WO2000022149A1 - Procede de transformation - Google Patents

Procede de transformation Download PDF

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Publication number
WO2000022149A1
WO2000022149A1 PCT/IB1999/001676 IB9901676W WO0022149A1 WO 2000022149 A1 WO2000022149 A1 WO 2000022149A1 IB 9901676 W IB9901676 W IB 9901676W WO 0022149 A1 WO0022149 A1 WO 0022149A1
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WO
WIPO (PCT)
Prior art keywords
transformed
plant seed
wetting agent
surfactant
agrobacterium strain
Prior art date
Application number
PCT/IB1999/001676
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English (en)
Inventor
Jacoba Adriana De Ronde
William Cress
Original Assignee
Protein Research Trust
Priority date (The priority date 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 date listed.)
Filing date
Publication date
Application filed by Protein Research Trust filed Critical Protein Research Trust
Priority to AU59937/99A priority Critical patent/AU762964B2/en
Priority to EP99970425A priority patent/EP1121452A1/fr
Priority to CA002346004A priority patent/CA2346004A1/fr
Priority to BR9914572-3A priority patent/BR9914572A/pt
Publication of WO2000022149A1 publication Critical patent/WO2000022149A1/fr

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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
    • 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)
    • C12N15/8201—Methods for introducing genetic material into plant cells, e.g. DNA, RNA, stable or transient incorporation, tissue culture methods adapted for transformation
    • C12N15/8209—Selection, visualisation of transformants, reporter constructs, e.g. antibiotic resistance markers
    • 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)
    • C12N15/8201—Methods for introducing genetic material into plant cells, e.g. DNA, RNA, stable or transient incorporation, tissue culture methods adapted for transformation
    • C12N15/8202—Methods for introducing genetic material into plant cells, e.g. DNA, RNA, stable or transient incorporation, tissue culture methods adapted for transformation by biological means, e.g. cell mediated or natural vector
    • C12N15/8205—Agrobacterium mediated transformation
    • 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
    • C12N15/8251—Amino acid content, e.g. synthetic storage proteins, altering amino acid biosynthesis
    • 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

