CN108070597B - 一种杨树nac基因启动子及其应用 - Google Patents
一种杨树nac基因启动子及其应用 Download PDFInfo
- Publication number
- CN108070597B CN108070597B CN201810146879.9A CN201810146879A CN108070597B CN 108070597 B CN108070597 B CN 108070597B CN 201810146879 A CN201810146879 A CN 201810146879A CN 108070597 B CN108070597 B CN 108070597B
- Authority
- CN
- China
- Prior art keywords
- poplar
- dna
- plant
- promoter
- sequence
- 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
- 101150077549 nac gene Proteins 0.000 title abstract description 5
- 241000196324 Embryophyta Species 0.000 claims abstract description 37
- 108020004414 DNA Proteins 0.000 claims abstract description 36
- 241000219000 Populus Species 0.000 claims abstract description 30
- 239000012634 fragment Substances 0.000 claims abstract description 20
- 108090000623 proteins and genes Proteins 0.000 claims abstract description 13
- 102000053602 DNA Human genes 0.000 claims abstract description 9
- 230000014509 gene expression Effects 0.000 claims abstract description 9
- 108091026890 Coding region Proteins 0.000 claims abstract description 3
- 239000013598 vector Substances 0.000 claims description 12
- 230000009261 transgenic effect Effects 0.000 claims description 9
- 241000894006 Bacteria Species 0.000 claims description 3
- 238000012214 genetic breeding Methods 0.000 claims description 3
- 230000002792 vascular Effects 0.000 abstract description 6
- 108091028043 Nucleic acid sequence Proteins 0.000 abstract description 5
- 238000010353 genetic engineering Methods 0.000 abstract description 2
- 238000000034 method Methods 0.000 description 8
- 230000000694 effects Effects 0.000 description 6
- 239000007788 liquid Substances 0.000 description 6
- 238000010186 staining Methods 0.000 description 6
- 108091023040 Transcription factor Proteins 0.000 description 5
- 102000040945 Transcription factor Human genes 0.000 description 5
- 239000000706 filtrate Substances 0.000 description 5
- 239000000047 product Substances 0.000 description 5
- 238000013518 transcription Methods 0.000 description 5
- 230000035897 transcription Effects 0.000 description 5
- 241000589158 Agrobacterium Species 0.000 description 4
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 description 4
- 238000012408 PCR amplification Methods 0.000 description 4
- 230000001580 bacterial effect Effects 0.000 description 4
- 230000033228 biological regulation Effects 0.000 description 4
- 210000002421 cell wall Anatomy 0.000 description 4
- 238000006243 chemical reaction Methods 0.000 description 4
- 230000008569 process Effects 0.000 description 4
- 238000007400 DNA extraction Methods 0.000 description 3
- OKKJLVBELUTLKV-UHFFFAOYSA-N Methanol Chemical compound OC OKKJLVBELUTLKV-UHFFFAOYSA-N 0.000 description 3
- 230000015572 biosynthetic process Effects 0.000 description 3
- 239000001913 cellulose Substances 0.000 description 3
- 229920002678 cellulose Polymers 0.000 description 3
- 238000012258 culturing Methods 0.000 description 3
- 239000013604 expression vector Substances 0.000 description 3
- 239000001963 growth medium Substances 0.000 description 3
- 238000005286 illumination Methods 0.000 description 3
- 239000000463 material Substances 0.000 description 3
- 239000000203 mixture Substances 0.000 description 3
- 239000000843 powder Substances 0.000 description 3
- 230000001105 regulatory effect Effects 0.000 description 3
- 230000008844 regulatory mechanism Effects 0.000 description 3
- 238000011160 research Methods 0.000 description 3
- 230000034655 secondary growth Effects 0.000 description 3
- 239000007787 solid Substances 0.000 description 3
- 238000001179 sorption measurement Methods 0.000 description 3
- 229940027257 timentin Drugs 0.000 description 3
- 238000011144 upstream manufacturing Methods 0.000 description 3
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Chemical compound O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 3
- IJGRMHOSHXDMSA-UHFFFAOYSA-N Atomic nitrogen Chemical compound N#N IJGRMHOSHXDMSA-UHFFFAOYSA-N 0.000 description 2
- 239000002028 Biomass Substances 0.000 description 2
- 229920002488 Hemicellulose Polymers 0.000 description 2
- 241000095444 Populus davidiana Species 0.000 description 2
- 241000183024 Populus tremula Species 0.000 description 2
- 239000004098 Tetracycline Substances 0.000 description 2
- OJOBTAOGJIWAGB-UHFFFAOYSA-N acetosyringone Chemical compound COC1=CC(C(C)=O)=CC(OC)=C1O OJOBTAOGJIWAGB-UHFFFAOYSA-N 0.000 description 2
