgtgtgaaca tggaacagaa agtgttgatt 600
aattattatc ctatatgtcc ccagccggac ttgacatttg gaatcagaaa gcatacggat 660
ccgggtatga ttatcatttt gcttcaggac atggttgggg ggttataggc tatcagggat 720
ggcggcagaa cttggattat tgttcagcct gtcgaaggaa cttttgtcgt cagtttgggt 780
gatcatggtc attatttgaa cagtggaaag ttcagaaatg ccgatcatca agcagtagtg 840
aattcagctt ctagcagatt gtctatcgca actttccaga acccggctca gaatgctata 900
gtgtatccat taaaaatcag ggaaggcgaa aaaccagttc ttgaagaagc catgacatat 960
gccgaaatgt ataaaaaaaa catgactaaa catattgaag tggctatcca gaaaaaattg 1020
gccagaaaga aaaaattgca gaacgaaaaa gccagactgg agacgaaatt ttag 1074
<210> 4
<211> 1074
<212> DNA
<213> Artificial Sequence
<400> 4
atggctccaa caactattac tgcattggcc aaggtaaaaa catttaactc tgattttgtc 60
cgggttgagg atgagcgtcc caaagttgcc tacaatcaat tcagcattga aattcccatt 120
atttctttaa ctggtatcga tgatgattcc aatggcaggt gacctgaggt gtgtcgtaaa 180
ataatggtgg ccttcgaaga ctgggggttt ttccaggtaa ttgatcatgg tattgacagc 240
ggtttgatcg cggtaatgtc tcgtctgtct cgtgaattct ttgctttgcc tgccgaggtg 300
aaacttcggt atgatactac tggtggtaag agaggcggct tcattatctc cattcatctt 360
cagggtgacg atgtgaaggt ttggcgtgag tttgttgttt atttttcgta cccagtcgat 420
gctcgggtct actcgagatg gcctgataag ccagagggtt ggtggtctgt tacggtggtt 480
tataatgaga agttgatggc gctaagtgcc aagttactgg aagtgctatc agaggccatg 540
gggcttgaaa aagaggctct tacagaggct tgtgtgaaca tggtacagaa agtgttgatt 600
aattactatc ctacatgtcc ccaaccggtc ttgacatttg gagtcagaag gcatacggtt 660
ccgggtacga ttaccatttt gcttcaggtc atggttgggg ggttacaggc taccagggtt 720
ggcggcaaaa cttggtttac tgttcagcct gtcgagggtg cttttgtcgt caatttgggt 780
gatcatggtc attatttgag caatggtagg ttcaagaatg ccgatcatca agcagtaatg 840
aattcaactt ctaacagatt gtctatcgca actttccaga acccggctca gaatgctata 900
gtgtatccat taaagatcag ggtgggcgag aagccaattc ttgaggtggc catgacatac 960
gccgagatgt ataagaaaaa catgactaaa catattgagg tggctaccca gaagaaattg 1020
gccaaggtga aaagattgca gaacgagaag gccaagctgg agacgaaatt ttaa 1074
Claims (7)
1. An optimized flavone synthase, wherein the coding gene of the flavone synthase is shown as any one of SEQ ID NO. 2-4.
2. A recombinant host cell expressing the gene encoding flavone synthase of claim 1.
3. The recombinant host cell of claim 2, wherein the recombinant host cell is e.
4. Use of the flavone synthase of claim 1 or the recombinant host cell of any one of claims 2-3 for the production of apigenin and its downstream products.
5. The use as claimed in claim 4, wherein the downstream product is a biofiber material containing apigenin.
6. The use of claim 5, wherein the biofiber material is polyester fiber.
7. A method for efficiently producing apigenin, comprising transforming the recombinant host cell of claim 2 or 3 in a substrate comprising naringenin.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202210643356.1A CN114940982B (en) | 2022-06-08 | 2022-06-08 | Apigenin prepared from genetically engineered bacteria and application thereof in polyester fiber manufacturing |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202210643356.1A CN114940982B (en) | 2022-06-08 | 2022-06-08 | Apigenin prepared from genetically engineered bacteria and application thereof in polyester fiber manufacturing |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN114940982A CN114940982A (en) | 2022-08-26 |
| CN114940982B true CN114940982B (en) | 2023-01-03 |
Family
ID=82909884
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN202210643356.1A Active CN114940982B (en) | 2022-06-08 | 2022-06-08 | Apigenin prepared from genetically engineered bacteria and application thereof in polyester fiber manufacturing |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN114940982B (en) |
Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2001005981A1 (en) * | 1999-07-19 | 2001-01-25 | Suntory Limited | Genes coding for flavone synthases |
| CN110616204A (en) * | 2019-09-27 | 2019-12-27 | 佛山市汇腾生物技术有限公司 | Flavone synthase and application thereof |
| CN110616162A (en) * | 2019-09-27 | 2019-12-27 | 佛山市汇腾生物技术有限公司 | Pichia pastoris for expressing flavone synthase |
| CN112725256A (en) * | 2021-02-22 | 2021-04-30 | 湖南省农产品加工研究所 | Recombinant escherichia coli and method for biosynthesizing diosmetin by using recombinant escherichia coli |
Family Cites Families (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US7338791B2 (en) * | 2004-07-10 | 2008-03-04 | The Research Foundation Of State University Of New York | Production of flavonoids by recombinant microorganisms |
-
2022
- 2022-06-08 CN CN202210643356.1A patent/CN114940982B/en active Active
Patent Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2001005981A1 (en) * | 1999-07-19 | 2001-01-25 | Suntory Limited | Genes coding for flavone synthases |
| CN110616204A (en) * | 2019-09-27 | 2019-12-27 | 佛山市汇腾生物技术有限公司 | Flavone synthase and application thereof |
