CN116042663A - 一种铁皮石斛β-紫罗兰酮合成关键酶基因DoCCD1及其克隆方法和应用 - Google Patents
一种铁皮石斛β-紫罗兰酮合成关键酶基因DoCCD1及其克隆方法和应用 Download PDFInfo
- Publication number
- CN116042663A CN116042663A CN202210915168.XA CN202210915168A CN116042663A CN 116042663 A CN116042663 A CN 116042663A CN 202210915168 A CN202210915168 A CN 202210915168A CN 116042663 A CN116042663 A CN 116042663A
- Authority
- CN
- China
- Prior art keywords
- doccd1
- beta
- dendrobium candidum
- ionone
- key enzyme
- 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.)
- Pending
Links
- PSQYTAPXSHCGMF-BQYQJAHWSA-N β-ionone Chemical compound CC(=O)\C=C\C1=C(C)CCCC1(C)C PSQYTAPXSHCGMF-BQYQJAHWSA-N 0.000 title claims abstract description 112
- SFEOKXHPFMOVRM-UHFFFAOYSA-N (+)-(S)-gamma-ionone Natural products CC(=O)C=CC1C(=C)CCCC1(C)C SFEOKXHPFMOVRM-UHFFFAOYSA-N 0.000 title claims abstract description 56
- 241000026010 Dendrobium candidum Species 0.000 title claims abstract description 53
- 108090000790 Enzymes Proteins 0.000 title claims abstract description 37
- 230000015572 biosynthetic process Effects 0.000 title claims abstract description 35
- 238000003786 synthesis reaction Methods 0.000 title claims abstract description 35
- 238000000034 method Methods 0.000 title claims abstract description 14
- 238000010367 cloning Methods 0.000 title claims abstract description 13
- 108090000623 proteins and genes Proteins 0.000 claims abstract description 33
- 125000003275 alpha amino acid group Chemical group 0.000 claims abstract description 25
- 102000004169 proteins and genes Human genes 0.000 claims abstract description 14
- 239000002773 nucleotide Substances 0.000 claims abstract description 13
- 125000003729 nucleotide group Chemical group 0.000 claims abstract description 13
- UPYKUZBSLRQECL-UKMVMLAPSA-N Lycopene Natural products CC(=C/C=C/C=C(C)/C=C/C=C(C)/C=C/C1C(=C)CCCC1(C)C)C=CC=C(/C)C=CC2C(=C)CCCC2(C)C UPYKUZBSLRQECL-UKMVMLAPSA-N 0.000 claims abstract description 10
- 150000001746 carotenes Chemical class 0.000 claims abstract description 10
- 235000005473 carotenes Nutrition 0.000 claims abstract description 10
- NCYCYZXNIZJOKI-UHFFFAOYSA-N vitamin A aldehyde Natural products O=CC=C(C)C=CC=C(C)C=CC1=C(C)CCCC1(C)C NCYCYZXNIZJOKI-UHFFFAOYSA-N 0.000 claims abstract description 10
- 229920001184 polypeptide Polymers 0.000 claims abstract description 6
- 102000004196 processed proteins & peptides Human genes 0.000 claims abstract description 6
- 108090000765 processed proteins & peptides Proteins 0.000 claims abstract description 6
- 108091028043 Nucleic acid sequence Proteins 0.000 claims abstract description 4
- 241000196324 Embryophyta Species 0.000 claims description 16
- 235000002637 Nicotiana tabacum Nutrition 0.000 claims description 14
- 239000013598 vector Substances 0.000 claims description 14
- 238000012408 PCR amplification Methods 0.000 claims description 13
- 241000208125 Nicotiana Species 0.000 claims description 11
- 239000002299 complementary DNA Substances 0.000 claims description 10
- 230000009466 transformation Effects 0.000 claims description 10
- 230000001052 transient effect Effects 0.000 claims description 10
- 238000006467 substitution reaction Methods 0.000 claims description 7
- 125000000539 amino acid group Chemical group 0.000 claims description 6
- 238000012217 deletion Methods 0.000 claims description 6
- 230000037430 deletion Effects 0.000 claims description 6
- 230000002194 synthesizing effect Effects 0.000 claims description 6
- 230000009261 transgenic effect Effects 0.000 claims description 6
- 238000012163 sequencing technique Methods 0.000 claims description 5
- 241000894006 Bacteria Species 0.000 claims description 4
- 230000000694 effects Effects 0.000 claims description 4
- 238000001962 electrophoresis Methods 0.000 claims description 4
- 102000004190 Enzymes Human genes 0.000 claims description 3
- 238000000246 agarose gel electrophoresis Methods 0.000 claims description 3
- 150000001413 amino acids Chemical class 0.000 claims description 3
- 230000003321 amplification Effects 0.000 claims description 3
- 230000014509 gene expression Effects 0.000 claims description 3
- 238000003199 nucleic acid amplification method Methods 0.000 claims description 3
- 238000000746 purification Methods 0.000 claims description 3
- 238000010839 reverse transcription Methods 0.000 claims description 3
- 229930002839 ionone Natural products 0.000 abstract description 10
- 150000002499 ionone derivatives Chemical class 0.000 abstract description 10
- 101150028438 CCD1 gene Proteins 0.000 abstract description 2
- 230000006870 function Effects 0.000 abstract description 2
- 241000588724 Escherichia coli Species 0.000 description 10
- IJGRMHOSHXDMSA-UHFFFAOYSA-N Atomic nitrogen Chemical compound N#N IJGRMHOSHXDMSA-UHFFFAOYSA-N 0.000 description 8
- 241001076416 Dendrobium tosaense Species 0.000 description 8
- 239000007788 liquid Substances 0.000 description 6
- 241000589158 Agrobacterium Species 0.000 description 5
