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WO2014205599A1 - Antiport sodium-hydrogène de tonoplaste dans nhx3 dethellungiella halophila, gène codant et son utilisation - Google Patents

Antiport sodium-hydrogène de tonoplaste dans nhx3 dethellungiella halophila, gène codant et son utilisation Download PDF

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Publication number
WO2014205599A1
WO2014205599A1 PCT/CN2013/000752 CN2013000752W WO2014205599A1 WO 2014205599 A1 WO2014205599 A1 WO 2014205599A1 CN 2013000752 W CN2013000752 W CN 2013000752W WO 2014205599 A1 WO2014205599 A1 WO 2014205599A1
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WO
WIPO (PCT)
Prior art keywords
plant
seq
gene
expression vector
plants
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.)
Ceased
Application number
PCT/CN2013/000752
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English (en)
Chinese (zh)
Inventor
何云蔚
王建胜
王君丹
刘捷
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BIOCENTURY TRANSGENE (CHINA) Co Ltd
Original Assignee
BIOCENTURY TRANSGENE (CHINA) Co Ltd
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Publication date
Application filed by BIOCENTURY TRANSGENE (CHINA) Co Ltd filed Critical BIOCENTURY TRANSGENE (CHINA) Co Ltd
Priority to CN201380074520.0A priority Critical patent/CN105189534A/zh
Priority to PCT/CN2013/000752 priority patent/WO2014205599A1/fr
Publication of WO2014205599A1 publication Critical patent/WO2014205599A1/fr
Anticipated expiration legal-status Critical
Ceased legal-status Critical Current

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    • 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)
    • C12N15/8241—Phenotypically and genetically modified plants via recombinant DNA technology
    • C12N15/8261—Phenotypically and genetically modified plants via recombinant DNA technology with agronomic (input) traits, e.g. crop yield
    • C12N15/8271—Phenotypically and genetically modified plants via recombinant DNA technology with agronomic (input) traits, e.g. crop yield for stress resistance, e.g. heavy metal resistance
    • C12N15/8273—Phenotypically and genetically modified plants via recombinant DNA technology with agronomic (input) traits, e.g. crop yield for stress resistance, e.g. heavy metal resistance for drought, cold, salt resistance

Definitions

  • Two tester cDNAs with different adaptors were mixed with an excess of Driver for the first forward subtractive hybridization.
  • the products of the two first forward subtractive hybridizations were mixed, and a second forward subtractive hybridization was performed with the newly denatured Driver cDNA, and the differentially expressed genes were amplified by two inhibitory PCR amplifications (PCR). Before, the second forward subtractive hybridization product is end-filled).
  • the first round of PCR amplification was carried out using SEQ ID NO: 4 and the universal primer AUAP (provided with the kit), and the cDNA obtained by reverse transcription of the mRNA extracted from the salt-treated group was used as a template. Specific steps are as follows:
  • PCR amplification product plus A tail 2.5 times the volume of absolute ethanol was added to the PCR product, placed at -20 ° C for 10 minutes, centrifuged, the supernatant was removed, air-dried, and the resulting precipitate was dissolved in 21 ⁇ L of double distilled water. Then add 2.5 ⁇ to it ⁇ Buffer 0.5 ⁇ 5 mM dATP, 1.0 lExTaq. Reaction conditions: The reaction was carried out at 70 ° C for 30 minutes. The obtained 1400 bp DNA fragment was recovered (Omega recovery kit), and ligated into the pGEM T-easy vector to obtain a ThNHX3-pGEM plasmid, and then the ligation product was transformed into E.
  • Amino acid sequence of NHX3 protein SEQ ID NO: 1
  • TCAGAATTCCCAGTGAATTCCCGATCTAGTA The 35S promoter was amplified using the primers SEQ ID NO: 16 and SEQ ID NO: 17 using the pCAMBIA2300 plasmid as a template.
  • TAKARA's PrimeSTAR HS DNA polymerase was used. 50 ⁇ PCR reaction system: 10 ⁇ 5 xPS Buffer 3 ⁇ 2.5 mM dNTP, 1.0 ⁇ pCAMBIA2300 plasmid, 1.0 ⁇ PrimeSTAR HS DNA polymerase, 10 ⁇ primers SEQ ID NO: 16 and SEQ ID NO: 17 each 2.0 ⁇ and 31 ⁇ double distilled water.
  • M is the DNA Ladder Marker (DL2000, purchased from Shenzhen Ruizhen Biotechnology Co., Ltd.), and 1-8 is the salt-tolerant T1 transgenic Arabidopsis plants ( belonging to ⁇ 1 ⁇ , Tlf7, TlflO, respectively).
  • Three strains) 9 is the 35S-ThNHX3-2300 plasmid PCR positive control, and 10-13 is the non-transgenic control Arabidopsis plant.
  • the size of the band shown is the same as the size of the positive control (approximately 750 bp).
  • the results showed that the salt-tolerant T1 transgenic Arabidopsis plants had significant transcription in 73 ⁇ 4 ⁇ 3 ⁇ 4 ⁇ 5, and there was no transcription of ThNHJG in non-transgenic control Arabidopsis plants.