- 230000003321 amplification Effects 0.000 description 2
- 239000000872 buffer Substances 0.000 description 2
- 239000012881 co-culture medium Substances 0.000 description 2
- 238000010276 construction Methods 0.000 description 2
- 238000005520 cutting process Methods 0.000 description 2
- 238000013461 design Methods 0.000 description 2
- 238000001514 detection method Methods 0.000 description 2
- 238000010586 diagram Methods 0.000 description 2
- 235000013399 edible fruits Nutrition 0.000 description 2
- 238000002474 experimental method Methods 0.000 description 2
- 230000004927 fusion Effects 0.000 description 2
- 101150054900 gus gene Proteins 0.000 description 2
- 208000015181 infectious disease Diseases 0.000 description 2
- 229930027917 kanamycin Natural products 0.000 description 2
- 229960000318 kanamycin Drugs 0.000 description 2
- 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 2
- 229930182823 kanamycin A Natural products 0.000 description 2
- 229920005610 lignin Polymers 0.000 description 2
- 238000009630 liquid culture Methods 0.000 description 2
- 238000004519 manufacturing process Methods 0.000 description 2
- 239000003550 marker Substances 0.000 description 2
- 239000002609 medium Substances 0.000 description 2
- 239000013642 negative control Substances 0.000 description 2
- 238000003199 nucleic acid amplification method Methods 0.000 description 2
- 238000012257 pre-denaturation Methods 0.000 description 2
- 102000004169 proteins and genes Human genes 0.000 description 2
- JQXXHWHPUNPDRT-WLSIYKJHSA-N rifampicin Chemical compound O([C@](C1=O)(C)O/C=C/[C@@H]([C@H]([C@@H](OC(C)=O)[C@H](C)[C@H](O)[C@H](C)[C@@H](O)[C@@H](C)\C=C\C=C(C)/C(=O)NC=2C(O)=C3C([O-])=C4C)C)OC)C4=C1C3=C(O)C=2\C=N\N1CC[NH+](C)CC1 JQXXHWHPUNPDRT-WLSIYKJHSA-N 0.000 description 2
- 229960001225 rifampicin Drugs 0.000 description 2
- 238000012163 sequencing technique Methods 0.000 description 2
- 239000000243 solution Substances 0.000 description 2
- 230000001954 sterilising effect Effects 0.000 description 2
- 238000004659 sterilization and disinfection Methods 0.000 description 2
- 239000012085 test solution Substances 0.000 description 2
- 229960002180 tetracycline Drugs 0.000 description 2
- 229930101283 tetracycline Natural products 0.000 description 2
- 235000019364 tetracycline Nutrition 0.000 description 2
- 150000003522 tetracyclines Chemical class 0.000 description 2
- 230000008719 thickening Effects 0.000 description 2
- 239000002023 wood Substances 0.000 description 2
- JXCKZXHCJOVIAV-UHFFFAOYSA-N 6-[(5-bromo-4-chloro-1h-indol-3-yl)oxy]-3,4,5-trihydroxyoxane-2-carboxylic acid;cyclohexanamine Chemical compound [NH3+]C1CCCCC1.O1C(C([O-])=O)C(O)C(O)C(O)C1OC1=CNC2=CC=C(Br)C(Cl)=C12 JXCKZXHCJOVIAV-UHFFFAOYSA-N 0.000 description 1
- 229920001817 Agar Polymers 0.000 description 1
- 241000219194 Arabidopsis Species 0.000 description 1
- 229930192334 Auxin Natural products 0.000 description 1
- 108091062157 Cis-regulatory element Proteins 0.000 description 1
- 230000004568 DNA-binding Effects 0.000 description 1
- 102000016928 DNA-directed DNA polymerase Human genes 0.000 description 1
- 108010014303 DNA-directed DNA polymerase Proteins 0.000 description 1
- 241000759904 Davidia Species 0.000 description 1
- 102000016911 Deoxyribonucleases Human genes 0.000 description 1
- 108010053770 Deoxyribonucleases Proteins 0.000 description 1
- ZGTMUACCHSMWAC-UHFFFAOYSA-L EDTA disodium salt (anhydrous) Chemical compound [Na+].[Na+].OC(=O)CN(CC([O-])=O)CCN(CC(O)=O)CC([O-])=O ZGTMUACCHSMWAC-UHFFFAOYSA-L 0.000 description 1
- 108090000790 Enzymes Proteins 0.000 description 1
- 102000004190 Enzymes Human genes 0.000 description 1
- 108700039691 Genetic Promoter Regions Proteins 0.000 description 1
- 241000282414 Homo sapiens Species 0.000 description 1
- 108700001094 Plant Genes Proteins 0.000 description 1
- 241000249899 Populus tomentosa Species 0.000 description 1
- 241000218976 Populus trichocarpa Species 0.000 description 1
- 102000006382 Ribonucleases Human genes 0.000 description 1
- 108010083644 Ribonucleases Proteins 0.000 description 1
- 241001052560 Thallis Species 0.000 description 1
- 229920004890 Triton X-100 Polymers 0.000 description 1
- 239000013504 Triton X-100 Substances 0.000 description 1
- 230000009471 action Effects 0.000 description 1
- 230000004913 activation Effects 0.000 description 1
- 239000008272 agar Substances 0.000 description 1