  • THIS INVENTION relates to a method for obtaining transformed or genetically modified plant seed . It also relates to a transformation composition.
  • a method for obtaining transformed or genetically modified plant seed including contacting germinating plant seed with a wetting agent or surfactant and an Agrobacterium strain to transform or genetically modify the plant seed .
  • the wetting agent or surfactant may be any suitable wetting agent or surfactant which facilitates or enhances penetration and transformation of germinating plant seed by the Agrobacterium strain.
  • the term "wetting agent” includes surfactants with wetting properties.
  • the wetting agent may be a non-oil based wetting agent, and may include a polyether polymethyl siloxane copolymer.
  • a suitable wetting agent is Break-Thru (available from Goldschmidt Chemical Corporation in Hopewell, USA) . It is believed that the active component of Break-Thru is polyether polymethyl siloxane copolymer, Break-Thru being a non-oil wetting agent.
  • the Agrobacterium strain and the wetting agent may be in the form of an admixture or suspension.
  • the wetting agent and the Agrobacterium strain may be present in the admixture in a mass ratio of the wetting agent: Agrobacterium strain of between 1 :99 and 1 : 10000, e.g. 1 : 1000.
  • the germinating plant seed may be subjected to vacuum infiltration while they are being contacted with the wetting agent and the Agrobacterium strain .
  • the germinating plant seed may be subjected to vacuum infiltration for a period of between 5 minutes and 40 minutes, e.g . 20 minutes, at a pressure of between 1 50 Pa (a) and 750 Pa (a), e.g . 585 Pa (a) .
  • the vacuum infiltration may be carried out at a temperature of between 1 5 °C and 35 °C, e.g. 25 °C.
  • the germinating plant seed may then be contacted with the admixture for a period of between 2 hours and 48 hours, e.g. 24 hours, at a temperature of between 1 5 °C and 35 °C, e.g . room or ambient temperature.
  • the Agrobacterium strain may be any suitable strain such as Agrobacterium tumefaciens, for example, Agrobacterium tumefaciens strain
  • the germinating plant seed may be transformed by the introduction of foreign DNA via the Agrobacterium strain.
  • the germinating plant seed may be transformed by exposing or contacting the germinating plant seed with a culture of Agrobacterium, said Agrobacterium strain being transformation competent and including a construct comprising a foreign gene, the foreign gene including appropriate regulatory sequences so as to be expressed in the cells of a plant which is cultivated from the transformed or genetically modified plant seed .
  • the foreign gene may be any suitable gene, such as a foreign gene which confers disease resistance and/or drought resistance to the plant which is cultivated from the transformed or genetically modified plant seed .
  • the Agrobacterium strain may include a suitable plasmid to facilitate transformation of the plant seed .
  • the plasmid may include a vector, such as vector pBI 1 21 .
  • the method may include inducing further growth of the transformed plant seed and selecting for a transformant in the presence of a selecting agent.
  • the Agrobacterium strain may include a plasmid which includes both said foreign gene and a selection agent resistance gene, the selection agent resistance gene also including appropriate regulatory sequences so as to be expressed in the cells of the plant which is cultivated from the transformed or genetically modified plant seed .
  • the selection agent resistance gene may code for antibiotic resistance, thus imparting resistance to an antibiotic selection agent to the plant which is cultivated from the transformed or genetically modified plant seed.
  • the antibiotic selection agent may be selected from the group consisting of at least one of kanamycin and rifampicin, and the selection agent resistance gene may be a GUS-intron gene.
  • the plant seed may be from the family Leguminoseae or any other dicotyledonous plant, for example, soybean or lupin seed .
  • soybean seed may be allowed to germinate until it has a small plumule, easily removable seed coat and cotyledons which are not appressed against each other before the germinating soybean seed is contacted with the wetting agent and the Agrobacterium strain.
  • lupin seed may be allowed to germinate until the plumule is between 1 0 - 20 mm in size, before the germinating lupin seed is contacted with the wetting agent and the Agrobacterium strain.
  • the method may include germinating plant seed at a temperature of between 22 °C and 32 °C, e.g. about 29 °C, for a period of between 2 days and
  • the invention extends to a transformed or genetically modified plant seed produced by the method as herein before described, to a transformed or genetically modified plant cultivated from said plant seed, to a plant seed produced by said transformed or genetically modified plant, and to a plant which is the progeny of said transformed or genetically modified plant.
  • the transformed or genetically modified plant may comprise cells which comprise in their genome at least one preselected foreign gene which produces a foreign cellular product encoded by the foreign gene.
  • the foreign gene may code for at least one of disease resistance and drought resistance.
  • a transformation composition which includes, in admixture, an Agrobacterium strain and a wetting agent.
  • the wetting agent may be a non-oil based wetting agent.
  • the wetting agent may include a polyether polymethyl siloxane copolymer, and the Agrobacterium strain may be Agrobacterium tumefaciens.
  • the wetting agent and the Agrobacterium strain may be present in a mass ratio of the wetting agent: Agrobacterium strain of between 1 :99 and 1 : 1 0000, e.g . 1 : 1 000.
  • Figure 1 A shows X-GLUC histochemical localization of GUS enzyme activity in GUS transformed soybean cultivar Carnia 2233 leaf tissue
  • Figure 1 B shows X-GLUC histochemical localization of GUS enzyme activity in GUS transformed soybean cultivar Carnia 2233 root tissue
  • Figure 2 shows X-GLUC histochemical localization of GUS enzyme activity in GUS-INT transformed soybean cultivar Carnia 2233 stomata;
  • Figure 3 shows the effect of mannitol stress on proline synthesis in third generation transformed soybean cultivar Carnia 2233 with an antisense P5CR construct.
  • Figure 4 shows a woodenbox screening of control soybean cultivar Carnia 2233 (without Arabidopsis P CR gene) compared to transformed soybean cultivar Carnia 2233 (containing P5CR gene in antisense orientation);
  • Figure 6 shows transformed lupin leaves of initial transformed seed, together with first generation seed compared to control plants leaves and seeds.
  • Soybean seed (Carnia 2233) was sterilised for 5 minutes in 3.5 %
  • soybean seeds were then sorted and soybean seeds having a small plumule, easily removable seed coat and cotyledons which were not appressed against each other, were selected .
  • the selected germinating or germinated soybean seeds were then contacted with the Agrobacterium/ wetting agent suspension and vacuum infiltrated for 20 minutes under a pressure of 78 millitorr.
  • the germinating soybean seeds were then incubated for a further period of 24 hours in contact with the Agrobacterium/wett ⁇ ig agent suspension solution at room or ambient temperature to obtain transformed soybean seeds.
  • the transformed soybean seeds were then planted in a soil mixture comprising soil, sand, vermiculite (5:5:3) and grown in a greenhouse to obtain transformed soybean plants. Percentage success rate of transformation was determined by detecting GUS-gene activity using a fluorometric and a histochemical assay.
