EP0169230A1 - Procede de production de cultures de tissus mixotrophiques et de plantes resistant aux herbicides - Google Patents

Procede de production de cultures de tissus mixotrophiques et de plantes resistant aux herbicides

Info

Publication number
EP0169230A1
EP0169230A1 EP85900726A EP85900726A EP0169230A1 EP 0169230 A1 EP0169230 A1 EP 0169230A1 EP 85900726 A EP85900726 A EP 85900726A EP 85900726 A EP85900726 A EP 85900726A EP 0169230 A1 EP0169230 A1 EP 0169230A1
Authority
EP
European Patent Office
Prior art keywords
photosynthesis
sugar
culture medium
cultivation
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.)
Withdrawn
Application number
EP85900726A
Other languages
German (de)
English (en)
Inventor
Agnes Cseplo
Pál MALIGA
Péter MEDGYESY
László MENCZEL
László MARTON
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
MTA SZEGEDI BIOLOGIAI KOEZPONTJA
MTA Szegedi Biologiai Kozpontja
Magyar Tudomanyos Akademia Szegedi Biologiai Kozpont
Original Assignee
MTA SZEGEDI BIOLOGIAI KOEZPONTJA
MTA Szegedi Biologiai Kozpontja
Magyar Tudomanyos Akademia Szegedi Biologiai Kozpont
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 MTA SZEGEDI BIOLOGIAI KOEZPONTJA, MTA Szegedi Biologiai Kozpontja, Magyar Tudomanyos Akademia Szegedi Biologiai Kozpont filed Critical MTA SZEGEDI BIOLOGIAI KOEZPONTJA
Publication of EP0169230A1 publication Critical patent/EP0169230A1/fr
Withdrawn legal-status Critical Current

Links

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
    • C12N15/01—Preparation of mutants without inserting foreign genetic material therein; Screening processes therefor
    • A—HUMAN NECESSITIES
    • A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
    • A01H—NEW PLANTS OR NON-TRANSGENIC PROCESSES FOR OBTAINING THEM; PLANT REPRODUCTION BY TISSUE CULTURE TECHNIQUES
    • A01H4/00—Plant reproduction by tissue culture techniques ; Tissue culture techniques therefor
    • 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
    • C12N5/00—Undifferentiated human, animal or plant cells, e.g. cell lines; Tissues; Cultivation or maintenance thereof; Culture media therefor
    • C12N5/0018—Culture media for cell or tissue culture
    • C12N5/0025—Culture media for plant cell or plant tissue culture
    • 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
    • C12N5/00—Undifferentiated human, animal or plant cells, e.g. cell lines; Tissues; Cultivation or maintenance thereof; Culture media therefor
    • C12N5/04—Plant cells or tissues

