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WO1991000358A1 - Procede d'introduction de molecules, notamment de matiere genetique, dans des cellules vegetales - Google Patents

Procede d'introduction de molecules, notamment de matiere genetique, dans des cellules vegetales Download PDF

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Publication number
WO1991000358A1
WO1991000358A1 PCT/DK1990/000166 DK9000166W WO9100358A1 WO 1991000358 A1 WO1991000358 A1 WO 1991000358A1 DK 9000166 W DK9000166 W DK 9000166W WO 9100358 A1 WO9100358 A1 WO 9100358A1
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WO
WIPO (PCT)
Prior art keywords
plant cells
cells
molecules
genetic material
ultrasound treatment
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/DK1990/000166
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English (en)
Inventor
Morten JØRSBOE
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International N&H Denmark ApS
Original Assignee
Danisco AS
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Filing date
Publication date
Application filed by Danisco AS filed Critical Danisco AS
Publication of WO1991000358A1 publication Critical patent/WO1991000358A1/fr
Anticipated expiration legal-status Critical
Ceased legal-status Critical Current

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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
    • C12N15/8206—Methods for introducing genetic material into plant cells, e.g. DNA, RNA, stable or transient incorporation, tissue culture methods adapted for transformation by physical or chemical, i.e. non-biological, means, e.g. electroporation, PEG mediated
    • 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
    • C12N13/00—Treatment of microorganisms or enzymes with electrical or wave energy, e.g. magnetism, sonic waves
    • 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
    • C12N15/64—General methods for preparing the vector, for introducing it into the cell or for selecting the vector-containing host
    • 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
    • C12N15/87—Introduction of foreign genetic material using processes not otherwise provided for, e.g. co-transformation