- 238000006243 chemical reaction Methods 0.000 description 5
- 210000004027 cell Anatomy 0.000 description 4
- 238000010586 diagram Methods 0.000 description 4
- 239000013604 expression vector Substances 0.000 description 4
- 239000000796 flavoring agent Substances 0.000 description 4
- 235000013305 food Nutrition 0.000 description 4
- 239000012634 fragment Substances 0.000 description 4
- 229910052757 nitrogen Inorganic materials 0.000 description 4
- 241000207746 Nicotiana benthamiana Species 0.000 description 3
- 244000061176 Nicotiana tabacum Species 0.000 description 3
- 238000004458 analytical method Methods 0.000 description 3
- 230000001580 bacterial effect Effects 0.000 description 3
- 235000013355 food flavoring agent Nutrition 0.000 description 3
- 239000007789 gas Substances 0.000 description 3
- 239000000203 mixture Substances 0.000 description 3
- 238000009395 breeding Methods 0.000 description 2
- 230000001488 breeding effect Effects 0.000 description 2
- 239000003795 chemical substances by application Substances 0.000 description 2
- 239000002537 cosmetic Substances 0.000 description 2
- 238000001514 detection method Methods 0.000 description 2
- 239000000835 fiber Substances 0.000 description 2
- 230000008014 freezing Effects 0.000 description 2
- 238000007710 freezing Methods 0.000 description 2
- 239000011521 glass Substances 0.000 description 2
- 238000004519 manufacturing process Methods 0.000 description 2
- 239000000463 material Substances 0.000 description 2
- 230000001404 mediated effect Effects 0.000 description 2
- 239000002304 perfume Substances 0.000 description 2
- 239000013612 plasmid Substances 0.000 description 2
- 238000002470 solid-phase micro-extraction Methods 0.000 description 2
- 239000000243 solution Substances 0.000 description 2
- 210000001519 tissue Anatomy 0.000 description 2
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 2
- FPIPGXGPPPQFEQ-UHFFFAOYSA-N 13-cis retinol Natural products OCC=C(C)C=CC=C(C)C=CC1=C(C)CCCC1(C)C FPIPGXGPPPQFEQ-UHFFFAOYSA-N 0.000 description 1
- 241000167854 Bourreria succulenta Species 0.000 description 1
- 244000025254 Cannabis sativa Species 0.000 description 1
- OKTJSMMVPCPJKN-UHFFFAOYSA-N Carbon Chemical compound [C] OKTJSMMVPCPJKN-UHFFFAOYSA-N 0.000 description 1
- 208000002177 Cataract Diseases 0.000 description 1
- 241000207199 Citrus Species 0.000 description 1
- 240000008067 Cucumis sativus Species 0.000 description 1
- 235000010799 Cucumis sativus var sativus Nutrition 0.000 description 1
- 108020004414 DNA Proteins 0.000 description 1
- 244000000626 Daucus carota Species 0.000 description 1
- 235000002767 Daucus carota Nutrition 0.000 description 1
- 241001523681 Dendrobium Species 0.000 description 1
- 240000001008 Dimocarpus longan Species 0.000 description 1
- 235000000235 Euphoria longan Nutrition 0.000 description 1
- 229930182566 Gentamicin Natural products 0.000 description 1
- CEAZRRDELHUEMR-URQXQFDESA-N Gentamicin Chemical compound O1[C@H](C(C)NC)CC[C@@H](N)[C@H]1O[C@H]1[C@H](O)[C@@H](O[C@@H]2[C@@H]([C@@H](NC)[C@@](C)(O)CO2)O)[C@H](N)C[C@@H]1N CEAZRRDELHUEMR-URQXQFDESA-N 0.000 description 1
- 101150075239 L1 gene Proteins 0.000 description 1
- 101150007425 L4 gene Proteins 0.000 description 1
- 241000209510 Liliopsida Species 0.000 description 1
- 108060001084 Luciferase Proteins 0.000 description 1
- 235000007688 Lycopersicon esculentum Nutrition 0.000 description 1
- 240000004658 Medicago sativa Species 0.000 description 1
- 235000017587 Medicago sativa ssp. sativa Nutrition 0.000 description 1
- 206010028980 Neoplasm Diseases 0.000 description 1
- 241000233855 Orchidaceae Species 0.000 description 1
- 240000007594 Oryza sativa Species 0.000 description 1
- 235000007164 Oryza sativa Nutrition 0.000 description 1
- ISWSIDIOOBJBQZ-UHFFFAOYSA-N Phenol Chemical compound OC1=CC=CC=C1 ISWSIDIOOBJBQZ-UHFFFAOYSA-N 0.000 description 1
- 241000219000 Populus Species 0.000 description 1
- 241001092459 Rubus Species 0.000 description 1
- 235000017848 Rubus fruticosus Nutrition 0.000 description 1
- 235000011034 Rubus glaucus Nutrition 0.000 description 1
- 244000235659 Rubus idaeus Species 0.000 description 1
- 235000009122 Rubus idaeus Nutrition 0.000 description 1
- 240000003768 Solanum lycopersicum Species 0.000 description 1
- 235000021307 Triticum Nutrition 0.000 description 1
- 244000098338 Triticum aestivum Species 0.000 description 1
- FPIPGXGPPPQFEQ-BOOMUCAASA-N Vitamin A Natural products OC/C=C(/C)\C=C\C=C(\C)/C=C/C1=C(C)CCCC1(C)C FPIPGXGPPPQFEQ-BOOMUCAASA-N 0.000 description 1
- 240000008042 Zea mays Species 0.000 description 1
- 235000005824 Zea mays ssp. parviglumis Nutrition 0.000 description 1
- 235000002017 Zea mays subsp mays Nutrition 0.000 description 1
- 229930013930 alkaloid Natural products 0.000 description 1
- 150000003797 alkaloid derivatives Chemical class 0.000 description 1
- FPIPGXGPPPQFEQ-OVSJKPMPSA-N all-trans-retinol Chemical compound OC\C=C(/C)\C=C\C=C(/C)\C=C\C1=C(C)CCCC1(C)C FPIPGXGPPPQFEQ-OVSJKPMPSA-N 0.000 description 1