- 238000000246 agarose gel electrophoresis Methods 0.000 description 1
- 238000004458 analytical method Methods 0.000 description 1
- 239000002363 auxin Substances 0.000 description 1
- 210000004027 cell Anatomy 0.000 description 1
- 238000005119 centrifugation Methods 0.000 description 1
- 239000007795 chemical reaction product Substances 0.000 description 1
- 239000003153 chemical reaction reagent Substances 0.000 description 1
- 238000005352 clarification Methods 0.000 description 1
- 238000010924 continuous production Methods 0.000 description 1
- 238000011161 development Methods 0.000 description 1
- 230000004069 differentiation Effects 0.000 description 1
- 239000012153 distilled water Substances 0.000 description 1
- 239000012154 double-distilled water Substances 0.000 description 1
- 230000007613 environmental effect Effects 0.000 description 1
- 238000001976 enzyme digestion Methods 0.000 description 1
- 239000000446 fuel Substances 0.000 description 1
- 238000012215 gene cloning Methods 0.000 description 1
- PCHJSUWPFVWCPO-UHFFFAOYSA-N gold Chemical compound [Au] PCHJSUWPFVWCPO-UHFFFAOYSA-N 0.000 description 1
- 239000010931 gold Substances 0.000 description 1
- 229910052737 gold Inorganic materials 0.000 description 1
- 230000012010 growth Effects 0.000 description 1
- 230000001744 histochemical effect Effects 0.000 description 1
- SEOVTRFCIGRIMH-UHFFFAOYSA-N indole-3-acetic acid Chemical compound C1=CC=C2C(CC(=O)O)=CNC2=C1 SEOVTRFCIGRIMH-UHFFFAOYSA-N 0.000 description 1
- 238000002955 isolation Methods 0.000 description 1
- 239000002029 lignocellulosic biomass Substances 0.000 description 1
- 239000006166 lysate Substances 0.000 description 1
- 230000007246 mechanism Effects 0.000 description 1
- 239000012528 membrane Substances 0.000 description 1
- 238000002156 mixing Methods 0.000 description 1
- 239000004570 mortar (masonry) Substances 0.000 description 1
- 229910052757 nitrogen Inorganic materials 0.000 description 1
- 239000002773 nucleotide Substances 0.000 description 1
- 125000003729 nucleotide group Chemical group 0.000 description 1
- 239000013612 plasmid Substances 0.000 description 1
- 239000013641 positive control Substances 0.000 description 1
- 238000002360 preparation method Methods 0.000 description 1
- 238000004537 pulping Methods 0.000 description 1
- 239000002994 raw material Substances 0.000 description 1
- 238000003259 recombinant expression Methods 0.000 description 1
- 230000022532 regulation of transcription, DNA-dependent Effects 0.000 description 1
- 108091008146 restriction endonucleases Proteins 0.000 description 1
- 239000012882 rooting medium Substances 0.000 description 1
- 230000006574 secondary cell wall biogenesis Effects 0.000 description 1
- 239000012064 sodium phosphate buffer Substances 0.000 description 1
- 239000008223 sterile water Substances 0.000 description 1
- 239000006228 supernatant Substances 0.000 description 1
- 239000000725 suspension Substances 0.000 description 1
- 238000003786 synthesis reaction Methods 0.000 description 1
- 239000004753 textile Substances 0.000 description 1
- 230000002103 transcriptional effect Effects 0.000 description 1
- 238000012546 transfer Methods 0.000 description 1
- 238000003260 vortexing Methods 0.000 description 1
- 238000005406 washing Methods 0.000 description 1
Images
Classifications
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
- C07K14/415—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from plants
-
- 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)
- C12N15/8216—Methods for controlling, regulating or enhancing expression of transgenes in plant cells
- C12N15/8222—Developmentally regulated expression systems, tissue, organ specific, temporal or spatial regulation
- C12N15/8223—Vegetative tissue-specific promoters
- C12N15/8225—Leaf-specific, e.g. including petioles, stomata
-
- 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)
- C12N15/8216—Methods for controlling, regulating or enhancing expression of transgenes in plant cells
- C12N15/8222—Developmentally regulated expression systems, tissue, organ specific, temporal or spatial regulation
- C12N15/8223—Vegetative tissue-specific promoters
- C12N15/8226—Stem-specific, e.g. including tubers, beets
-
- 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)
- C12N15/8216—Methods for controlling, regulating or enhancing expression of transgenes in plant cells