Definitions

  • Tissue cultures are sterile, homogeneous, can be maintained both in a stationary and exceptionally growing phase, and are suitable for micro-testing. The problems of penetration and transport as associated with the whole plant can be avoided with the use of cell cultures.
  • tissue culture techniques The main problem with tissue culture techniques is that the tissue cultures of plants are heterotrophic; plant cells cannot grow in tissue cultures without sugar. Even the green tissue cultures are not capable of supporting their growth from photosynthesis (their photosynthetic activity is very low). Photosynthesis-inhibiting herbicides are therefore in-effective in tissue-cultures under standard laboratory conditions (a culture medium containing 3 % sugar and a light intensity of. 1000 to 2000 Ix). More particularly, in the effective concentrations the herbicides also inhibit the growth of cultures grown in the dark; the effect observed is not the primary photosynthesis inhibiting activity.
  • the invention relates to a process for the preparation of raixotrophic tissue cultures induced from seeds, plant parts, protoplasts or single cells on a known culture medium containing less than 1 % sugar which is capable of the needed amount of photosynthesis for selection.
  • raixotrophic tissue culture is used to refer to cultures in which the growth can substantially be attributed to both sugar content and light conditions of the culture medium.
  • seed When preparing the cultures, one can start from any plant material conventionally used for this purpose. It should be noted that the term "seed" referred to as a possible starting material includes the embryo as well.
  • a plant part for example, leaves, stems, roots, blooms, etc. or part thereof can be used.
  • Protoplasts are plant cells without a cell wall which are referred to from herein as to include the protoplasts modified by fusion or other cell genetic methods.
  • the single cells are plant cells having a cell wall, including pollen.
  • sugars accessible to the plants e.g., saccharose and glucose (Plant Sci. Letters, 21, 209-214 (1981) 7.) .
  • the raixotrophic tissue cultures are preferably prepared by starting from tissue cultures prepared from sterile leaf or stem parts, on a known culture medium having a normal sugar concentration. More specifically, on a culture medium containing approximately 1 % sugar, callus formation is induced under a light intensity of approximately 1000 Ix at a temperature of approximately 25 After several weeks of cultivation inoculae selected from the calli are placed on a culture medium having a low (less than 1 %) sugar concentration and cultivation is continued under light intensity of about
  • Cultivation is performed on a known culture medium containing 0.2 to 0.8 % , more preferably 0.2 to 0.5 %, most preferably 0.2 to 0.3 % of sugar.
  • a known culture medium containing 0.2 to 0.8 % , more preferably 0.2 to 0.5 %, most preferably 0.2 to 0.3 % of sugar.
  • the process can be carried out under conventional, sterile laboratory conditions, i.e. under approximately 1000 Ix of continuous light at about 25 oC.
  • the process is equally suitable for the cultivation of tissues capable or uncapable of autotrophic growth.
  • the raixotrophic tissue cultures obtained proved suitable for testing the primary effects of photosynthesis-inhibiting herbicides including fading due to photodestruction.
  • tissue cultures can be used to distinguish the effect of photosynthesis-inhibiting herbicides from the effect of other types of herbicides. Thus, it can be determined with a very simple method if a given herbicide exerts its activity via inhibition of photosynthesis and in an affirmative case if it has a secondary effect at higher concentrations.
  • the mixotrophic tissue cultures can be used for the production of herbicide-resistant plants which is the most important achievement of the invention.
  • tissue cultures prepared by the process, according to the invention are suitable for testing the effect of a large variety of herbicides such as urea triazine, uracyl, diphenyl ether, pyridazine, hydroxybenzonitrile, nitrophenol, tridiazine, bispyridine, amide, anilide, triazole, thiocarbamate, glycine and chlorosulfurone and for the production of plants which are resistant to photosynthesis inhibiting herbicides.
  • herbicides such as urea triazine, uracyl, diphenyl ether, pyridazine, hydroxybenzonitrile, nitrophenol, tridiazine, bispyridine, amide, anilide, triazole, thiocarbamate, glycine and chlorosulfurone
  • the calli cultivated under a defined light intensity, on a culture medium having a low sugar concentration fade under the effect of photosynthesis inhibiting herbicides; except for herbicide-resistant calli which can easily be selected on the basis of this property.
  • the selection method according to the invention is equally suitable for the production of mono- and dicotyledonous herbicide-resistant plants.
  • plants illustrated by the Examples include practically all cultivated plants such as cereals (corns), maize, tomato, potato, sugar beet, sun-flower, soya, cotton, grapes, etc. Most of our tests have, however, been performed on tobacco plants since tobacco plant is the most widely used model plant for such kind of experiments.
  • resistancy can be transplanted from resistant plant species into sensitive plants., preferably by protoplast fusion.