- 230000004075 alteration Effects 0.000 description 1
- 230000009286 beneficial effect Effects 0.000 description 1
- 239000000440 bentonite Substances 0.000 description 1
- 229910000278 bentonite Inorganic materials 0.000 description 1
- SVPXDRXYRYOSEX-UHFFFAOYSA-N bentoquatam Chemical compound O.O=[Si]=O.O=[Al]O[Al]=O SVPXDRXYRYOSEX-UHFFFAOYSA-N 0.000 description 1
- 235000013361 beverage Nutrition 0.000 description 1
- 230000003115 biocidal effect Effects 0.000 description 1
- 238000010170 biological method Methods 0.000 description 1
- 235000021029 blackberry Nutrition 0.000 description 1
- 239000008280 blood Substances 0.000 description 1
- 210000004369 blood Anatomy 0.000 description 1
- 229910052799 carbon Inorganic materials 0.000 description 1
- 239000012159 carrier gas Substances 0.000 description 1
- 230000015556 catabolic process Effects 0.000 description 1
- 235000019693 cherries Nutrition 0.000 description 1
- 235000020971 citrus fruits Nutrition 0.000 description 1
- 238000010276 construction Methods 0.000 description 1
- 235000005822 corn Nutrition 0.000 description 1
- 238000012258 culturing Methods 0.000 description 1
- 238000006731 degradation reaction Methods 0.000 description 1
- 238000011161 development Methods 0.000 description 1
- 230000029087 digestion Effects 0.000 description 1
- 239000004205 dimethyl polysiloxane Substances 0.000 description 1
- 235000013870 dimethyl polysiloxane Nutrition 0.000 description 1
- 239000003814 drug Substances 0.000 description 1
- 230000007613 environmental effect Effects 0.000 description 1
- 239000000686 essence Substances 0.000 description 1
- 241001233957 eudicotyledons Species 0.000 description 1
- 229930003935 flavonoid Natural products 0.000 description 1
- 150000002215 flavonoids Chemical class 0.000 description 1
- 235000017173 flavonoids Nutrition 0.000 description 1
- 235000019634 flavors Nutrition 0.000 description 1
- 238000001502 gel electrophoresis Methods 0.000 description 1
- 238000010353 genetic engineering Methods 0.000 description 1
- 229960002518 gentamicin Drugs 0.000 description 1
- 150000004676 glycans Chemical class 0.000 description 1
- 239000001963 growth medium Substances 0.000 description 1
- 101150054900 gus gene Proteins 0.000 description 1
- 239000001307 helium Substances 0.000 description 1
- 229910052734 helium Inorganic materials 0.000 description 1
- SWQJXJOGLNCZEY-UHFFFAOYSA-N helium atom Chemical compound [He] SWQJXJOGLNCZEY-UHFFFAOYSA-N 0.000 description 1
- 230000001939 inductive effect Effects 0.000 description 1
- 208000015181 infectious disease Diseases 0.000 description 1
- 238000002347 injection Methods 0.000 description 1
- 239000007924 injection Substances 0.000 description 1
- 229930027917 kanamycin Natural products 0.000 description 1
- 229960000318 kanamycin Drugs 0.000 description 1
- SBUJHOSQTJFQJX-NOAMYHISSA-N kanamycin Chemical compound O[C@@H]1[C@@H](O)[C@H](O)[C@@H](CN)O[C@@H]1O[C@H]1[C@H](O)[C@@H](O[C@@H]2[C@@H]([C@@H](N)[C@H](O)[C@@H](CO)O2)O)[C@H](N)C[C@@H]1N SBUJHOSQTJFQJX-NOAMYHISSA-N 0.000 description 1
- 229930182823 kanamycin A Natural products 0.000 description 1
- 238000000520 microinjection Methods 0.000 description 1
- 238000012986 modification Methods 0.000 description 1
- 230000004048 modification Effects 0.000 description 1
- 210000005036 nerve Anatomy 0.000 description 1
- CXQXSVUQTKDNFP-UHFFFAOYSA-N octamethyltrisiloxane Chemical compound C[Si](C)(C)O[Si](C)(C)O[Si](C)(C)C CXQXSVUQTKDNFP-UHFFFAOYSA-N 0.000 description 1
- 230000002018 overexpression Effects 0.000 description 1
- 230000000144 pharmacologic effect Effects 0.000 description 1
- 238000004987 plasma desorption mass spectroscopy Methods 0.000 description 1
- 229920000435 poly(dimethylsiloxane) Polymers 0.000 description 1
- 229920001282 polysaccharide Polymers 0.000 description 1
- 239000005017 polysaccharide Substances 0.000 description 1
- 230000008569 process Effects 0.000 description 1
- 239000002994 raw material Substances 0.000 description 1
- 210000003705 ribosome Anatomy 0.000 description 1
- 235000009566 rice Nutrition 0.000 description 1
- 238000005070 sampling Methods 0.000 description 1
- 238000000926 separation method Methods 0.000 description 1
- 239000000126 substance Substances 0.000 description 1
- 238000010257 thawing Methods 0.000 description 1
- 230000005026 transcription initiation Effects 0.000 description 1
- 230000001131 transforming effect Effects 0.000 description 1
- 230000010474 transient expression Effects 0.000 description 1
- 238000003146 transient transfection Methods 0.000 description 1
- 239000013603 viral vector Substances 0.000 description 1
- 235000019155 vitamin A Nutrition 0.000 description 1
- 239000011719 vitamin A Substances 0.000 description 1
- 229940045997 vitamin a Drugs 0.000 description 1
- 239000003039 volatile agent Substances 0.000 description 1
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.)