  • the assay buffer contained 50 mM NaP0 4 (pH7.00), 10mM EDTA, 0.1 % (v/v) Triton X-100, 10mM mercapto ethanol and 2 M methyl umbilliferyl glucuronide (Sigma) . Small pieces of plant tissue were crushed in this buffer and incubated overnight at a temperature of 37 °C in the dark. Reactions were visualised on a long wave length UV light box. A histochemical assay was also performed in testing putative transformants. Plant tissue, which tested positive in the fluorescence assay, was incubated overnight at a temperature of 37 °C in the dark in a histochemical staining solution.
  • the staining solution contained 50mM NaP0 4 (pH7.00), 0.1 % (v/v) Triton X-100, 1 .04 mM X-Gluc and 0.5% (v/v) DMSO.
  • the plant tissue was subsequently washed in FAA for 1 0 minutes, followed with a wash in 50% ethanol. The tissue was dehydrated with 100% ethanol and hydrated slowly up to 100% h ⁇ O.
  • GUS expression patterns of soybean cultivar Carnia 2233 transformed with the CaMV 35S gene were thus observed in leaf ( Figure 1 A) and root ( Figure 1 B) tissue of the plant. Plants tested positive for the GUS gene up to the third generation.
  • soybean seed (Carnia 2233) was transformed using an Agrobacterium strain LBA4404 containing a p35S GUS INT gene.
  • a GUS-intron gene has the ability to discriminate between prokaryotic organisms such as Agrobacterium tumefaciens and eukaryotic organisms such as plants. Only plant tissue containing the GUS-INT gene turns blue in colour in association with a histochemical assay. Any Agrobacterium possibly still present in the plant tissue does not turn blue in colour.
  • Figure 2 shows transformed plant cells which have been stained with X-
  • soybean seed (Carnia)
  • P5CR gene was cloned in antisense orientation into plasmid HB1 01 pMA445, containing a heat inducible promoter which was subsequently triparental mated to Agrobacterium tumefaciens (Armitage, P. 1 988. Transformation of dicotyl plant cells using the Ti plasmid of Agrobacterium tumefaciens and Ri plasmid of A. rhizogenes. In: Plant Genetic Engineering and Gene Expression: A laboratory Manual. Draper, J ., Scott, R., Armitage, P. and Welden, R. (Eds) . Blackwell Scientific Publications, Oxford . pp69-1 60) .
  • the construct included a kanamycin resistant gene which can be used in screening of transformants. Putative transformed seed (with the P5CR gene) were tested for the presence of kanamycin resistance to indicate transformation. Third generation transformed soybean seed and untransformed soybean seed were tested for germination viability on agar plates supplemented with kanamycin. It was noted that all the seeds germinated on agar plates with Omg/f kanamycin (see Table 1 below) . It was noted that as the concentration of kanamycin in the agar plates was increased, the percentage of germinating plants decreased. Some of the seed initially started to germinate for a short period before dying off and some of the germinating plants showed deformities.
  • the untransformed control plants and the transformed plants were subjected to a variety of tests including different physiological techniques e.g. proline accumulation and anatomical techniques e.g . woodenbox screening to compare the untransformed control plants with the transformed plants under drought and osmotic stress.
  • Inactivation of the P5CR gene resulted in decreased proline synthesis.
  • the application of a mannitol stress test resulted in the untransformed control plants showing a significant increase in proline concentration whereas the antisense transformed plants displayed a significant decrease in proline concentration, indicating that the P5CR gene had been inactivated in the transformed plants and the transformed plants were unable to synthesise proline in response to the osmotic stress test.
  • Control plants and fourth generation antisense transformed plants were subjected to a woodenbox experiment. Seed was planted in a box and watered until all plants reached the six leaf stage. At this stage the promoter was activated and drought stress was applied. The control plants survived a six day drought stress significantly more than most of the antisense transformed plants which died . The results indicate that soybean plants were successfully transformed with the antisense construct, as the transformed plants were unable to survive a drought stress test with a lower copy number of the proline gene (see Figure 4) .
  • yo A measurement of the ability of a trapped excition to move an electron into electron transport.
  • ABS/RC The absorption flux of photons per photosystem II (PSII) reaction centre (RC) .
  • ABS/CSm The absorption flux of photons per PSII cross section (CS) .
  • TRo/RC The rate at which an excition is trapped by the RC.
  • ETo/RC The electron transport per RC.
  • Dlo/RC The energy flux, which is wasted per RC as heat or transfer to other systems.
  • RC/CSm An indication of the number of active RCAEs (density) per CS.
  • TRo/CSm The rate at which an exciton is trapped by the CS.
  • the antisense transformed plants were subjected to stress tests which resulted in the antisense transformed plants increasing active reaction centres, absorption, trapping and electron transport per cross section in compensating for stress imposed on the antisense transformed plants, whereas in contrast the sense transformed plants (see Figure 5A) when subjected to stress tests shut down certain reaction centres and there was a decrease in trapping, absorption and electron transport.
  • the sense transformed plants contain a higher P5CR copy number and are therefore able to use energy more efficiently than the antisense transformed plants having a lower P5CR copy number. It appears that the sense transformed plants are more drought tolerant whereas the antisense transformed plants are more drought sensitive. These tests indicated that transformation of Ibis was successful.
  • Lupinus a/bus seed (cultivar Esta) was transformed with Agrobacterium strain LBA4404 containing a p35S GUS INT gene.
  • Figure 6 shows transformed lupin leaves of initial transformed seed, together with first generation seed compared to control plants, leaves and seeds. The transformed leaves and seed demonstrate blue colouring representing GUS-gene activity. This indicates that lupin was successivefully transformed using the method according to the invention and that the GUS INT gene was successfully transferred at least to the first generation.
  • the methods in accordance with the invention resulted in a transformation success rate of approximately 35 % of the soybean seeds. This is a relatively high transformation success rate in that conventional techniques usually only have a transformation success rate of less than 5% .
  • the soybean seed By inserting a foreign gene or genes into a plasmid in Agrobacterium tumefaciens, the soybean seed may be transformed with the foreign gene. The foreign gene is then included in the cells of a soybean plant which grows from the transformed soybean seed and may then be inherited by its progeny.
  • the methods are relatively easy to carry out and relatively inexpensive compared to conventional transformation procedures and techniques. As no tissue culture steps are used in the methods according to the invention, it is believed that there will be little or no loss of genetic traits which would usually occur as a result of somatic mutations.
  • the methods in accordance with the invention can be used for transforming any suitable plant seed with genes of interest or agricultural usefulness, for example, drought resistant or disease resistant genes.
  • the methods in accordance with the invention can also be used for producing transgenic plants of other species where routine tissue culture procedures have not yet been established.