Definitions

  • the present invention relates to a method for introducing molecules / particularly genetic material, into intact plant cells.
  • introducing molecules such as genetic material, for instance plasmid DNA, RNA, vira or fragments thereof, into animal cells and protoplasts.
  • Such known methods include inter alia chemical methods, electroporation and microinjection. It has been shown that such known methods could be implemented in both transient and stable transformation. In general all methods disclosed for obtaining transient expression have turned out to be usable for obtaining stable transformation.
  • polyethylene glycol polyethylene glycol
  • PEG polyethylene glycol
  • polyethylene glycol usually 40% polyethylene glycol 6000, is added to plant protoplasts and genetic material ( rens et al., Nature, 296, 72-74, 1982).
  • Polyethylene i ine or poly-L-ornithin can be used correspondingly.
  • Electroporation In this case a suspension of animal cells or plant protoplasts is exposed to a short electric pulse of high field strength in the presence of genetic material (Neuman et al., EMBO J., 1, 841-845, 1982; Fromm et al., Nature, 319, 791-793, 1986).
  • WO Publication No. 89/02464 discloses the transformation of animal cells with DNA fragments by subjecting the cells to ultrasound treatment sufficient to traumatize the cells without killing them. Due to the similarities between animal cells and plant protoplasts mentioned above it has to be considered obvious that this method can also be used to introduce DNA fragments in plant protoplasts.
  • the object of the present invention is to provide a method for introducing molecules, particularly genetic material, into intact plant cells, in an inexpensive and effective way.
  • the method according to the invention is a useful alternative to previously known introduction methods.
  • the method provides a new approach involving only moderate costs, said method being carried out quickly and simply. Based on introduction experiments already conducted it can be predicted that the method will also turn out to be superior when used for a great number of other biological materials, where known methods are insufficient.
  • the plant cells are subjected to an ultrasound treatment considerably milder than the ultrasound treatment used for homogenizing or lysing cells. It is important that the treatment is suitably mild so that a sufficient number of plant cells remain viable.
  • the plant cell suspension can advantageously be exposed to ultrasonic waves of a frequency range of from 5 kHz to 10 MHz, particularly from 10 to 100 kHz, and an electric output power (i.e.
  • Examples of molecules introducable into plant cells by the method according to the invention include DNA, plasmid DNA, RNA, vira, proteins, lipids, pharmaceutical compositions, small molecules, organelles or fragments of such materials.
  • the method according to the invention has advantageously been used to introduce molecules into cells of sugar beet and tobacco plants.
  • a particularly suitable medium for plant cells and molecules during ultrasound treatment is CPW comprising from 21 to 28% sucrose.
  • the mild ultrasound treatment of the method according to the invention is advantageously carried out using a sound-emitting means having an acute point, said means being only immersed in the upper portion of the medium.
  • the method according to the invention is advantageously carried out by using a device comprising a sound-emitting means capable of emitting ultrasound in a medium for a period of up to 10.000 ms.
  • a device comprising a sound-emitting means capable of emitting ultrasound in a medium for a period of up to 10.000 ms.
  • Such a device is advantageously provided in such a way that it emits ultrasonic waves of a frequency in the range of from 10 to 100 kHz.
  • the device according to the invention can be equipped so that the electric power supplied to the sound-emitting means can be adjusted to any given value in the range of from 5 to 300 W and so that the duration of the ultrasou ⁇ treatment can be adjusted within a range of from 100 to 10.000 ms.
  • the sound source of the device is advantageously formed in such a way that said device can be immersed in a medium in a suitable vessel, for instance an Eppendorf tube.
  • a suitable vessel for instance an Eppendorf tube.
  • the present invention has been developed using a device for lysing cells, said device being commercially available from Branson, Eagle Road, Danbury, Connecticut, USA, under the name of Sonifier B 15.
  • the device can emit ultrasound of a frequency of 20 kHz.
  • the electric power values expressed in watts mentioned in the present specification with claims are electric output powers as read from the output control of the electric power supply unit.