- C12N9/0069—Oxidoreductases (1.) acting on single donors with incorporation of molecular oxygen, i.e. oxygenases (1.13)
-
- 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
- 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
- C12N15/79—Vectors or expression systems specially adapted for eukaryotic hosts
- C12N15/82—Vectors or expression systems specially adapted for eukaryotic hosts for plant cells, e.g. plant artificial chromosomes (PACs)
- C12N15/8241—Phenotypically and genetically modified plants via recombinant DNA technology
- C12N15/8242—Phenotypically and genetically modified plants via recombinant DNA technology with non-agronomic quality (output) traits, e.g. for industrial processing; Value added, non-agronomic traits
- C12N15/8243—Phenotypically and genetically modified plants via recombinant DNA technology with non-agronomic quality (output) traits, e.g. for industrial processing; Value added, non-agronomic traits involving biosynthetic or metabolic pathways, i.e. metabolic engineering, e.g. nicotine, caffeine
-
- 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
- C12P7/00—Preparation of oxygen-containing organic compounds
- C12P7/24—Preparation of oxygen-containing organic compounds containing a carbonyl group
- C12P7/26—Ketones
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12Y—ENZYMES
- C12Y113/00—Oxidoreductases acting on single donors with incorporation of molecular oxygen (oxygenases) (1.13)
- C12Y113/11—Oxidoreductases acting on single donors with incorporation of molecular oxygen (oxygenases) (1.13) with incorporation of two atoms of oxygen (1.13.11)
- C12Y113/11051—9-Cis-epoxycarotenoid dioxygenase (1.13.11.51)
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12R—INDEXING SCHEME ASSOCIATED WITH SUBCLASSES C12C - C12Q, RELATING TO MICROORGANISMS
- C12R2001/00—Microorganisms ; Processes using microorganisms
- C12R2001/01—Bacteria or Actinomycetales ; using bacteria or Actinomycetales
- C12R2001/185—Escherichia
- C12R2001/19—Escherichia coli
-
- Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
- Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
- Y02A—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE
- Y02A50/00—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE in human health protection, e.g. against extreme weather
- Y02A50/30—Against vector-borne diseases, e.g. mosquito-borne, fly-borne, tick-borne or waterborne diseases whose impact is exacerbated by climate change
Landscapes
- Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Genetics & Genomics (AREA)
- Chemical & Material Sciences (AREA)
- Engineering & Computer Science (AREA)
- Organic Chemistry (AREA)
- Zoology (AREA)
- Wood Science & Technology (AREA)
- Bioinformatics & Cheminformatics (AREA)
- Biotechnology (AREA)
- General Engineering & Computer Science (AREA)
- Biomedical Technology (AREA)