  • Herbicide-resistance is preferably tested by protoplast technique since this method is suitable for testing the most cells in a time unit. In a Petri dish only about 9 calli can be tested, whereas by the protoplast method about 1000 colonies can be examined simultaneously.
  • a further advantage of the colonies which originate from protoplasts is that they are derived from a single cell therefore, they are much more homogeneous than the calli of multiple cell origin which are mixed genetically. Accordingly, the most advantageous way to produce resistant plants is the cultivation in cultures of protoplast origin.
  • Example 1 Example 1
  • saccharose 3 % (w/v) agar 0.8 % (w/v) 1 1 of culture medium is prepared by admixing 250 ml of the stock-solution RM I, 5 ml of the stock-solution RM II and 10 ml of the stock-solution RM III.
  • RM-salts Components RM I, RM II and RM III together are called RM-salts (Murashige et al. : Physiol. Plant 15, 473-497 (1982). Plants were cultivated under a light intensity of 1000 1x and a relative humidity of 70 % at 25 °C.
  • RM-salts (RM I + RM II + RM III) in the above proportions, i.e. 250 ml of RM I + 5.0 ml of RM II + 10 ml of RM III for one 1 of culture medium; inosite 100 mg/1 thiamine 1 mg/1 naphthylacetic acid 0.1 mg/1 benzyladenine 1.0 mg/1 saccharose 1 % (w/v)
  • the controls are cultures grown under the same conditions but without light.
  • the growth rate is satisfactory for experimental purposes, for example on a culture medium containing 0.3 % of saccharose about half of the weight measured after growth on a culture medium containing 2.0 % of saccharose is obtained.
  • the experimental data clearly show that on culture media having a low saccharose concentration the calli, although require an external saccharose supply for their growth, show a substantial photosynthesis-dependant growth, i.e. are mixotrophic.
  • Example 5 Regeneration of plants resistant to photosynthesis-inhibiting herbicides.
  • 1 g of sterile Nicotiana plumbaginifolia leaves was cut to about 1 mm pieces and were sterilized in 10 ml of a filter sterile enzyme solution in dark at a temperature of 25 oC without shaking. The incubation time was 18 hours.
  • KCL 400 pH 5.6 The suspension obtained was centrifuged again (50 g, 2 minutes).
  • the protoplasts collected from the bottom of the centrifuge tube were cultivated in a K3 nutrient solution (1 x 10 5 protoplasts/ml) under illumination ( 100 1x, 25 oC).
  • the cultures were diluted with K3 nutrient solutions having gradually decreasing glucose concentrations (0.4, 0.3, 0.2, 0.1 M) every 7 to 10th days until a 30-40 times final dilution.
  • Example 1 About 6 weeks after starting the cultivation the cell colonies were added to a culture medium containing 0.3 % of saccharose and 0.4 M of terbutryne, and otherwise having the same composition as given in Example 1 ( 10-fold dilution which in the end corresponded to a colony density of about 1000 colonies/Petri dish having a diameter of 10 cm).
  • the colonies were cultured at 28 oC in a light intensity of 1500 1x. After 1 to 2 months the colonies capable of greening were isolated and grown to calli on the same medium. From the calli obtained plants were regenerated on a culture medium containing 2 % of saccharose, no terbutryne and otherwise having the same composition as the culture medium used for the selection. After a one-month growth, sprouts were appearing which were rooted and cultivated on a culture medium described in Example 1. Cultivation was continued until 10 cm plants were obtained. Small pieces of the leaves of 10 cm plants were placed on a culture medium containing 0 3 % of saccharose and 0.4 M of terbutyne but otherwise having the same composition as given in Example l.
  • the test was repeated using methobrorauron as a herbicide in place of terbutryne. There could be isolated resistant cell lines also in this case and the resistance was expressed in the regenerated plants as well. The frequency of the appearance of resistant cell lines was about 1/10 5 colonies.
  • the herbicide sensitivities of TBR plants and wild-type N. plumbaginifolia were compared in isolated chloroplasts by measuring the inhibition of the photosynthetic electron transport from water to ferricyanide (Hill-reaction) in the presence of various herbicides.
  • the half inhibitory concentrations (I 50 ) and the relative resistances (R/S) are shown in Table 6.
  • RF 1 resistance in the TBR plants was inherited maternally.
  • Chloroplasts were isolated as described by Reeves and
  • Example 6 Transmission of herbicide-resistance between plant species by protoplast fusion
  • atrazine-resistant chloro plasts were transplanted from a mutant weed (Solanum nigrum) into tobacco (Nicotiana plumbaginifolia) and potato, respectively.
  • the transmission was carried out by the process described in Example 5 for the cultivation of protoplasts by a chloroplast transfer method based on protoplast fusion (Menczel et al. : Genetics, 100, 487-95 (1982); Medgyesy et al. : Molec. Gen. Genet. 179, 693-698 (1980)).
  • This procedure we made use of the possibility of the selection of cell lines containing resistant chloroplasts on low sugar concentrations.
  • the appearance of the resistance in the cultivated plants was controlled as disclosed in Example 5.