- C12N15/8222—Developmentally regulated expression systems, tissue, organ specific, temporal or spatial regulation
- C12N15/8223—Vegetative tissue-specific promoters
- C12N15/8227—Root-specific
Landscapes
- Health & Medical Sciences (AREA)
- Genetics & Genomics (AREA)
- Life Sciences & Earth Sciences (AREA)
- Engineering & Computer Science (AREA)
- Biomedical Technology (AREA)
- Chemical & Material Sciences (AREA)
- Organic Chemistry (AREA)
- Bioinformatics & Cheminformatics (AREA)
- Biotechnology (AREA)
- General Engineering & Computer Science (AREA)
- Zoology (AREA)
- Wood Science & Technology (AREA)
- General Health & Medical Sciences (AREA)
- Molecular Biology (AREA)
- Biophysics (AREA)
- Biochemistry (AREA)
- Microbiology (AREA)
- Plant Pathology (AREA)
- Physics & Mathematics (AREA)
- Cell Biology (AREA)
- Botany (AREA)
- Gastroenterology & Hepatology (AREA)
- Medicinal Chemistry (AREA)
- Proteomics, Peptides & Aminoacids (AREA)
- Micro-Organisms Or Cultivation Processes Thereof (AREA)
Abstract
本发明公开了一种杨树NAC基因启动子及其应用。本发明提供了DNA片段，为如下1)‑3)中任一所述的DNA分子：1)编码区为序列表中序列1所示的DNA分子；2)在严格条件下与1)限定的DNA序列杂交且具有启动子功能的DNA分子；3)与1)限定的DNA序列至少具有70％、至少具有75％、至少具有80％、至少具有85％、至少具有90％、至少具有95％、至少具有96％、至少具有97％、至少具有98％或至少具有99％同源性且具有启动子功能的DNA分子。应用本发明提供的启动子，可以在植物基因工程中，诱导相应目的基因在根、茎、叶维管组织中特异表达。
Description
技术领域
本发明属于生物技术领域，涉及一种杨树NAC基因启动子及其应用。
背景技术
木质纤维素是地球上最丰富、最廉价且符合可持续发展要求的可再生资源，利用它生产纤维素燃料乙醇，是世界生物能源产业的主流技术路线。木质纤维素类生物质包括木质素、纤维素和半纤维素，主要是木本植物、草本植物难以利用的次生细胞壁成分。
植物的次生生长是一项重要的生命活动。次生生长是指植物的初生生长结束后，由于次生分生组织，特别是维管形成层的活动，不断产生次生韧皮部和次生木质部，使细胞壁加厚、植物根和茎加粗的过程，主要指次生细胞壁的形成过程。次生木质部是木材的主要组成部分，主要由纤维素、木质素和半纤维素组成，是制浆造纸、建筑业以及纺织业的原材料，也是地球上最丰富的可再生资源，对人类的生产和生活起着重要作用。因此阐明植物次生细胞壁生物合成的分子生物学调控机理，对于提高木本植物生物质产量及提高木质纤维素生物质的转化效率至关重要。生长素和多个转录因子家族如MYB、NAC、AP2/EREBP、homeobox、MADS等参与了这一过程的调控。
转录因子是一类具有同真核基因启动区特异DNA序列(顺式元件)结合活性的蛋白质分子，或是具有DNA结合域及转录激活域等结构特征的蛋白质分子。高等植物自身生长发育和对环境变化响应是通过转录因子调控目的基因表达而实现的，转录因子在这些过程中起了“开关”的作用。
近几年的研究表明，转录调节是林木次生生长的核心，在木材形成中起着关键作用。模式植物拟南芥中的研究表明，植物次生细胞壁合成的转录调控涉及多基因参与的网络系统，NAC类转录因子广泛参与其中，而目前只有少数NAC蛋白上游或者下游基因得到鉴定，国内外有关NAC转录因子的研究成果多集中在基因的克隆和功能鉴定，在转录调控方面进展缓慢。大多数NAC蛋白所涉及的调控网络还没有被揭示，同时NAC蛋白在植物体内的转录调控机制还不明确。
植物基因启动子中包含多种重要的顺式作用元件，在转录水平上参与调控下游相应基因表达。因此分离与弄清楚植物启动子的分子本质不仅是研究基因表达调控机制的重要内容，而且也是构建基因工程表达载体的关键。
发明内容
本发明的一个目的是提供DNA片段。
本发明提供的DNA片段，为如下1)-3)中任一所述的DNA分子：
1)编码区为序列表中序列1所示的DNA分子；
2)在严格条件下与1)限定的DNA序列杂交且具有启动子功能的DNA分子；
3)与1)限定的DNA序列至少具有70％、至少具有75％、至少具有80％、至少具有85％、至少具有90％、至少具有95％、至少具有96％、至少具有97％、至少具有98％或至少具有99％同源性且具有启动子功能的DNA分子。
含有上述DNA片段的重组载体、表达盒、转基因细胞系或重组菌也是本发明保护的范围。
扩增上述DNA片段的引物对也是本发明保护的范围。
上述引物对中的一条引物的序列为序列2，另一条引物的序列为序列3。
上述DNA片段在使目的基因在植物组织中的表达中的应用也是本发明保护的范围。
上述DNA片段在作为启动子的应用也是本发明保护的范围。
上述应用中，所述植物组织为根、茎和/或叶片。
上述应用中，所述植物为双子叶植物。
上述应用中，所述双子叶植物为杨树。
上述DNA片段在植物的遗传育种中的应用也是本发明保护的范围。
上述应用中个，所述植物为双子叶植物；所述双子叶植物具体为杨树。
上述遗传育种包括研究维管组织基因表达模式或转录调控机制、构建转基因植物、改良植物性状、培养植物新品种等。
本发明的实验证明，本发明发现了启动子PtrNAC150，通过农杆菌介导将PtrNAC150启动子转入山新杨(Populus davidianaxPopulus bolleana)，转PtrNAC150启动子杨树GUS染色结果显示，GUS基因在根、茎、叶维管组织中特异表达，因此，应用本发明提供的启动子，可以在植物基因工程中，诱导相应目的基因在根、茎、叶维管组织中特异表达。
附图说明
图1为本发明植物表达载体PYBA-1121结构示意图。
图2为本发明转PtrNAC150pro杨树的GUS基因PCR检测结果。
图3为转基因植株GUS染色图。
具体实施方式
下述实施例中所使用的实验方法如无特殊说明，均为常规方法。
下述实施例中所用的材料、试剂等，如无特殊说明，均可从商业途径得到。
下面结合具体实施例对本发明作进一步说明，但本发明并不限于以下实施例。
下述实施例中，所述百分含量如无特殊说明，均为质量百分含量。
实施例1、源于杨树DNA片段PtrNAC150pro的获得
1、毛果杨DNA提取
以毛果杨叶片为材料，利用Qiagen公司植物基因组DNA提取试剂盒提取毛果杨叶片基因组DNA,具体步骤如下：
a.叶片组织用液氮冷冻，再用研钵研磨成很细的粉末，取100mg该粉末放入1.5ml离心管中。
b.加入400ul裂解液AP1盒4ul RNase A，使用涡旋振荡器振荡混匀，65℃孵育10分钟，期间颠倒2-3次。
c.加入130ul缓冲液P3，充分混匀，冰上放置5分钟。
d.20000×g室温离心5分钟，移取上清至旋转柱，20000×g，离心2分钟。
e.将滤液移至新的离心管，加入1.5倍体积缓冲液AW1，混合均匀。
f.取650ul混合物至DNeasy Mini吸附柱，6000×g，离心1分钟，弃滤液。如还有混合物，重复此步骤。
g.向吸附柱加入500ul漂洗液AW2，6000×g，离心1分钟,弃滤液。
h.再向吸附柱加入500ul漂洗液AW2，20000×g，离心2分钟,弃滤液。
i.将吸附柱放入新的1.5ml离心管中，向吸附膜的中部加入100ul灭菌ddH2O,室温静置5分钟，6000×g，离心1分钟，滤液即为基因组DNA。
2.引物设计与PCR扩增
(1)引物设计与合成
设计特异性引物，引物序列如下：
NAC150pF：5’GGAGATAACATAAGATTCAGGAGCC 3’
NAC150pR：5’GCTTCTTTCTCAGAGACAGCTGAT 3’
引物购自生工生物工程(上海)股份有限公司。
(2)PCR扩增
以上述1提取的毛果杨基因组DNA为模板，在高保真Phusion DNA酶的作用下，利用引物NAC150pF和NAC150pR进行PCR扩增。
反应体系为50ul，由下列成分组成：Phusion DNA聚合酶0.5ul(2U/ul),5xPhusion HF Buffer 10ul，预混dNTP(2.5mM each)1ul，模板DNA 5ul(200ng)，NAC150pF(10uM)2.5ul，NAC150pR(10uM)2.5ul，加无菌ddH2O补足50ul。
扩增条件为：98℃预变性30s；98℃，10s，65℃,40s，72℃,1min30s，进行30个循环；最后72℃继续延伸5min。
得到2387bp的PCR产物。
经过测序，该PCR产物具有序列表中序列1所示的核苷酸，将其命名为PtrNAC150pro。
将该PCR产物与载体Peasy-Blunt Zero(全式金，货号：CB501)中，得到中间载体Peasy-Blunt Zero-PtrNAC150pro。
实施例2、DNA片段PtrNAC150pro在作为启动子中的应用
1、重组表达载体pYBA-1121-PtrNAC150pro-GUS的构建
用NotⅠ和PstⅠ限制性内切酶Peasy-Blunt Zero-PtrNAC150pro，回收2431bp的PtrNAC150pro启动子片段；将该片段与经过同样酶切的pYBA-1121载体(GenBank号：KF876811；序列2)片段连接，得到重组载体pYBA-1121-PtrNAC150pro-GUS，其结构示意图见图1。