- Biochemistry (AREA)
- General Health & Medical Sciences (AREA)
- Microbiology (AREA)
- Molecular Biology (AREA)
- Plant Pathology (AREA)
- Physics & Mathematics (AREA)
- Biophysics (AREA)
- Nutrition Science (AREA)
- Cell Biology (AREA)
- Medicinal Chemistry (AREA)
- Chemical Kinetics & Catalysis (AREA)
- General Chemical & Material Sciences (AREA)
- Enzymes And Modification Thereof (AREA)
Abstract
本发明公开了一种铁皮石斛β‑紫罗兰酮合成关键酶基因DoCCD1及其克隆方法和应用,所述的铁皮石斛β‑紫罗兰酮合成关键酶基因具有如下核苷酸序列:(1)由SEQ ID NO:1所示的核苷酸序列;或(2)由序列SEQ ID NO:1所示的核苷酸序列同源性在70%以上且编码相同功能蛋白质的氨基酸序列;或(3)在高严谨条件下可与SEQ ID No:1限定的DNA序列杂交的核苷酸序列。本发明的CCD1基因能够通过降解胡萝卜素生成紫罗兰酮,为了提高铁皮石斛的经济价值,鉴定出具有真正功能的DoCCD1是非常有意义的工作。
Description
技术领域
本发明涉及遗传工程技术领域,具体涉及一种铁皮石斛β-紫罗兰酮合成关键酶基因DoCCD1及其克隆方法和应用。
背景技术
铁皮石斛(Dendrobium officinale)是兰科石斛属多年生附生植物,其茎既可作为食品,也是我国特有的名贵药材。《本草经》和《本草纲目》中均将铁皮石斛列入在内,多年来被誉为“中华九大仙草之首”,民间称其为“救命仙草”,有延年益寿之功效。铁皮石斛主要分布在我国南方省份。铁皮石斛是药用范围较广的中药,2020版《中华人民共和国药典》将其列为药用石斛的主要来源。其主要含有生物碱类、多糖类、黄酮类、酚类等多种化学成分,具有降血糖、改善记忆、保护神经、抗白内障、抗肿瘤等药理作用。此外,铁皮石斛还具有较高的观赏价值。
紫罗兰酮是一种高级香料,是调香中不可或缺的重要的原香料,用途广,需求量大。紫罗兰酮的分子式为C13H20O,根据双键位置的不同,可将紫罗兰酮划分为α体,β体和γ体3种同分异构体,而自然界中多以α体、β体这两种异构的混合形式存在,其中β-紫罗兰酮是用于高档级精细日用化妆品的调香剂和增香剂,也是医药工业上合成维生素A的重要原料。在食品行业中,紫罗兰酮常用作高档食品、饮料的增香剂,是中国GB2760-2011规定允许使用的食用香料,主要用以配制龙眼、树莓、黑莓、樱桃、柑橘等香精,全球年需求量达近万吨。由于国内目前的生产工艺限制,缺口超过1000吨,特别是医药级的,基本全靠进口。产品的市场潜力大,也急需高效的高产方法。
紫罗兰酮是铁皮石斛中最重要的挥发性成分,而这也是铁皮石斛具有重要美容及医用价值的基础。
目前,缺乏一种铁皮石斛β-紫罗兰酮合成关键酶基因DoCCD1及其克隆方法和应用。
发明内容
针对现有技术的问题,本发明的第一目的在于提供一种铁皮石斛β-紫罗兰酮合成关键酶基因DoCCD1;第二目的在于提供所述的铁皮石斛β-紫罗兰酮合成关键酶基因DoCCD1的应用。
本发明的第二目的是这样实现的,所述的铁皮石斛β-紫罗兰酮合成关键酶基因DoCCD1在获得高β-紫罗兰酮含量的瞬时转化烟草植株中的应用,即在烟草植株体内瞬时过表达DoCCD1基因,可提高烟草中β-紫罗兰酮的含量。
为达到上述目的,本发明采用了下列技术方案:本发明的一种铁皮石斛β-紫罗兰酮合成关键酶基因DoCCD1,所述的铁皮石斛β-紫罗兰酮合成关键酶基因具有如下核苷酸序列:
(1)由SEQ ID NO:1所示的核苷酸序列;或
(2)由序列SEQ ID NO:1所示的核苷酸序列同源性在70%以上且编码相同功能蛋白质的氨基酸序列;或
(3)在高严谨条件下可与SEQ ID No:1限定的DNA序列杂交的核苷酸序列。
本发明所述的铁皮石斛β-紫罗兰酮合成关键酶基因DoCCD1编码的多肽,所述多肽具有如下氨基酸序列:
(1)由SEQ ID No.2所示的氨基酸序列组成的蛋白质;或
(2)与序列SEQ ID No.2限定的氨基酸序列同源性在80%至100%编码相同功能蛋白质的氨基酸序列;或
(3)SEQ ID No.2所示的氨基酸序列经增加、缺失或替换一个或多个氨基酸具有同等活性的由(1)衍生的蛋白。
进一步地,一个或几个氨基酸残基的取代和/或缺失和/或添加是指不超过10个氨基酸残基的取代和/或缺失和/或添加。
本发明的一种含有所述的铁皮石斛β-紫罗兰酮合成关键酶基因DoCCD1的重组载体。
本发明的一种含有所述的铁皮石斛β-紫罗兰酮合成关键酶基因DoCCD1的表达盒。
本发明的一种含有所述的铁皮石斛β-紫罗兰酮合成关键酶基因DoCCD1的转基因细胞系或重组菌。
本发明的所述的铁皮石斛β-紫罗兰酮合成关键酶基因DoCCD1的克隆方法,包括如下步骤:
(1)铁皮石斛叶片cDNA合成:提取铁皮石斛叶片总RNA,反转录得到第一链cDNA;所述的引物序列的正向引物的氨基酸序列如SEQ ID:No.3所示;所述的引物序列的反向引物的氨基酸序列如SEQ ID:No.4所示;
(2)DoCCD1基因的PCR扩增:以铁皮石斛叶片cDNA为模板,根据DoCCD1基因序列设计引物,进行PCR扩增,回收和纯化PCR扩增产物,并测序。
进一步地,在步骤(2)中,所述的测序的具体操作是将扩增获得的PCR产物在0.8%的琼脂糖凝胶电泳,电泳结束后,采用Qiagen公司PCR产物纯化试剂盒,按照产品说明回收纯化所述的PCR产物,并送Invitrogen测序,验证序列结果。
本发明的一种所述的铁皮石斛β-紫罗兰酮合成关键酶基因DoCCD1在获得高β-紫罗兰酮烟草瞬时转化植株中的应用。
本发明的一种铁皮石斛β-紫罗兰酮合成关键酶基因DoCCD1的应用为在含有胡萝卜素的大肠杆菌中表达该基因,可获得β-紫罗兰酮。
有益效果:本发明的CCD1基因能够通过降解胡萝卜素生成紫罗兰酮,为了提高铁皮石斛的经济价值,鉴定出具有真正功能的DoCCD1是非常有意义的工作。
与现有技术相比,本发明具有如下优点:DoCCD1基因在构建到植物表达载体中时,在其转录起始核苷酸前可加上任何一种增强启动子或诱导型启动子。携带有本发明Ribosomal L4/L1基因的表达载体可通过使用Ti质粒、Ri质粒、植物病毒载体、直接DNA转化、显微注射、电导、农杆菌介导等常规生物学方法转化植物细胞或组织,并将转化的植物经组织培育成植株。