  • the ultrasound-emitting means was immersed for approx. 2 to 3 mm measured from the surface.
  • Preliminary experiments with a calorimeter have shown that the ultrasonic power imparted to the liquid during experimental procedure is approx. 5 to 10% of the given electric output power.
  • the genetic material in question is especially DNA or fragments thereof, such as plasmid DNA.
  • the method is also suitable for introducing RNA or fragments thereof as well as vira, for instance for pathological tests.
  • the method according to the invention is probably also suitable for introducing proteins, lipids, pharmaceutical compositions, small molecules as well as organelles and virus particles into cells.
  • the medium can be any suitable, conventional medium for cells or protoplasts.
  • the method according to the invention uses a technique presumbably resulting in a temporary moderate weakening of both the cell membrane and the cell wall, which is shown in intact plant cells in the following examples. The examples thus show that plasmid DNA can penetrate plant cells having been treated with ultrasound.
  • a cell suspension culture of sugar beets (Beta vulgaris L.) of the genotype Ml (available from DANISCO A/S, Copenhagen, Denmark) is prepared from callus obtained from embryos.
  • the cell suspension is cultured in darkness at 25°C on a rotary shaking table.
  • the cells are maintained by sub-culturing in a medium according to Murashige and Skoog (Physiol. Plant., 15, 473-497, 1962) to which 5.7 ⁇ M indol acetic acid and 4.4 ⁇ M benzyladenine was added.
  • CPW is an aqueous solution of a mixture of inorganic salts comprising i.a. approx. 10 mM Ca , described by Frearson et al., (Dev. Biol., 33, 130-137, 1973).
  • a cell suspension for ultrasound treatment is prepared by removing cells 3 to 4 days after sub-culturing and washing them twice with CPW 13S (i.e. CPW containing 13% sorbitol), finally suspending said cells in CPW 13S at a ratio of 1 part by volume cells to 4 parts by volume CPW 13S. Then plasmid DNA is added to the suspension in 0.35 ml CPW containing 21% sucrose and plant cells (500.000/ml) in an Eppendorf tube, the final plasmid concentration being 45 ⁇ g/ml.
  • CPW 13S i.e. CPW containing 13% sorbitol
  • the plasmid DNA used is a plasmid coding for the marker enzyme chloramphenicol-acetyltransferase (CAT) , in this case the plasmid pCaMVCN having the code 27-4909, available from Pharmacia LKB Biotechnology, Uppsala, Sweden.
  • CAT chloramphenicol-acetyltransferase
  • the cells treated by ultrasound are then transferred to petri dishes containing a MS-medium (Murashige and Skoog, cf. above) and are incubated for 2 days at 23°C.
  • the presence of CAT-activity is shown by adding ⁇ C-marked chloramphenicol to an extract obtained from the treated cells, whereupon the sample is heated to 60°C for 6 min. After cooling acetyl-coenzyme A is added to the sample, the final concentration of acetyl-coenzyme A being 0.71 mM.
  • the introduction of plasmids is assessed by measuring the percentage transformation of chloramphenicol (CA).
  • the method used is a modification of the method described by Gorman et al. (Mol. Cell. Biol., 2, 1044-1051, 1982).
  • the present example illustrates the introduction of plasmid DNA into intact sugar beet cells of the genotype Ml.
  • the suspension culture of sugar beet cells is maintained by sub-culturing as described above and is treated with ultrasound, cultured and analysed in the manner described.
  • the results of the measured CAT-activity appear from Table 1.
  • the present example illustrates the introduction of plasmid DNA into intact tobacco cells.
  • a suspension culture of tobacco cells is maintained by sub-culturing in the manner described above for sub-culturing of cell suspensions with sugar beets, the culture medium, however, being a medium according to Murashige and Skoog (Physiol. Plant. 15, 473-497, 1962), to which were added 0.2 mg/1 2,4-dichlorophenoxy acetic acid, 0.1 mg/1 kinetin, 0.9 mg/1 thiamin hydrochloride and 0.2 g/1 H2PO4, pH 6.0.
  • the cells are removed 3 to 4 days after sub-culturing and washed twice with CPW 13S (i.e. CPW containing 13% sorbitol), finally suspending said cells in CPW 13S at a ratio of 1 part by volume cells to 4 parts by volume CPW 13S.
  • Samples of 0.35 ml each are taken out and the plasmid pCaMVCN is added to each sample, the final plasmid concentration being 100 ⁇ g/ml.
  • the cells are then subjected to ultrasound treatment under the conditions appearing from Table 2. After culturing for 2 days in the above-mentioned medium the cells are extracted and their CAT-activity is measured. The results appear from Table 2.
  • the method according to the invention can be used to introduce molecules into intact plant cells.