Abstract

La présente invention concerne un procédé d'obtention de graines transformées ou modifiées génétiquement qui consiste à mettre en contact des graines en germination avec un agent mouillant ou un tensioactif et avec une souche d'Agrobacterium afin de transformer ou de modifier génétiquement les graines. L'invention concerne aussi une composition de transformation comprenant, en mélange, une souche d'Agrobacterium et un agent mouillant ou un tensioactif.
PCT/IB1999/001676 1998-10-15 1999-10-14 Procede de transformation WO2000022149A1 (fr)

Priority Applications (4)

Application Number Priority Date Filing Date Title
AU59937/99A AU762964B2 (en) 1998-10-15 1999-10-14 Transformation process
EP99970425A EP1121452A1 (fr) 1998-10-15 1999-10-14 Procede de transformation
CA002346004A CA2346004A1 (fr) 1998-10-15 1999-10-14 Procede de transformation
BR9914572-3A BR9914572A (pt) 1998-10-15 1999-10-14 Processo de transformação

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
ZA989427 1998-10-15
ZA98/9427 1998-10-15

Publications (1)

Publication Number Publication Date
WO2000022149A1 true WO2000022149A1 (fr) 2000-04-20

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PCT/IB1999/001676 WO2000022149A1 (fr) 1998-10-15 1999-10-14 Procede de transformation

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EP (1) EP1121452A1 (fr)
AU (1) AU762964B2 (fr)
BR (1) BR9914572A (fr)
CA (1) CA2346004A1 (fr)
WO (1) WO2000022149A1 (fr)