附图说明
图1为本发明的铁皮石斛叶片中β-紫罗兰酮含量图;
图2为本发明的DoCCD1的PCR产物电泳图;图中,M-分子量标记;1-PCR产物;
图3为本发明的DoCCD1基因植物瞬时表达载体图;
图4为本发明的瞬时转化烟草中β-紫罗兰酮含量图;图中,本氏烟草野生型对照;DoCCD1为瞬时转化烟株;
图5为本发明的DoCCD1基因植物转化大肠杆菌表达载体图;
图6为本发明的含胡萝卜素大肠杆菌中转化DoCCD1基因后,胡萝卜素发生降解图;图中,BL21-野生型对照;DoCCD1为转基因大肠杆菌。
图7为本发明的含胡萝卜素大肠杆菌中转化DoCCD1基因后,β-紫罗兰酮含量图;图中,Control为BL21野生型对照;DoCCD1为转基因大肠杆菌。
具体实施方式
下面结合实施例和附图对本发明作进一步的说明,但不以任何方式对本发明加以限制,基于本发明教导所作的任何变换或替换,均属于本发明的保护范围。
本发明的一种铁皮石斛β-紫罗兰酮合成关键酶基因DoCCD1,所述的铁皮石斛β-紫罗兰酮合成关键酶基因具有如下核苷酸序列:
(1)由SEQ ID NO:1所示的核苷酸序列;或
(2)由序列SEQ ID NO:1所示的核苷酸序列同源性在70%以上且编码相同功能蛋白质的氨基酸序列;或
(3)在高严谨条件下可与SEQ ID No:1限定的DNA序列杂交的核苷酸序列。
本发明所述的铁皮石斛β-紫罗兰酮合成关键酶基因DoCCD1编码的多肽,所述多肽具有如下氨基酸序列:
(1)由SEQ ID No.2所示的氨基酸序列组成的蛋白质;或
(2)与序列SEQ ID No.2限定的氨基酸序列同源性在80%至100%编码相同功能蛋白质的氨基酸序列;或
(3)SEQ ID No.2所示的氨基酸序列经增加、缺失或替换一个或多个氨基酸具有同等活性的由(1)衍生的蛋白。一个或几个氨基酸残基的取代和/或缺失和/或添加是指不超过10个氨基酸残基的取代和/或缺失和/或添加。
本发明的一种含有所述的铁皮石斛β-紫罗兰酮合成关键酶基因DoCCD1的重组载体。
本发明的一种含有所述的铁皮石斛β-紫罗兰酮合成关键酶基因DoCCD1的表达盒。
本发明的一种含有所述的铁皮石斛β-紫罗兰酮合成关键酶基因DoCCD1的转基因细胞系或重组菌。
本发明的一种所述的铁皮石斛β-紫罗兰酮合成关键酶基因DoCCD1在获得高β-紫罗兰酮烟草瞬时转化植株中的应用。
实施例中植物组织材料取自铁皮石斛和烟草植株的生长和发育阶段都是在人工气候室中,并保持生长温度在22-25℃之间,以尽量减少外界环境因素对β-紫罗兰酮合成过程中的影响。
实施例1
检测铁皮石斛中β-紫罗兰酮含量
将铁皮石斛叶片用液氮磨碎,称取1g样品放入玻璃瓶中,将SPME纤维(95μm×10mmCarbon WR/PDMS,CTC Analytics AG,Switzerland)通过隔膜引入小瓶中,40℃下将顶部空间挥发物吸附30分钟。将SPME纤维引入气相色谱仪并进行分析。分析仪器为Agilent7890-5975气相色谱质谱仪;分离色谱柱为DB-5MS气相色谱柱(30m×0.25mm×0.25μm),载气(氦气)流速1.1mL/min,进样口温度为250℃,不分流进样。温度程序为40℃持续1分钟,2℃/min升至60℃,10℃/min升至325℃。离子源温度为250℃。扫描范围:m/z 33-500。
实施例2
克隆DOCCD1基因
本发明的所述的铁皮石斛β-紫罗兰酮合成关键酶基因DoCCD1的克隆方法,包括如下步骤:
(1)铁皮石斛叶片cDNA合成:提取铁皮石斛叶片总RNA,反转录得到第一链cDNA;所述的引物序列的正向引物的氨基酸序列如SEQ ID:No.3所示;所述的引物序列的反向引物的氨基酸序列如SEQ ID:No.4所示;
(2)DoCCD1基因的PCR扩增:以铁皮石斛叶片cDNA为模板,根据DoCCD1基因序列设计引物,进行PCR扩增,回收和纯化PCR扩增产物,并测序。
以铁皮石斛叶片cDNA为模板,根据数据库信息设计引物,进行DoCCD1基因的PCR扩增,得到PCR扩增产物。设计引物如下所示:
正向引物:5’-ATGGAGAGAGATTTTATGGGTTTGAC-3’
反向引物:5’-TCACCTTGTGTCTACATTATTTTGCC-3’。
PCR反应体系和扩增条件如表1所示。
表1
将扩增获得的PCR产物在0.8%的琼脂糖凝胶电泳,凝胶电泳结果如图1所示。电泳结束后,采用Qiagen公司PCR产物纯化试剂盒,按照产品说明回收纯化所述的PCR产物,并送Invitrogen测序,验证序列结果。
实施例3
植物DoCCD1基因瞬时转化载体的构建
以实施例2中DoCCD1全长片段为模板,用含有接头序列的引物进行PCR扩增,扩增产物经PCR产物纯化后,BsaI酶切载体,回收载体骨架片段。经过诺唯赞IIOne Step Cloning Kit重组反应插入到pHREAC载体(图3)中。
(1)反应引物序列如下:
DoCCD1_F:5’-taaacgtctctaaaaATGGGGGAATTGGCAGCG-3’;
DoCCD1_R:5’-aatgaaaccagagcgTCATATTTCTGATTGCTGCTGCTG-3’。
实施例4
农杆菌介导的烟草瞬时转化
(1)冻融法转化农杆菌
将1μg(200ng/μL)的pHREAC重组载体加入到100μL感受态农杆菌GV3101中,混匀后在冰上静置5min,放入液氮中冷冻5min,然后从液氮中取出,放入37℃水浴锅中水浴5min,再在冰上静置5min后,加入500μL LB溶液,在28℃、充分震荡条件下恢复培养4h,最后将菌液均匀涂抹于选择性平板培养基上,28℃下培养48h。
(2)瞬时转化本氏烟。
具体方法如下:将载体分别转入农杆菌GV3101中,注射约50天大的本氏烟草(Nicotiana benthamiana)叶片,侵染三天后采样,经液氮速冻。用于后期分析。
(3)烟草叶片中β-紫罗兰酮检测
烟草瞬时转染叶片中β-紫罗兰酮含量的检测方法同实施例1.可以观测到瞬时转化DoCCD1的本氏烟叶片中β-紫罗兰酮含量显著增加。
实施例5
DoCCD1能够催化胡萝卜素生成β-紫罗兰酮
将pACCAR16△crt载体转换到BL21-AI菌株中,此时大肠杆菌能够生成胡萝卜素。
以实施例2中DoCCD1全长片段为模板,用含有接头序列的引物进行PCR扩增,扩增产物经PCR产物纯化后,Xhol和XbaI双酶切载体,回收载体骨架片段。经过诺唯赞II One Step Cloning Kit重组反应插入到pThio_Dan1载体(图5)中。
(1)反应引物序列如下:
DoCCD1_F:5’-gcccttggcgaattcctcgagATGGGGGAATTGGCAGCG-3’;