经过测序，重组载体pYBA-1121-PtrNAC150pro-GUS为将序列表中序列1所示的PtrNAC150pro替换pYBA-1121载体的NotⅠ和PstⅠ酶切位点间的DNA片段，得到的重组载体。
2、转PtrNAC150pro杨树的获得
将重组载体pYBA-1121-PtrNAC150pro-GUS转入农杆菌EHA105中，得到重组菌EHA105/pYBA-1121-PtrNAC150pro-GUS。
将重组菌EHA105/pYBA-1121-PtrNAC150pro-GUS培养得到菌液，划线于YEP固体培养基(利福平25mg/L,四环素5mg/L，卡那霉素50mg/L),28℃暗培养2天，取单菌落接种于5mlYEP液体培养基(利福平25mg/L,四环素5mg/L，卡那霉素50mg/L)中，28℃，200rpm过夜振荡培养；次日扩大培养于50ml的YEP液体培养基中，振荡培养至OD600＝0.6-0.8，离心收集菌体，重悬于MS侵染培养液中，侵染前加入200μM乙酰丁香酮，得活化的农杆菌悬液。
外植体选择山新杨(Populus davidiana×P.bolleana)(以下称为非转基因杨树)无菌苗，取新鲜、幼嫩的无菌苗叶片，用解剖刀切成1cm2大小的叶盘，将叶盘与活化的EHA105/pYBA-1121-PtrNAC150pro-GUS菌液混合，室温下侵染15min。
在固体共培养基上放置灭菌滤纸，将农杆菌侵染后的外植体表面菌液吸净后放置在所述灭菌滤纸上，在22-23℃条件下黑暗培养3-4天。固体共培养基的制备方法为：蒸馏水加入0.7％(W/V)的琼脂粉，高压灭菌后加入200uM AS。滤纸为Whatman No.1定性滤纸(直径9cm)。
用附加250mg/L Timentin的无菌水洗外植体3-4遍，再用无菌滤纸吸去残留液体，于分化培养基上光照培养5-7天，然后转移至选择培养基(MS+NAA0.1mg/L+6-BA0.2mg/L+TDZ0.01mg/L+Km50mg/L+Timentin 250mg/L)上培养，待抗性芽长至1-2cm时转至生根培养基(MS+NAA 0.1mg/L+Km 50mg/L+Timentin 250mg/L)上生根。温度25℃，光照14h/d，光照强度1600-2000lux。待植株长出根系，得到转PtrNAC150pro杨树。
载体pYBA-1121中含有GUS,但是GUS前面没有启动子因此不表达。
3、转PtrNAC150pro杨树的PCR检测
利用Qiagen基因组DNA提取试剂盒，提取转PtrNAC150pro杨树和非转基因杨树叶片的基因组DNA,上游引物从PtrNAC150pro序列中设计，下游引物从GUS序列中设计，PCR引物分别为：NAC150-Pro-F：5’GCTCGCAGGATATGAATAGCA 3’和GUS-R：5’TTGCCCGGCTTTCTTGTAAC 3’，进行PCR扩增。以非转基因杨树作为阴性对照，不加DNA的作为空白对照。
反应体系：基因组DNA 50ng；rTaqPreMix 12.5ul；上游引物(10uM)1.0ul；下游引物(10uM)1.0ul；加ddH2O至25ul。反应条件：94℃预变性3min；94℃，30s，55℃,40s，72℃,1min，进行30个循环；最后72℃继续延伸5min。
反应产物在1.0％琼脂糖凝胶电泳上检测。
结果如图2所示，M：Marker；1：空白对照；2-3：WT；4：pYBA-1121-PtrNAC150pro-GUS质粒阳性对照；5-11：转PtrNAC150pro杨树；可以看出，7株转PtrNAC150pro杨树4株得到343bp的产物，为阳性转PtrNAC150pro杨树，说明外源基因已经整合到杨树基因组DNA中。
非转基因杨树未得到扩增产物。
4、转PtrNAC150pro杨树的GUS染色
根据Jefferson等(Jefferson RA,Kavanagh TA,Bevan MW(1987)GUS fusions:beta-glucuronidase as a sensitive and versatile gene fusion marker in higherplants.EMBO J6:3901.)的方法对阳性转PtrNAC150pro杨树材料进行GUS组织化学染色分析，将材料浸泡于GUS活性检测液，37℃过夜，然后用70％乙醇脱色，观察并拍照，其中GUS活性检测液包含：0.1mol/L K4Fe(CN)6、0.1mol/L K3Fe(CN)6、50mmol/L磷酸钠缓冲液(pH7.0)、10mmol/L Na2EDTA、0.001％(v/v)Triton X-100、0.5mg/ml X-Gluc和20％甲醇。以非转基因杨树作为对照。
结果如图3所示，；a1-c1:转PtrNAC150pro杨树GUS染色；a2-c2：野生型GUS染色；a：叶；b：茎；c：根；可以看出，阳性转PtrNAC150pro杨树中，PtrNAC150启动子驱动的GUS基因在叶片、茎和根的维管组织中特异表达，阴性对照(非转基因杨树)无GUS表达活性。
序列表
<110>北京农业生物技术研究中心
<120> 一种杨树NAC基因启动子及其应用
<160> 2
<170> PatentIn version 3.5
<210> 1
<211> 2387
<212> DNA
<213> 人工序列
<220>
<223>
<400> 1
ggagataaca taagattcag gagcctatat gaagagttat gttacttctc taggttctag 60
ttagccttgg tgatatatat ttaactatag gacaggggtc ctgttgcttt gttgagattg 120
tctatctcaa actgtcatgg aaggtagatc tatggcacta gggagaaccc cttccaagga 180
gactgtgcaa tcacactagg gagagcacat tctaaggaga ttgtgtaatc ccactaggaa 240
aaaacccttt tcaggagagt gtgtaagagt ccagaagcct ccattaaaga tatgtagata 300
gaaagtgtat gttagaaaga taattatttt aatttaaaat taaacttact tcctatcatg 360
gataatattt tccaaggtga tttattggac tctagggagc tattagagag tggtgctatt 420
gcactataga gacccctgat caatcccttc cgagggtggg tccgccatga agacttgcta 480
aggcagcaaa acctgatttt gcacttgagt ggccaacaca actcgggtga ggggttccac 540
ttggttgata agctgttgga agttcgaatg agcagaagtg ctcgcgataa cgggtaaatc 600
catcaagcgt tctagctcag gtgtgtcatg gttgtttgcc atgtaatgac ttggaaatgt 660
cattagaaag aatttgagta ctttttttta tcagtattca atggatggtg caattgatga 720
tgttctagaa aattcaagtg gcttaggggc aatacttata ttggataatc aattaatctc 780
ttagttttag taatttaatc cattcttcct agctaaggtg gctggaggca ggtgcctaat 840
acctgtaaaa gatccccttt ggtgaattta gagattttct aatgattaag tcggtgactt 900
tttaagagaa aatgcaataa tattacaaag ttttttttta tgaagattct gatttaagtt 960
agtgattaag tctatttata gctcatgagt ttttttatgg agatgagatg ttaaagtgta 1020
atattgtctt gataatatta aaaagagaaa atatctctga cgtatatatt aatgttgatg 1080
agaaatatcc ctgcatatgt attaatgaga gaaaatatcc ctgacatatg tattaatttg 1140
gatgaaaata tggtgagctc ttaaaggact ttgacacaat atcttttaca caacattgtt 1200
atcgatgaaa atatgatagg tcattgaagg acttttatac atttaatgag tgtagagaaa 1260
gtaggatcca aaaaatatag ctaaactcaa ggaagatgag tcatttcttg gctgaacgga 1320
aaggaggtgg atcgtttctt tagctgagtc taatgaaagt taggtttttt atggtttttt 1380
gggttattaa atttgagttt atcaaataaa aaataagaaa attatattgt tgtgatttgt 1440
aaaaagaaga agaagaataa gataattatg tactagagta aacgtttata tgtatatata 1500
taacatgctt ttgattgtag ggcttaagta acagctatga aatctgtttc tttccttttg 1560
ctttagtagt caagccctcg aaacaaaagg agtatgagag agaaggttcc ttaagcttta 1620
gaaaatccaa ctggaatgta tttttttctc tctctcacat aaaccgctta cagcgggcta 1680
cttaaattgg caaagactct taaaggtgag tgcctagcta ggcttgaggg aaccttttcc 1740
tttttctcac tgacttcggt taaagtaaga gactagagag agcgatttct ggtgatttca 1800