Landscapes

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

Abstract

Procédé de préparation de cultures de tissus mixotrophiques en les cultivant dans une culture de tissus capable de photosynthèse ou contenant des parties capables de photosynthèse, préparée à partir de semences, d'une partie quelconque de la plante, de protoplastes ou d'une cellule unique sur un bouillon de culture connu contenant du sucre et exposé à un éclairage. Le procédé se caractérise par le fait que la culture est exécutée sur un bouillon de culture contenant moins de 1% de sucre, par exemple du saccharose ou du glucose. Est également décrit un nouveau procédé de sélection et de reproduction de plantes résistant aux herbicides inhibant la photosynthèse. Pour la sélection de plantes résistant aux herbicides ou utilise le bouillon de culture susmentionné.
EP85900726A 1984-01-16 1985-01-16 Procede de production de cultures de tissus mixotrophiques et de plantes resistant aux herbicides Withdrawn EP0169230A1 (fr)

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
HU84152A HU190751B (en) 1984-01-16 1984-01-16 Process for producing mixotroph tissue-cultures and herbicide-resisting plants
HU15284 1985-07-26

Publications (1)

Publication Number Publication Date
EP0169230A1 true EP0169230A1 (fr) 1986-01-29

Family

ID=10948200

Family Applications (1)

Application Number Title Priority Date Filing Date
EP85900726A Withdrawn EP0169230A1 (fr) 1984-01-16 1985-01-16 Procede de production de cultures de tissus mixotrophiques et de plantes resistant aux herbicides

Country Status (7)

Country Link
EP (1) EP0169230A1 (fr)
JP (1) JPS61501187A (fr)
DK (1) DK418185D0 (fr)
ES (1) ES539629A0 (fr)
HU (1) HU190751B (fr)
IT (1) IT1183269B (fr)
WO (1) WO1985003085A1 (fr)

Families Citing this family (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
GB2234663B (en) * 1987-03-23 1991-12-04 Imp Tobacco Co Ltd "process for producing tobacco cells"
GB2203022B (en) * 1987-03-23 1991-11-20 Imp Tobacco Co Ltd Smoking material and process for making the same
PH24574A (en) * 1987-06-01 1990-08-03 Hokko Chem Ind Co Method of breeding rice plants of new variety having resistance to herbicides
GB9123159D0 (en) * 1991-10-31 1991-12-18 Ici Plc Herbicide resistant plants
NZ252209A (en) * 1992-05-12 1995-09-26 Moet & Chandon Stabilised culture of proembryogenic cellular aggregates for use in vine regeneration techniques

Family Cites Families (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
AU529693B2 (en) * 1978-08-16 1983-06-16 British Petroleum Company Limited, The Undifferentiated plant cell cultivation
JPS5716692A (en) * 1980-07-01 1982-01-28 Nippon Paint Co Ltd Cultivating method of plant cell
FR2492404A1 (fr) * 1980-10-22 1982-04-23 Synthelabo Procede de production ou de biotransformation de metabolites par des cellules vegetales in vitro
ATE8797T1 (de) * 1980-11-06 1984-08-15 Albright & Wilson Limited Schwielenkultur in einer nahrhaften stroemung.
DE3270112D1 (en) * 1981-08-11 1986-04-30 Mitsui Petrochemical Ind Method for producing secondary metabolites of plants

Non-Patent Citations (1)

* Cited by examiner, † Cited by third party
Title
See references of WO8503085A1 *