DoCCD1_R:5’-tgcctgcaggtcgactctagaTATTTCTGATTGCTGCTGCTGC-3’。
将构建好的pThio_Dan1载体转化含pACCAR16△crt载体的BL21-AI菌株。可以观测到含有DoCCD1的大肠杆菌变成白色,而没有转化DoCCD1的大肠杆菌呈现胡萝卜色(图6)
之后进行摇菌,提取5ml菌液放入玻璃瓶。之后按照实施例1中的方法测定菌液中紫罗兰酮含量。可以观察到转化DoCCD1的大肠杆菌中β-紫罗兰酮含量显著增加(图7),表明DoCCD1能够催化胡萝卜素生成β-紫罗兰酮。
为了便于对转基因植物细胞或植物进行鉴定及筛选,可对所使用的载体进行加工,如加入植物可选择性标记(GUS基因、荧光素酶基因等)或具有抗性的抗生素标记物(庆大霉素,卡那霉素等)。被转化的植物宿主既可以是单子叶植物,也可以是双子叶植物,如:烟草、水稻、小麦、玉米、黄瓜、番茄、杨树、草坪草或苜蓿等。
本发明的β-紫罗兰酮合成关键酶相关蛋白及其编码基因为农作物尤其是β-紫罗兰酮含量育种提供基因与技术的支持。
以上显示和描述了本发明的基本原理、主要特征和本发明的优点。本行业的技术人员应该了解,本发明不受上述实施例的限制,上述实施例和说明书中描述的只是说明本发明的原理,在不脱离本发明精神和范围的前提下,本发明还会有各种变化和改进,本发明要求保护范围由所附的权利要求书、说明书及其等效物界定。
Claims (10)
1.一种铁皮石斛β-紫罗兰酮合成关键酶基因DoCCD1,其特征在于:所述的铁皮石斛β-紫罗兰酮合成关键酶基因具有如下核苷酸序列:
(1)由SEQ ID NO:1所示的核苷酸序列;或
(2)由序列SEQ ID NO:1所示的核苷酸序列同源性在70%以上且编码相同功能蛋白质的氨基酸序列;或
(3)在高严谨条件下可与SEQ ID No:1限定的DNA序列杂交的核苷酸序列。
2.权利要求1所述的铁皮石斛β-紫罗兰酮合成关键酶基因DoCCD1编码的多肽,其特征在于:所述多肽具有如下氨基酸序列:
(1)由SEQ ID No.2所示的氨基酸序列组成的蛋白质;或
(2)与序列SEQ ID No.2限定的氨基酸序列同源性在80%至100%编码相同功能蛋白质的氨基酸序列;或
(3)SEQ ID No.2所示的氨基酸序列经增加、缺失或替换一个或多个氨基酸具有同等活性的由(1)衍生的蛋白。
3.根据权利要求2所述的铁皮石斛β-紫罗兰酮合成关键酶基因DoCCD1编码的多肽,其特征在于:一个或几个氨基酸残基的取代和/或缺失和/或添加是指不超过10个氨基酸残基的取代和/或缺失和/或添加。
4.一种含有权利要求1或2所述的铁皮石斛β-紫罗兰酮合成关键酶基因DoCCD1的重组载体。
5.一种含有权利要求1或2所述的铁皮石斛β-紫罗兰酮合成关键酶基因DoCCD1的表达盒。
6.一种含有权利要求1或2所述的铁皮石斛β-紫罗兰酮合成关键酶基因DoCCD1的转基因细胞系或重组菌。
7.权利要求1或2所述的铁皮石斛β-紫罗兰酮合成关键酶基因DoCCD1的克隆方法,其特征在于包括如下步骤:
(1)铁皮石斛叶片cDNA合成:提取铁皮石斛叶片总RNA,反转录得到第一链cDNA;所述的引物序列的正向引物的氨基酸序列如SEQ ID:No.3所示;所述的引物序列的反向引物的氨基酸序列如SEQ ID:No.4所示;
(2)DoCCD1基因的PCR扩增:以铁皮石斛叶片cDNA为模板,根据DoCCD1基因序列设计引物,进行PCR扩增,回收和纯化PCR扩增产物,并测序。
8.根据权利要求7所述的铁皮石斛β-紫罗兰酮合成关键酶基因DoCCD1的克隆方法,其特征在于:在步骤(2)中,所述的测序的具体操作是将扩增获得的PCR产物在0.8%的琼脂糖凝胶电泳,电泳结束后,采用Qiagen公司PCR产物纯化试剂盒,按照产品说明回收纯化所述的PCR产物,并送Invitrogen测序,验证序列结果。
9.一种权利要求1或2所述的铁皮石斛β-紫罗兰酮合成关键酶基因DoCCD1在获得高β-紫罗兰酮烟草瞬时转化植株中的应用。
10.一种权利要求1或2铁皮石斛β-紫罗兰酮合成关键酶基因DoCCD1的应用为在含有胡萝卜素的大肠杆菌中表达该基因,可获得β-紫罗兰酮。
Applications Claiming Priority (2)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
CN2022107955658 | 2022-07-06 | ||
CN202210795565 | 2022-07-06 |
Publications (1)
Publication Number | Publication Date |
---|---|
CN116042663A true CN116042663A (zh) | 2023-05-02 |
Family
ID=86116928
Family Applications (1)
Application Number | Title | Priority Date | Filing Date |
---|---|---|---|
CN202210915168.XA Pending CN116042663A (zh) | 2022-07-06 | 2022-08-01 | 一种铁皮石斛β-紫罗兰酮合成关键酶基因DoCCD1及其克隆方法和应用 |
Country Status (1)
Country | Link |
---|---|
CN (1) | CN116042663A (zh) |
Citations (6)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
WO2005118812A1 (ja) * | 2004-06-04 | 2005-12-15 | Marine Biotechnology Institute Co., Ltd. | カロテノイドケトラーゼ及びカロテノイドヒドロキシラーゼ遺伝子を利用したアスタキサンチンまたはその代謝物の製造法 |
CN105368850A (zh) * | 2015-11-25 | 2016-03-02 | 天津大学 | 产生β-紫罗酮香气物质的枸杞类胡萝卜素裂解双加氧酶基因及应用 |
CN106282137A (zh) * | 2016-08-02 | 2017-01-04 | 郑州轻工业学院 | 一种类胡萝卜素9, 10’双加氧酶的制备方法及其应用 |
CN108949599A (zh) * | 2018-06-29 | 2018-12-07 | 华南理工大学 | 一种产β-紫罗兰酮基因工程菌及其构建方法与应用 |