atggttgaag aaccaggctt atacctaaat tccacagcca tagtttcctc ttttcgggct 1860
cttcctcctc caacttcgtt cctatcccct cttactacaa gttttctttt cccttaacga 1920
atttaaaaca ttcatacttc tgttttggag atatttctca tctgggtttt tttttttttt 1980
tctgactact ttttcgtctg ttgaaagtat cgaggctgta tagtactcgt atatttccgt 2040
tccacacata tatatatagt tgttgagtta tttttatgat tcttgattga atcaacatct 2100
gcaagtcttt caagttgttt ttctttgaat cttatctcaa aagaaagccg tatattatat 2160
gcatattttt gttttcaaga caaatattaa agcttattct taattaatag ctataattaa 2220
tatacgtgct tttgttaaca tgagttcata aatcaaattc atgatttgtg ctgtatagat 2280
agctcgcagg atatgaatag caagctgttg tttaagtttt taaaagttag gcttgggaga 2340
aagaagatac ttgtacacat agcatcagct gtctctgaga aagaagc 2387
<210> 2
<211> 7242
<212> DNA
<213> 人工序列
<220>
<223>
<400> 2
tttccatagg ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg 60
gcgaaacccg acaggactat aaagatacca ggcgtttccc cctggaagct ccctcgtgcg 120
ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc cttcgggaag 180
cgtggcgctt tctcatagct cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc 240
caagctgggc tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct tatccggtaa 300
ctatcgtctt gagtccaacc cggtaagaca cgacttatcg ccactggcag cagccactgg 360
taacaggatt agcagagcga ggtatgtagg cggtgctaca gagttcttga agtggtggcc 420
taactacggc tacactagaa gaacagtatt tggtatctgc gctctgctga agccagttac 480
cttcggaaaa agagttggta gctcttgatc cggcaaacaa accaccgctg gtagcggtgg 540
tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag ctagcttgag 600
tatactcact agactttgct tcgcaaagtc gtgaccgcct acggcggctg cggcgcccta 660
cgggcttgct ctccgggctt cgccctgcgc ggtcgctgcg ctcccttgcc agcccgtgga 720
tatgtggacg atggccgcga gcggccaccg gctggctcgc ttcgctcggc ccgtggacaa 780
ccctgctgga caagctgatg gacaggctgc gcctgcccac gagcttgacc acagggattg 840
cccaccggct acccagcctt cgaccacata cccaccggct ccaactgcgc ggcctgcggc 900
cttgccccat caattttttt aattttctct ggggaaaagc ctccggcctg cggcctgcgc 960
gcttcgcttg ccggttggac accaagtgga aggcgggtca aggctcgcgc agcgaccgcg 1020
cagcggcttg gccttgacgc gcctggaacg acccaagcct atgcgagtgg gggcagtcga 1080
aggcgaagcc cgcccgcctg ccccccgagc ctcacggcgg cgagtgcggg ggttccaagg 1140
gggcagcgcc accttgggca aggccgaagg ccgcgcagtc gatcaacaag ccccggaggg 1200
gccacttttt gccggagggg gagccgcgcc gaaggcgtgg gggaaccccg caggggtgcc 1260
cttctttggg caccaaagaa ctagatatag ggcgaaatgc gaaagactta aaaatcaaca 1320
acttaaaaaa ggggggtacg caacagctca ttgcggcacc ccccgcaata gctcattgcg 1380
taggttaaag aaaatctgta attgactgcc acttttacgc aacgcataat tgttgtcgcg 1440
ctgccgaaaa gttgcagctg attgcgcatg gtgccgcaac cgtgcggcac cctaccgcat 1500
ggagataagc atggccacgc agtccagaga aatcggcatt caagccaaga acaagcccgg 1560
tcactgggtg caaacggaac gcaaagcgca tgaggcgtgg gccgggctta ttgcgaggaa 1620
acccacggcg gcaatgctgc tgcatcacct cgtggcgcag atgggccacc agaacgccgt 1680
ggtggtcagc cagaagacac tttccaagct catcggacgt tctttgcgga cggtccaata 1740
cgcagtcaag gacttggtgg ccgagcgctg gatctccgtc gtgaagctca acggccccgg 1800
caccgtgtcg gcctacgtgg tcaatgaccg cgtggcgtgg ggccagcccc gcgaccagtt 1860
gcgcctgtcg gtgttcagtg ccgccgtggt ggttgatcac gacgaccagg acgaatcgct 1920
gttggggcat ggcgacctgc gccgcatccc gaccctgtat ccgggcgagc agcaactacc 1980
gaccggcccc ggcgaggagc cgcccagcca gcccggcatt ccgggcatgg aaccagacct 2040
gccagccttg accgaaacgg aggaatggga acggcgcggg cagcagcgcc tgccgatgcc 2100
cgatgagccg tgttttctgg acgatggcga gccgttggag ccgccgacac gggtcacgct 2160
gccgcgccgg tagcacttgg gttgcgcagc aacccgtaag tgcgctgttc cagactatcg 2220
gctgtagccg cctcgccgcc ctataccttg tctgcctccc cgcgttgcgt cgcggtgcat 2280
ggagccgggc cacctcgacc tgaatggaag ccggcggcac ctcgctaacg gattcaccgt 2340
ttttatcagg ctctgggagg cagaatacaa acaaacacat acagcgactt agtttacccg 2400
ccaatatatc ctgtcaacgc gtttaaactg gattttggtt ttaggaatta gaaattttat 2460
tgatagaagt attttacaaa tacaaataca tactaagggt ttcttatatg ctcaacacat 2520
gagcgaaacc ctataagaac cctaattccc ttatctggga actactcaca cattattctg 2580
gagaaaaata gagagagata gatttgtaga gagagactgg tgatttgctc gcgatcattg 2640
tttgcctccc tgctgcggtt tttcaccgaa gttcatgcca gtccagcgtt tttgcagcag 2700
aaaagccgcc gacttcggtt tgcggtcgcg agtgaagatc cctttcttgt taccgccaac 2760
gcgcaatatg ccttgcgagg tcgcaaaatc ggcgaaattc catacctgtt caccgacgac 2820
ggcgctgacg cgatcaaaga cgcggtgata catatccagc catgcacact gatactcttc 2880
actccacatg tcggtgtaca ttgagtgcag cccggctaac gtatccacgc cgtattcggt 2940
gatgataatc ggctgatgca gtttctcctg ccaggccaga agttcttttt ccagtacctt 3000
ctctgccgtt tccaaatcgc cgctttggac ataccatccg taataacggt tcaggcacag 3060
cacatcaaag agatcgctga tggtatcggt gtgagcgtcg cagaacatta cattgacgca 3120