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  • Health & Medical Sciences (AREA)
  • Genetics & Genomics (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Engineering & Computer Science (AREA)
  • Biomedical Technology (AREA)
  • Zoology (AREA)
  • Bioinformatics & Cheminformatics (AREA)
  • Biotechnology (AREA)
  • Organic Chemistry (AREA)
  • Chemical & Material Sciences (AREA)
  • Wood Science & Technology (AREA)
  • General Engineering & Computer Science (AREA)
  • Microbiology (AREA)
  • Molecular Biology (AREA)
  • Biochemistry (AREA)
  • General Health & Medical Sciences (AREA)
  • Plant Pathology (AREA)
  • Biophysics (AREA)
  • Physics & Mathematics (AREA)
  • Cell Biology (AREA)
  • Micro-Organisms Or Cultivation Processes Thereof (AREA)
  • Breeding Of Plants And Reproduction By Means Of Culturing (AREA)
  • Preparation Of Compounds By Using Micro-Organisms (AREA)

Abstract

Procédé d'introduction de molécules, notamment de matière génétique dans des cellules végétales intactes, consistant à soumettre un milieu contenant lesdites céllules végétales ainsi que lesdites molécules à un traitement ultrasonique doux.
PCT/DK1990/000166 1989-06-29 1990-06-28 Procede d'introduction de molecules, notamment de matiere genetique, dans des cellules vegetales Ceased WO1991000358A1 (fr)

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
DK325189A DK168302B1 (da) 1989-06-29 1989-06-29 Fremgangsmåde til indføring af molekyler, især genetisk materiale i planteceller
DK3251/89 1989-06-29

Publications (1)

Publication Number Publication Date
WO1991000358A1 true WO1991000358A1 (fr) 1991-01-10

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PCT/DK1990/000166 Ceased WO1991000358A1 (fr) 1989-06-29 1990-06-28 Procede d'introduction de molecules, notamment de matiere genetique, dans des cellules vegetales