CN113088498A (zh) * | 2020-01-08 | 2021-07-09 | 中国医学科学院药用植物研究所 | 一种参与挥发性成分形成的类胡萝卜素裂解双加氧酶编码基因的筛选鉴定及应用 |
CN113512556A (zh) * | 2021-06-11 | 2021-10-19 | 南京大学 | 一种与β-紫罗兰酮合成相关的类胡萝卜素裂解双加氧酶基因及其编码蛋白和应用 |
-
2022
- 2022-08-01 CN CN202210915168.XA patent/CN116042663A/zh active Pending
Patent Citations (6)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
WO2005118812A1 (ja) * | 2004-06-04 | 2005-12-15 | Marine Biotechnology Institute Co., Ltd. | カロテノイドケトラーゼ及びカロテノイドヒドロキシラーゼ遺伝子を利用したアスタキサンチンまたはその代謝物の製造法 |
CN105368850A (zh) * | 2015-11-25 | 2016-03-02 | 天津大学 | 产生β-紫罗酮香气物质的枸杞类胡萝卜素裂解双加氧酶基因及应用 |
CN106282137A (zh) * | 2016-08-02 | 2017-01-04 | 郑州轻工业学院 | 一种类胡萝卜素9, 10’双加氧酶的制备方法及其应用 |
CN108949599A (zh) * | 2018-06-29 | 2018-12-07 | 华南理工大学 | 一种产β-紫罗兰酮基因工程菌及其构建方法与应用 |
CN113088498A (zh) * | 2020-01-08 | 2021-07-09 | 中国医学科学院药用植物研究所 | 一种参与挥发性成分形成的类胡萝卜素裂解双加氧酶编码基因的筛选鉴定及应用 |
CN113512556A (zh) * | 2021-06-11 | 2021-10-19 | 南京大学 | 一种与β-紫罗兰酮合成相关的类胡萝卜素裂解双加氧酶基因及其编码蛋白和应用 |
Non-Patent Citations (3)
Title |
---|
ZHAO CONGHUI 等: "Functional Characterization of a Dendrobium officinale Geraniol Synthase DoGES1 Involved in Floral Scent Formation", 《INTERNATIONAL JOURNAL OF MOLECULAR SCIENCES》, vol. 21, no. 19, 23 September 2020 (2020-09-23) * |
无: "Genbank:XM_020824279.2", 《GENBANK》, 21 April 2019 (2019-04-21) * |
魏涛 等: "霍氏肠杆菌β-胡萝卜素9, 10\'双加氧酶的制备及其应用", 《中国酿造》, vol. 41, no. 1, 25 January 2022 (2022-01-25), pages 128 * |
Similar Documents
Publication | Publication Date | Title |
---|---|---|
Chen et al. | Root‐specific expression of opine genes and opine accumulation in some cultivars of the naturally occurring genetically modified organism Nicotiana tabacum | |
CN108048415B (zh) | 两个杨梅黄酮醇合成酶MrFLSs蛋白及其编码基因的应用 | |
CN107955067B (zh) | 参与桃黄酮醇生物合成调控的两个myb转录因子及其应用 | |
CN113416748A (zh) | 一种合成大麻二酚的表达载体、异源表达方法及应用 | |
CN111235046A (zh) | 异源合成α-檀香烯的重组解脂耶氏酵母及其构建方法 | |
CN106220719B (zh) | 一种青蒿bHLH类转录因子编码序列及克隆方法与应用 | |
CN111518818A (zh) | 一个参与杨梅素生物合成的羟化酶基因及其应用 | |
CN106047895B (zh) | 一种青蒿bZIP类转录因子编码序列及克隆方法与应用 | |
CN110484547B (zh) | 桃多胺氧化酶PpPAO1基因、其编码蛋白和应用 | |
CN109402080B (zh) | 蛋白质ugt142及其编码基因与应用 | |
CN115896137A (zh) | 一种南果梨PuAAT基因、其过表达载体及应用 | |
CN116042663A (zh) | 一种铁皮石斛β-紫罗兰酮合成关键酶基因DoCCD1及其克隆方法和应用 | |
CN113025594B (zh) | 多肽、核酸及其在合成香叶醇上的应用 | |
CN106047897B (zh) | 一种青蒿bHLH类转录因子编码序列及克隆方法与应用 | |
CN115992109A (zh) | 石吊兰素糖基转移酶蛋白及其编码基因与应用 | |
CN114480326B (zh) | 亚精胺衍生物糖基转移酶LbUGT73及其编码基因和应用 | |
CN116790545A (zh) | 用于重楼皂苷生物合成的糖基转移酶PpUGT2 | |
CN109810965B (zh) | 一种源于知母的β-葡萄糖苷酶、其编码基因、表达载体及其应用 | |
CN114507657B (zh) | 一种菊花脑单萜合酶基因CnTPS2及其应用 | |
CN106086038B (zh) | 一种青蒿wrky类转录因子编码序列及克隆方法与应用 | |
CN115927406B (zh) | 一种南果梨PuAOX1a基因、过表达载体及应用 | |
CN115976068B (zh) | 提高雪莲绿原酸含量的SiHQT基因及其编码产物和应用 | |
KR102551064B1 (ko) | 포도에서 분리된 신규 u6 프로모터 및 이의 용도 | |
CN116606832A (zh) | 双苄基丁烷型木脂素氧甲基转移酶及其应用 | |
CN116445443A (zh) | 联苯环辛烯型木脂素氧甲基转移酶及其应用 |
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 |