ggtgatcgga cgcgtcgggt cgagtttacg cgttgcttcc gccagtggcg cgaaatattc 3180
ccgtgcacct tgcggacggg tatccggttc gttggcaata ctccacatca ccacgcttgg 3240
gtggtttttg tcacgcgcta tcagctcttt aatcgcctgt aagtgcgctt gctgagtttc 3300
cccgttgact gcctcttcgc tgtacagttc tttcggcttg ttgcccgctt cgaaaccaat 3360
gcctaaagag aggttaaagc cgacagcagc agtttcatca atcaccacga tgccatgttc 3420
atctgcccag tcgagcatct cttcagcgta agggtaatgc gaggtacggt aggagttggc 3480
cccaatccag tccattaatg cgtggtcgtg caccatcagc acgttatcga atcctttgcc 3540
acgcaagtcc gcatcttcat gacgaccaaa gccagtaaag tagaacggtt tgtggttaat 3600
caggaactgt tcgcccttca ctgccactga ccggatgccg acgcgaagcg ggtagatatc 3660
acactctgtc tggcttttgg ctgtgacgca cagttcatag agataacctt cacccggttg 3720
ccagaggtgc ggattcacca cttgcaaagt cccgctagtg ccttgtccag ttgcaaccac 3780
ctgttgatcc gcatcacgca gttcaacgct gacatcacca ttggccacca cctgccagtc 3840
aacagacgcg tggttacagt cttgcgcgac atgcgtcacc acggtgatat cgtccaccca 3900
ggtgttcggc gtggtgtaga gcattacgct gcgatggatt ccggcatagt taaagaaatc 3960
atggaagtaa gactgctttt tcttgccgtt ttcgtcggta atcaccattc ccggcgggat 4020
agtctgccag ttcagttcgt tgttcacaca aacggtgata cgtacacttt tcccggcaat 4080
aacatacggc gtgacatcgg cttcaaatgg cgtatagccg ccctgatgct ccatcacttc 4140
ctgattattg acccacactt tgccgtaatg agtgaccgca tcgaaacgca gcacgatacg 4200
ctggcctgcc caacctttcg gtataaagac ttcgcgctga taccagacgt tgcccgcata 4260
attacgaata tctgcatcgg cgaactgatc gttaaaactg cctggcacag caattgcccg 4320
gctttcttgt aacgcgcttt cccaccaacg ctgatcaatt ccacagtttt cgcgatccag 4380
actgaatgcc cacaggccgt cgagtttttt gatttcacgg gttggggttt ctacaggacg 4440
taacatggta ccgggccccc cctcgaggtc gacggtatcg ataagcttga tatcgaattc 4500
ctgcagcccg ggggatccac tagttctaga gcggccgcca ccgcggtgga gctccaattg 4560
gccctataga ccggtcgtac gaggcctatt aatcccgatc tagtaacata gatgacaccg 4620
cgcgcgataa tttatcctag tttgcgcgct atattttgtt ttctatcgcg tattaaatgt 4680
ataattgcgg gactctaatc ataaaaaccc atctcataaa taacgtcatg cattacatgt 4740
taattattac atgcttaacg taattcaaca gaaattatat gataatcatc gcaagaccgg 4800
caacaggatt caatcttaag aaactttatt gccaaatgtt tgaacgatct cagaagaact 4860
cgtcaagaag gcgatagaag gcgatgcgct gcgaatcggg agcggcgata ccgtaaagca 4920
cgaggaagcg gtcagcccat tcgccgccaa gctcttcagc aatatcacgg gtagccaacg 4980
ctatgtcctg atagcggtcc gccacaccca gccggccaca gtcgatgaat ccagaaaagc 5040
ggccattttc caccatgata ttcggcaagc aggcatcgcc atgggtcacg acgagatcct 5100
cgccgtcggg catgcgcgcc ttgagcctgg cgaacagttc ggctggcgcg agcccctgat 5160
gctcttcgtc cagatcatcc tgatcgacaa gaccggcttc catccgagta cgtgctcgct 5220
cgatgcgatg tttcgcttgg tggtcgaatg ggcaggtagc cggatcaagc gtatgcagcc 5280
gccgcattgc atcagccatg atggatactt tctcggcagg agcaaggtga gatgacagga 5340
gatcctgccc cggcacttcg cccaatagca gccagtccct tcccgcttca gtgacaacgt 5400
cgagcacagc tgcgcaagga acgcccgtcg tggccagcca cgatagccgc gctgcctcgt 5460
cctgaagttc attcagggca ccggacaggt cggtcttgac aaaaagaacc gggcgcccct 5520
gcgctgacag ccggaacacg gcggcatcag agcagccgat tgtctgttgt gcccagtcat 5580
agccgaatag cctctccacc caagcggccg gagaacctgc gtgcaatcca tcttgttcaa 5640
tcatagatcc ggtgcagatt atttggattg agagtgaata tgagactcta attggatacc 5700
gaggggaatt tatggaacgt cagtggagca tttttgacaa gaaatatttg ctagctgata 5760
gtgaccttag gcgacttttg aacgcgcaat aatggtttct gacgtatgtg cttagctcat 5820
taaactccag aaacccgcgg ctgagtggct ccttcaacgt tgcggttctg tcagttccaa 5880
acgtaaaacg gcttgtcccg cgtcatcggc gggggtcata acgtgactcc cttaattctc 5940
cgctcatgat cggcgcgcca cgtgacgcgt gtttacacca caatatatcc tgccacgatt 6000
tgcccatggg ggtgggcgaa gaactccagc atgagatccc cgcgctggag gatcatccag 6060
ccggcgtccc ggaaaacgat tccgaagccc aacctttcat agaaggcggc ggtggaatcg 6120
aaatctcgtg atggcaggtt gggcgtcgct tggtcggtca tttcgaaccc cagagtcccg 6180
ctcagaagaa ctcgtcaaga aggcgataga aggcgatgcg ctgcgaatcg ggagcggcga 6240
taccgtaaag cacgaggaag cggtcagccc attcgccgcc aagctcttca gcaatatcac 6300
gggtagccaa cgctatgtcc tgatagcggt ccgccacacc cagccggcca cagtcgatga 6360
atccagaaaa gcggccattt tccaccatga tattcggcaa gcaggcatcg ccatgggtca 6420
cgacgagatc ctcgccgtcg ggcatgcgcg ccttgagcct ggcgaacagt tcggctggcg 6480
cgagcccctg atgctcttcg tccagatcat cctgatcgac aagaccggct tccatccgag 6540
tacgtgctcg ctcgatgcga tgtttcgctt ggtggtcgaa tgggcaggta gccggatcaa 6600
gcgtatgcag ccgccgcatt gcatcagcca tgatggatac tttctcggca ggagcaaggt 6660
gagatgacag gagatcctgc cccggcactt cgcccaatag cagccagtcc cttcccgctt 6720
cagtgacaac gtcgagcaca gctgcgcaag gaacgcccgt cgtggccagc cacgatagcc 6780
gcgctgcctc gtcctgaagt tcattcaggg caccggacag gtcggtcttg acaaaaagaa 6840
ccgggcgccc ctgcgctgac agccggaaca cggcggcatc agagcagccg attgtctgtt 6900