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EP (1) EP0480971A1 (fr)
JP (1) JPH05500304A (fr)
AU (1) AU645260B2 (fr)
DK (1) DK168302B1 (fr)
WO (1) WO1991000358A1 (fr)

Cited By (38)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US5712134A (en) * 1990-05-09 1998-01-27 The Biological Research Center Of The Hungarian Academy Of Sciences Method of producing a cell carrying an excess of mammalian centromeres
US5851984A (en) * 1996-08-16 1998-12-22 Genentech, Inc. Method of enhancing proliferation or differentiation of hematopoietic stem cells using Wnt polypeptides
US5990281A (en) * 1996-09-30 1999-11-23 Genentech, Inc. Vertebrate smoothened proteins
US6025155A (en) * 1996-04-10 2000-02-15 Chromos Molecular Systems, Inc. Artificial chromosomes, uses thereof and methods for preparing artificial chromosomes
WO2000008195A1 (fr) * 1998-07-31 2000-02-17 Dornier Medtech Holding International Gmbh Procede et dispositif de transfert d'oligonucleotides dans des cellules
US6030945A (en) * 1996-01-09 2000-02-29 Genentech, Inc. Apo-2 ligand
US6077697A (en) * 1996-04-10 2000-06-20 Chromos Molecular Systems, Inc. Artificial chromosomes, uses thereof and methods for preparing artificial chromosomes
US6114603A (en) * 1998-03-27 2000-09-05 John Innes Center Genetic engineering of sugarbeet plants
US6159462A (en) * 1996-08-16 2000-12-12 Genentech, Inc. Uses of Wnt polypeptides
US6195936B1 (en) * 1999-02-22 2001-03-06 University Of Iowa Research Foundation Method for uptake of a substance into a seed
DE19962904A1 (de) * 1999-12-23 2001-08-09 Dornier Medizintechnik Verfahren zum Transfer von Molekülen in Zellen und Vorrichtung zur Durchführung des Verfahrens
US6291643B1 (en) 1997-06-05 2001-09-18 Board Of Reports, The University Of Texas System Apaf-1 an activator of caspase-3
EP0904362A4 (fr) * 1996-03-01 2001-09-26 Ohio State Res Found Procede de transformation de tissus de plantes
US6342369B1 (en) 1997-05-15 2002-01-29 Genentech, Inc. Apo-2-receptor
US6462176B1 (en) 1996-09-23 2002-10-08 Genentech, Inc. Apo-3 polypeptide
US6469144B1 (en) 1996-04-01 2002-10-22 Genentech, Inc. Apo-2LI and Apo-3 polypeptides
EP1382679A2 (fr) 1995-09-08 2004-01-21 Genentech, Inc. Antagonists du proteine apparentée à facteur de croissance endothélial vasculaire (VRP)
US6740739B1 (en) 1998-01-15 2004-05-25 Genentech, Inc. Substitutional variants of APO-2 ligand
US6746668B2 (en) 1996-01-09 2004-06-08 Genentech, Inc. Apo-2 ligand
EP1666052A1 (fr) 2000-02-16 2006-06-07 Genentech, Inc. Utilisation d'agonistes ou d'antagonistes pour moduler l'activite de molecules associees au tnf
US7267659B2 (en) 2002-05-24 2007-09-11 Dornier Medtech Systems Gmbh Method and apparatus for transferring medically effective substances into cells
EP1958965A2 (fr) 1997-08-25 2008-08-20 Genentech, Inc. Anticorps agonistes pour un récepteur musk, et leurs utilisations thérapeutiques
GB2452543A (en) * 2007-09-07 2009-03-11 Wei Huang Nucleic acid transfer techniques
EP2083079A1 (fr) 1997-06-18 2009-07-29 Genentech, Inc. Apo-2DcR
EP2233149A1 (fr) 2007-10-16 2010-09-29 ZymoGenetics, Inc. Combinaison de l'inhibition du BLYS et d'un agent anti-CD20 pour le traitement des maladies auto-immunes
EP2272868A2 (fr) 2003-06-05 2011-01-12 Genentech, Inc. Thérapie de combinaison pour des désordres de cellules B
WO2011019619A1 (fr) 2009-08-11 2011-02-17 Genentech, Inc. Production de protéines dans des milieux de culture cellulaire sans glutamine
EP2311960A2 (fr) 2001-08-29 2011-04-20 Genentech, Inc. Acides nucléiques Bv8 et polypeptides avec activité mitogénique
EP2311956A1 (fr) 1999-06-28 2011-04-20 Genentech, Inc. Methodes de production de ligand à APO-2 utilisant des ions métalliques divalents
EP2322165A1 (fr) 2001-11-13 2011-05-18 Genentech, Inc. Compositions à base de Apo2 ligand/TRAIL
EP2348043A1 (fr) 2001-10-02 2011-07-27 Genentech, Inc. Variantes du ligand APO-2 et leurs utilisations
EP2390256A1 (fr) 2001-05-30 2011-11-30 Agrisoma, Inc. Chromosomes artificiels de plantes, leurs utilisations et leurs procédés de préparation
EP2500032A1 (fr) 2002-06-24 2012-09-19 Genentech, Inc. Variantes du ligand/Trail APO-2 et leurs utilisations
WO2012151317A1 (fr) 2011-05-03 2012-11-08 Genentech, Inc. Agents d'interruption vasculaire et utilisations associées
EP2526960A1 (fr) 2003-03-12 2012-11-28 Genentech, Inc. Utilisation de BV8 et/ou de EG-VEGF pour promouvoir l'hématopoièse
US8669085B2 (en) 2009-02-05 2014-03-11 Ut-Battelle, Llc Transformation of gram positive bacteria by sonoporation
US9060915B2 (en) 2004-12-15 2015-06-23 Dornier MedTech Systems, GmbH Methods for improving cell therapy and tissue regeneration in patients with cardiovascular diseases by means of shockwaves
WO2016182336A1 (fr) * 2015-05-11 2016-11-17 대한민국(농촌진흥청장) Procédé pour retarder la maturation de fruits à l'aide d'ondes sonores