Also Published As

Publication number Publication date
DK418185A (da) 1985-09-13
ES8601646A1 (es) 1985-11-16
HUT37648A (en) 1986-01-23
IT1183269B (it) 1987-10-22
IT8519118A0 (it) 1985-01-16
ES539629A0 (es) 1985-11-16
JPS61501187A (ja) 1986-06-19
WO1985003085A1 (fr) 1985-07-18
DK418185D0 (da) 1985-09-13
IT8519118A1 (it) 1986-07-16
HU190751B (en) 1986-11-28

Similar Documents

Publication Publication Date Title
Behnke Selection of potato callus for resistance to culture filtrates of Phytophthora infestans and regeneration of resistant plants
RU2091010C1 (ru) Способ селекции пшеницы, устойчивой к гербицидам
Fitch High frequency somatic embryogenesis and plant regeneration from papaya hypocotyl callus
US5773702A (en) Imidazolinone herbicide resistant sugar beet plants
Heim et al. Mutation of a locus of Arabidopsis thaliana confers resistance to the herbicide isoxaben
Brar et al. Isolation, fusion and culture of Sorghum and corn protoplasts
Warner et al. Nitrate utilization by nitrate reductase-deficient barley mutants
Binarová et al. Selection for resistance to filtrates of Fusarium spp. in embryogenic cell suspension culture of Medicago sativa L.
Herath et al. Effect of culture media for anther culture of indica rice varieties and hybrids of indica and japonica
KR890004024B1 (ko) 원형질체로 부터 벼 식물체의 재생방법
Heath-Pagliuso et al. Somaclonal variation in celery: screening for resistance to Fusarium oxysporum f. sp. apii
Miladinova et al. The salinity effect on morphology and pigments content in three paulownia clones grown ex vitro
Kim et al. Selection of 5-methyltryptophan resistant rice mutants from irradiated calli derived from embryos
Lee et al. Selection and characterization of a rice mutant resistant to 5-methyltryptophan
WO2001080638A1 (fr) Systemes de commande de differenciation des cellules ou des organes et procede de commande de morphogenese mettant en oeuvre lesdits systemes de commande
Shimira et al. The assessment of androgenic response of two nematode resistant pepper (Capsicum annuum L.) genotypes
EP0169230A1 (fr) Procede de production de cultures de tissus mixotrophiques et de plantes resistant aux herbicides
JP2007528197A (ja) 綿植物を生産するための組織培養法
FR2628601A1 (fr) Nouvelles lignees parentales de brassica oleracea et procede de preparation de ces lignees parentales
Arihara et al. Aluminum-tolerance of carrot (Daucus carota L.) plants regenerated from selected cell cultures
Kang et al. Selection and characterization of a 5-methyltryptophan resistant mutant in Zea mays L.
Górecka et al. Carrot doubled haploids
Dotray et al. Effects of acetyl-coenzyme A carboxylase inhibitors on root cell transmembrane electric potentials in graminicide-tolerant and-susceptible corn (Zea mays L.)
US4795705A (en) Methods of producing herbicide resistant plant varieties and plants produced thereby
US4900676A (en) Method of producing herbicide resistant plant varieties and plants produced thereby

Legal Events

Date Code Title Description
PUAI Public reference made under article 153(3) epc to a published international application that has entered the european phase

Free format text: ORIGINAL CODE: 0009012

17P Request for examination filed

Effective date: 19850909

AK Designated contracting states

Designated state(s): AT BE CH DE FR GB LI LU NL SE

17Q First examination report despatched

Effective date: 19870406

STAA Information on the status of an ep patent application or granted ep patent

Free format text: STATUS: THE APPLICATION IS DEEMED TO BE WITHDRAWN

18D Application deemed to be withdrawn

Effective date: 19880209

RIN1 Information on inventor provided before grant (corrected)

Inventor name: MARTON, LASZLO

Inventor name: MALIGA, PAL

Inventor name: MENCZEL, LASZLO

Inventor name: CSEPLO, AGNES

Inventor name: MEDGYESY, PETER