gtgcccagtc atagccgaat agcctctcca cccaagcggc cggagaacct gcgtgcaatc 6960
catcttgttc aatcatgcga aacgatcctc atcctgtctc ttgatcagat cttgatcccc 7020
tgcgccatca gatccttggc ggcaagaaag ccatccagtt tactttgcag ggcttcccaa 7080
ccttaccaga gggcgcccca gctggcaatt ccggttcgct tgctgtccat aaaaccgccc 7140
agtctagcta tcgccatgta agcccactgc aagctacctg ctttctcttt gcgcttgcgt 7200
tttcccttgt ccagatagcc cagtagctga cattcatccc ag 7242
Claims (11)
1.DNA片段，为编码区为序列表中序列1所示的DNA分子。
2.含有权利要求1中所述DNA片段的重组载体、表达盒、转基因细胞系或重组菌。
3.扩增权利要求1中所述DNA片段的引物对。
4.根据权利要求3所述的引物对，其特征在于：所述引物对中的一条引物的序列为序列2，另一条引物的序列为序列3。
5.权利要求1所述DNA片段在使目的基因在植物组织中的表达中的应用；
或权利要求1所述DNA片段在作为启动子中的应用。
6.根据权利要求5中所述的应用，其特征在于：所述植物组织为根、茎和/或叶片。
7.根据权利要求5或6中所述的应用，其特征在于：所述植物为双子叶植物。
8.根据权利要求7所述的应用，其特征在于：所述双子叶植物为杨树。
9.权利要求1中所述DNA片段在植物的遗传育种中的应用。
10.根据权利要求9所述的应用，其特征在于：所述植物为双子叶植物。
11.根据权利要求10所述的应用，其特征在于：所述双子叶植物为杨树。
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201810146879.9A CN108070597B (zh) | 2018-02-12 | 2018-02-12 | 一种杨树nac基因启动子及其应用 |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201810146879.9A CN108070597B (zh) | 2018-02-12 | 2018-02-12 | 一种杨树nac基因启动子及其应用 |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN108070597A CN108070597A (zh) | 2018-05-25 |
| CN108070597B true CN108070597B (zh) | 2020-04-07 |
Family
ID=62155213
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201810146879.9A Active CN108070597B (zh) | 2018-02-12 | 2018-02-12 | 一种杨树nac基因启动子及其应用 |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN108070597B (zh) |
Family Cites Families (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102676458A (zh) * | 2011-06-02 | 2012-09-19 | 北京未名凯拓作物设计中心有限公司 | 一个组成型表达启动子及其应用 |
| CN102559682B (zh) * | 2012-01-19 | 2013-04-03 | 南京林业大学 | 一种杨树根原基特异表达启动子ProWOX11a及其应用 |
| US9738901B2 (en) * | 2012-05-10 | 2017-08-22 | The Regents Of The University Of California | Regulation of galactan synthase expression to modify galactan content in plants |
| US10774338B2 (en) * | 2014-01-16 | 2020-09-15 | The Regents Of The University Of California | Generation of heritable chimeric plant traits |
| CN106916828A (zh) * | 2017-05-03 | 2017-07-04 | 中国林业科学研究院林业研究所 | 一种调控杨树叶片发育的生长调节因子基因及其应用 |
-
2018
- 2018-02-12 CN CN201810146879.9A patent/CN108070597B/zh active Active
Also Published As
| Publication number | Publication date |
|---|---|
| CN108070597A (zh) | 2018-05-25 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| Long et al. | miR156-SPL modules regulate induction of somatic embryogenesis in citrus callus | |
| KR102223568B1 (ko) | 식물의 게놈 내의 외인성 서열의 통합을 위한 방법 및 조성물 | |
| CN105132425B (zh) | 用于增强植物中组成型基因表达的调节性核酸分子 | |
| CN109652440A (zh) | VQRn-Cas9&PmCDA1&UGI碱基编辑系统在植物基因编辑中的应用 | |
| CN106906214B (zh) | 新型植物终止子序列 | |
| Tao et al. | The NAC transcription factor ANAC017 regulates aluminum tolerance by regulating the cell wall-modifying genes | |
| Sinharoy et al. | Transformed hairy roots of arachis hypogea: A tool for studying root nodule symbiosis in a non–infection thread legume of the aeschynomeneae tribe | |
| Marjamaa et al. | Cloning, characterization and localization of three novel class III peroxidases in lignifying xylem of Norway spruce (Picea abies) | |
| Gupta et al. | The C4Ppc promoters of many C4 grass species share a common regulatory mechanism for gene expression in the mesophyll cell | |
| CN114164230B (zh) | 一种适用于甘蔗遗传转化的表达载体及其构建方法和应用 | |
| CN110628795B (zh) | 以失活的筛选剂抗性基因为报告体系的a·g碱基替换的细胞富集技术及其应用 | |
| CN114317590B (zh) | 一种将植物基因组中的碱基c突变为碱基t的方法 | |
| CN108070597B (zh) | 一种杨树nac基因启动子及其应用 | |
| CN111321167B (zh) | 一种异源蛋白表达的滚环复制重组载体构建方法及其应用 | |
| Tsuyama et al. | Coniferin β-glucosidase is ionically bound to cell wall in differentiating xylem of poplar | |
| EP3090054B1 (en) | A method for improving stem volume growth and biomass production in trees | |
| KR100974820B1 (ko) | 고추 아쿠아포린 유전자 유래 뿌리 특이적 발현 프로모터및 이를 포함하는 뿌리 특이적 발현 벡터 | |
| CN108624544B (zh) | 阿卡波糖工程菌及其制备方法和应用 | |
| Shi et al. | Programmable RNA N6‐methyladenosine editing with CRISPR/dCas13a in plants | |
| KR20090050189A (ko) | 톡소플라빈 분해 효소를 선별 마커로 이용한 형질전환 감자식물체 제조 | |
| CN113122554A (zh) | 在拟南芥中高效表达Bs3Cas12b蛋白的核酸分子及其在基因组编辑中的应用 | |
| CN113481233A (zh) | 构建依克多因生产菌的方法 | |
| Cho et al. | Evaluation of a novel promoter from Populus trichocarpa for mature xylem tissue specific gene delivery | |
| CN104109682B (zh) | 一种果胶裂解酶BnPL基因及其启动子和应用 | |
| CN114317518B (zh) | SpRYn-CBE碱基编辑系统在植物基因组碱基替换中的应用 |
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 | ||
| GR01 | Patent grant | ||
| GR01 | Patent grant |