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EP0290395A2 (fr) * 1987-05-05 1988-11-09 Sandoz Ag Transformation de tissus de plantes
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Cited By (62)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US5891691A (en) * 1990-05-09 1999-04-06 The Biological Research Center Of The Hungarian Academy Of Sciences Method of producing a cell carrying an excess of mammalian centromeres and the cell line carrying an excess of mammalian centromeres
US5712134A (en) * 1990-05-09 1998-01-27 The Biological Research Center Of The Hungarian Academy Of Sciences Method of producing a cell carrying an excess of mammalian centromeres
EP1382679A2 (fr) 1995-09-08 2004-01-21 Genentech, Inc. Antagonists du proteine apparentée à facteur de croissance endothélial vasculaire (VRP)
US6030945A (en) * 1996-01-09 2000-02-29 Genentech, Inc. Apo-2 ligand
US7285533B2 (en) 1996-01-09 2007-10-23 Genentech, Inc. Apo-2 ligand
US6998116B1 (en) 1996-01-09 2006-02-14 Genentech, Inc. Apo-2 ligand
US6746668B2 (en) 1996-01-09 2004-06-08 Genentech, Inc. Apo-2 ligand
EP0904362A4 (fr) * 1996-03-01 2001-09-26 Ohio State Res Found Procede de transformation de tissus de plantes
US6469144B1 (en) 1996-04-01 2002-10-22 Genentech, Inc. Apo-2LI and Apo-3 polypeptides
EP2314708A1 (fr) 1996-04-10 2011-04-27 Chromos Molecular Systems, Inc. Chromosomes artificiels, leurs utilisations et leurs procédés de préparation
US6077697A (en) * 1996-04-10 2000-06-20 Chromos Molecular Systems, Inc. Artificial chromosomes, uses thereof and methods for preparing artificial chromosomes
US6025155A (en) * 1996-04-10 2000-02-15 Chromos Molecular Systems, Inc. Artificial chromosomes, uses thereof and methods for preparing artificial chromosomes
US6743967B2 (en) 1996-04-10 2004-06-01 Chromos Molecular Systems Inc. Artificial chromosomes, uses thereof and methods for preparing artificial chromosomes
US6159462A (en) * 1996-08-16 2000-12-12 Genentech, Inc. Uses of Wnt polypeptides
US5851984A (en) * 1996-08-16 1998-12-22 Genentech, Inc. Method of enhancing proliferation or differentiation of hematopoietic stem cells using Wnt polypeptides
US6462176B1 (en) 1996-09-23 2002-10-08 Genentech, Inc. Apo-3 polypeptide
US5990281A (en) * 1996-09-30 1999-11-23 Genentech, Inc. Vertebrate smoothened proteins
US6407216B1 (en) 1996-09-30 2002-06-18 Genentech, Inc. Vertebrate smoothened antibodies
US6342369B1 (en) 1997-05-15 2002-01-29 Genentech, Inc. Apo-2-receptor
US8092799B2 (en) 1997-05-15 2012-01-10 Genentech, Inc. Antibodies to Apo-2 receptor polypeptides
US7749755B2 (en) 1997-05-15 2010-07-06 Genentech, Inc. Apo-2 receptor polynucleotides
US7939631B2 (en) 1997-05-15 2011-05-10 Genentech, Inc. APO-2 receptor polypeptides
US7750118B2 (en) 1997-05-15 2010-07-06 Genentech, Inc. Apo-2 receptor polypeptides
US7595046B2 (en) 1997-05-15 2009-09-29 Genentech, Inc. Treatment of cancer using anti-Apo-2 antibodies
US7807153B2 (en) 1997-05-15 2010-10-05 Genentech, Inc. Apo-2 receptor agonist antibodies
US7314619B2 (en) 1997-05-15 2008-01-01 Genentech, Inc. Inducing apoptosis using anti-Apo-2 antibodies
US6291643B1 (en) 1997-06-05 2001-09-18 Board Of Reports, The University Of Texas System Apaf-1 an activator of caspase-3
EP2083079A1 (fr) 1997-06-18 2009-07-29 Genentech, Inc. Apo-2DcR
EP1958965A2 (fr) 1997-08-25 2008-08-20 Genentech, Inc. Anticorps agonistes pour un récepteur musk, et leurs utilisations thérapeutiques
EP2017341A2 (fr) 1998-01-15 2009-01-21 Genentech, Inc. Ligand Apo-2
US6740739B1 (en) 1998-01-15 2004-05-25 Genentech, Inc. Substitutional variants of APO-2 ligand
US6114603A (en) * 1998-03-27 2000-09-05 John Innes Center Genetic engineering of sugarbeet plants
WO2000008195A1 (fr) * 1998-07-31 2000-02-17 Dornier Medtech Holding International Gmbh Procede et dispositif de transfert d'oligonucleotides dans des cellules
DE19834612A1 (de) * 1998-07-31 2000-02-24 Dornier Medtech Holding Int Gmbh Verfahren zum intrazellulären Transfer von Oligonukleotiden und Vorrichtung zur Durchführung desselben
US6195936B1 (en) * 1999-02-22 2001-03-06 University Of Iowa Research Foundation Method for uptake of a substance into a seed
EP2339003A2 (fr) 1999-06-28 2011-06-29 Genentech, Inc. Polypeptides variants substitutionnels du ligand APO-2
EP2311956A1 (fr) 1999-06-28 2011-04-20 Genentech, Inc. Methodes de production de ligand à APO-2 utilisant des ions métalliques divalents
DE19962904A1 (de) * 1999-12-23 2001-08-09 Dornier Medizintechnik Verfahren zum Transfer von Molekülen in Zellen und Vorrichtung zur Durchführung des Verfahrens
EP1666052A1 (fr) 2000-02-16 2006-06-07 Genentech, Inc. Utilisation d'agonistes ou d'antagonistes pour moduler l'activite de molecules associees au tnf
EP2390256A1 (fr) 2001-05-30 2011-11-30 Agrisoma, Inc. Chromosomes artificiels de plantes, leurs utilisations et leurs procédés de préparation
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DK325189A (da) 1990-12-30
AU645260B2 (en) 1994-01-13
AU6043290A (en) 1991-01-17
JPH05500304A (ja) 1993-01-28
EP0480971A1 (fr) 1992-04-22
DK325189D0 (da) 1989-06-29
DK168302B1 (da) 1994-03-07

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