Cited By (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2000037663A2 (fr) * 1998-12-23 2000-06-29 The Samuel Roberts Noble Foundation, Inc. Procede de transformation de vegetaux
EP1236801A2 (fr) * 2001-02-26 2002-09-04 The Agri-Biotechnology Research Center of Shanxi Méthode de transformation de plante par l'agrobacterium par le traitement de graines en germination
WO2003086053A1 (fr) * 2002-04-15 2003-10-23 Phytoculture Control Co., Ltd. Systeme de transformation de plantes
US20130157369A1 (en) * 2011-12-15 2013-06-20 Dow Agrosciences Llc Method for improved transformation using agrobacterium
US20140212387A1 (en) * 2011-05-27 2014-07-31 Bayer Intellectual Property Gmbh Liquid preparation for biological plant protection, method for producing it and use thereof

Citations (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
NL8801444A (nl) * 1988-06-06 1990-01-02 Solvay Werkwijze voor het genetisch transformeren van cellen van een multicellulair eukaryotisch organisme.
WO1997048814A2 (fr) * 1996-06-21 1997-12-24 Monsanto Company Procedes de production de ble fertile transforme stable faisant appel a une transformation mediee par l'agrobacterium et compositions derivees de ce dernier
WO1998037212A1 (fr) * 1997-02-20 1998-08-27 Plant Genetic Systems, N.V. Procede de transformation perfectionne pour vegetaux
WO1999014348A1 (fr) * 1997-09-12 1999-03-25 Performance Plants, Inc. TRANSFORMATION DE VEGETAUX $i(IN PLANTA)

Patent Citations (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
NL8801444A (nl) * 1988-06-06 1990-01-02 Solvay Werkwijze voor het genetisch transformeren van cellen van een multicellulair eukaryotisch organisme.
WO1997048814A2 (fr) * 1996-06-21 1997-12-24 Monsanto Company Procedes de production de ble fertile transforme stable faisant appel a une transformation mediee par l'agrobacterium et compositions derivees de ce dernier
WO1998037212A1 (fr) * 1997-02-20 1998-08-27 Plant Genetic Systems, N.V. Procede de transformation perfectionne pour vegetaux
WO1999014348A1 (fr) * 1997-09-12 1999-03-25 Performance Plants, Inc. TRANSFORMATION DE VEGETAUX $i(IN PLANTA)

Non-Patent Citations (1)

* Cited by examiner, † Cited by third party
Title
CLOUGH S J ET AL: "Floral dip: a simplified method for Agrobacterium-mediated transformation of Arabidopsis thaliana.", PLANT JOURNAL, (1998 DEC) 16 (6) 735-43. JOURNAL CODE: BRU., XP002132452 *

Cited By (8)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2000037663A2 (fr) * 1998-12-23 2000-06-29 The Samuel Roberts Noble Foundation, Inc. Procede de transformation de vegetaux
WO2000037663A3 (fr) * 1998-12-23 2000-11-09 Samuel Roberts Noble Found Inc Procede de transformation de vegetaux
EP1236801A2 (fr) * 2001-02-26 2002-09-04 The Agri-Biotechnology Research Center of Shanxi Méthode de transformation de plante par l'agrobacterium par le traitement de graines en germination
EP1236801A3 (fr) * 2001-02-26 2002-10-02 The Agri-Biotechnology Research Center of Shanxi Méthode de transformation de plante par l'agrobacterium par le traitement de graines en germination
WO2003086053A1 (fr) * 2002-04-15 2003-10-23 Phytoculture Control Co., Ltd. Systeme de transformation de plantes
US20140212387A1 (en) * 2011-05-27 2014-07-31 Bayer Intellectual Property Gmbh Liquid preparation for biological plant protection, method for producing it and use thereof
US9788551B2 (en) * 2011-05-27 2017-10-17 Bayer CropScience Biologies GmbH Liquid preparation for biological plant protection, method for producing it and use thereof
US20130157369A1 (en) * 2011-12-15 2013-06-20 Dow Agrosciences Llc Method for improved transformation using agrobacterium

Also Published As

Publication number Publication date
CA2346004A1 (fr) 2000-04-20
BR9914572A (pt) 2002-01-15
AU762964B2 (en) 2003-07-10
EP1121452A1 (fr) 2001-08-08
AU5993799A (en) 2000-05-01

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