| CN110616162A (en) * | 2019-09-27 | 2019-12-27 | 佛山市汇腾生物技术有限公司 | Pichia pastoris for expressing flavone synthase |
| CN112725256A (en) * | 2021-02-22 | 2021-04-30 | 湖南省农产品加工研究所 | Recombinant escherichia coli and method for biosynthesizing diosmetin by using recombinant escherichia coli |
Non-Patent Citations (2)
| Title |
|---|
| Daucus carota flavone synthase I mRNA, complete cds;GenBank: AY817675.1;《GenBank》;20050622;全文 * |
| Engineering co-culture system for production of apigetrin in Escherichia coli;Nguyen Huy Thuan 等;《J Ind Microbiol Biotechnol》;20180331;第45卷(第3期);175-185 * |
Also Published As
| Publication number | Publication date |
|---|---|
| CN114940982A (en) | 2022-08-26 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN100569938C (en) | Can produce the Cladosporium endogenetic fungus of trans-resveratrol | |
| CN107586795B (en) | Method for producing phloretin by fermentation of saccharomyces cerevisiae | |
| CN106148256B (en) | The genetic engineering bacterium and its construction method of production alpha-arbutin and application | |
| CN106754607B (en) | A kind of recombinant bacterial strain and its construction method producing tyrosol | |
| CN102206606A (en) | Recombinant escherichia coli and application thereof in production of 5-aminolevulinic acid (ALA) | |
| CN113416748A (en) | Expression vector for synthesizing cannabidiol, heterologous expression method and application | |
| CN103865862A (en) | Bacillus licheniformis strain loaded with serine acetyl transferase gene as well as building method and application thereof | |
| CN103509816A (en) | Coenzyme-Q10-production engineered bacteria construction method, engineered bacteria, and application thereof | |
| CN114940982B (en) | Apigenin prepared from genetically engineered bacteria and application thereof in polyester fiber manufacturing | |
| CN104130967A (en) | Escherichia coli with coexpression of L-lactate dehydrogenase and formate dehydrogenase as well as construction method and application of escherichia coli | |
| CN112080452B (en) | High-yield phenyllactic acid bacillus licheniformis genetically engineered bacterium, method for producing phenyllactic acid and application | |
| CN110004099B (en) | Fermentation production method of salidroside | |
| CN110305855A (en) | Rhizoma Gastrodiae GeCPR gene and its application | |
| CN106554932B (en) | Genencor engineering bacterium for producing gentamicin B and construction method thereof | |
| CN110938553A (en) | Monascus phy gene and application thereof in improvement of yield of yellow pigment | |
| CN113444737B (en) | Cytochrome P450 enzyme and application thereof in synthesis of ganoderma lucidum triterpenoid | |
| CN114134186A (en) | Method for synthesizing 5-hydroxy beta-indolyl alanine by using glucose as substrate through biological method | |
| CN114736918A (en) | Recombinant escherichia coli for producing salidroside through integrated expression and application thereof | |
| CN110066741B (en) | Mutant strain construction method for directionally improving metabolic yield of filamentous fungi through morphological optimization | |
| CN111254152B (en) | Acetylxylan esterase gene, coded product and preparation method thereof | |
| CN110029081B (en) | Engineering bacterium for over-expressing carbon catabolite repression effect transcription inhibitor gene and construction method thereof | |
| CN113817757A (en) | Recombinant yeast engineering strain for producing cherry glycoside and application | |
| CN107384923B (en) | Promoter pYLG and application thereof in construction of candida tropicalis with high yield of long-chain dicarboxylic acid | |
| CN101659960A (en) | Biological preparation method of chitin deacetylase | |
| CN105349480B (en) | The recombination bacillus coli of one plant of high efficient expression [2Fe2S] ferredoxin and its application |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| PB01 | Publication | ||
| PB01 | Publication | ||
| SE01 | Entry into force of request for substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| TA01 | Transfer of patent application right | ||
| TA01 | Transfer of patent application right |
Effective date of registration: 20221031 Address after: 266071 household B611, block B, No. 12, Hong Kong Middle Road, Shinan District, Qingdao City, Shandong Province Applicant after: BESTEE MATERIAL (TSINGTAO) Co.,Ltd. Applicant after: ZHONGKE TEXTILE RESEARCH INSTITUTE (QINGDAO) Co.,Ltd. Applicant after: QINGDAO BAICAO NEW MATERIAL CO.,LTD. Address before: 266000 611, block B, Fenghe Plaza, No. 12, Hong Kong Middle Road, Shinan District, Qingdao, Shandong Applicant before: Baicaobianda Biotechnology (Qingdao) Co.,Ltd. |
|
| GR01 | Patent grant | ||
| GR01 | Patent grant |