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  • Health & Medical Sciences (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Genetics & Genomics (AREA)
  • Chemical & Material Sciences (AREA)
  • Organic Chemistry (AREA)
  • Molecular Biology (AREA)
  • Engineering & Computer Science (AREA)
  • Bioinformatics & Cheminformatics (AREA)
  • Biophysics (AREA)
  • Biotechnology (AREA)
  • General Engineering & Computer Science (AREA)
  • General Health & Medical Sciences (AREA)
  • Biomedical Technology (AREA)
  • Biochemistry (AREA)
  • Wood Science & Technology (AREA)
  • Zoology (AREA)
  • Physics & Mathematics (AREA)
  • Microbiology (AREA)
  • Plant Pathology (AREA)
  • Cell Biology (AREA)
  • Botany (AREA)
  • Gastroenterology & Hepatology (AREA)
  • Medicinal Chemistry (AREA)
  • Proteomics, Peptides & Aminoacids (AREA)
  • Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)
  • Breeding Of Plants And Reproduction By Means Of Culturing (AREA)

Abstract

L'invention concerne un antiport sodium-hydrogène de tonoplaste dans NHX3 dérivé de Thellungiella halophila et son gène codant, son utilisation dans la culture de plantes transgéniques présentant une tolérance au sel accrue.
PCT/CN2013/000752 2013-06-24 2013-06-24 Antiport sodium-hydrogène de tonoplaste dans nhx3 dethellungiella halophila, gène codant et son utilisation Ceased WO2014205599A1 (fr)

Priority Applications (2)

Application Number Priority Date Filing Date Title
CN201380074520.0A CN105189534A (zh) 2013-06-24 2013-06-24 一种小盐芥液泡膜钠氢反向转运蛋白nhx3及其编码基因与应用
PCT/CN2013/000752 WO2014205599A1 (fr) 2013-06-24 2013-06-24 Antiport sodium-hydrogène de tonoplaste dans nhx3 dethellungiella halophila, gène codant et son utilisation

Applications Claiming Priority (1)

Application Number Priority Date Filing Date Title
PCT/CN2013/000752 WO2014205599A1 (fr) 2013-06-24 2013-06-24 Antiport sodium-hydrogène de tonoplaste dans nhx3 dethellungiella halophila, gène codant et son utilisation

Publications (1)

Publication Number Publication Date
WO2014205599A1 true WO2014205599A1 (fr) 2014-12-31

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PCT/CN2013/000752 Ceased WO2014205599A1 (fr) 2013-06-24 2013-06-24 Antiport sodium-hydrogène de tonoplaste dans nhx3 dethellungiella halophila, gène codant et son utilisation

Country Status (2)

Country Link
CN (1) CN105189534A (fr)
WO (1) WO2014205599A1 (fr)

Families Citing this family (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN118667833A (zh) * 2024-06-06 2024-09-20 山东中医药大学 一种忍冬耐盐基因LjNHX3及其应用

Citations (7)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US20030046729A1 (en) * 1998-03-18 2003-03-06 Eduardo Blumwald Increasing salt tolerance in plants by overexpression of vacuolar cation-proton antiporters
WO2006045829A1 (fr) * 2004-10-29 2006-05-04 Cropdesign N.V. Plantes à caractéristiques de croissance améliorées et procédé de production desdites plantes
WO2008129060A2 (fr) * 2007-04-23 2008-10-30 Basf Se Augmentation de la productivité d'une plante par combinaison d'agents chimiques avec des modifications transgéniques
CN101343316A (zh) * 2008-09-05 2009-01-14 中国科学院微生物研究所 一种钾离子转运相关蛋白系统及其编码基因簇与应用
CN101386854A (zh) * 2008-10-10 2009-03-18 哈尔滨师范大学 碱菀液泡膜Na+/H+反向转运蛋白的基因和氨基酸序列及基因制备方法和扩增引物
CN101456909A (zh) * 2009-01-13 2009-06-17 南京农业大学 一种大豆hkt类蛋白及其编码基因与应用
WO2012028646A1 (fr) * 2010-08-31 2012-03-08 Westfälische Wilhelms-Universität Münster Moyens destinés à améliorer des caractères agrobiologiques dans des plantes

Family Cites Families (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN1706951A (zh) * 2005-05-13 2005-12-14 山东大学 马蔺液泡膜Na+/H+逆向转运蛋白2基因及应用

Patent Citations (7)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US20030046729A1 (en) * 1998-03-18 2003-03-06 Eduardo Blumwald Increasing salt tolerance in plants by overexpression of vacuolar cation-proton antiporters
WO2006045829A1 (fr) * 2004-10-29 2006-05-04 Cropdesign N.V. Plantes à caractéristiques de croissance améliorées et procédé de production desdites plantes
WO2008129060A2 (fr) * 2007-04-23 2008-10-30 Basf Se Augmentation de la productivité d'une plante par combinaison d'agents chimiques avec des modifications transgéniques
CN101343316A (zh) * 2008-09-05 2009-01-14 中国科学院微生物研究所 一种钾离子转运相关蛋白系统及其编码基因簇与应用
CN101386854A (zh) * 2008-10-10 2009-03-18 哈尔滨师范大学 碱菀液泡膜Na+/H+反向转运蛋白的基因和氨基酸序列及基因制备方法和扩增引物
CN101456909A (zh) * 2009-01-13 2009-06-17 南京农业大学 一种大豆hkt类蛋白及其编码基因与应用
WO2012028646A1 (fr) * 2010-08-31 2012-03-08 Westfälische Wilhelms-Universität Münster Moyens destinés à améliorer des caractères agrobiologiques dans des plantes

Also Published As

Publication number Publication date
CN105189534A (zh) 2015-12-23

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