WO1999026470A1 - Nutritive media and manufactured seeds comprising same - Google Patents
Nutritive media and manufactured seeds comprising same Download PDFInfo
- Publication number
- WO1999026470A1 WO1999026470A1 PCT/US1998/024820 US9824820W WO9926470A1 WO 1999026470 A1 WO1999026470 A1 WO 1999026470A1 US 9824820 W US9824820 W US 9824820W WO 9926470 A1 WO9926470 A1 WO 9926470A1
- Authority
- WO
- WIPO (PCT)
- Prior art keywords
- nutritive medium
- manufactured seed
- medium further
- hcl
- manufactured
- Prior art date
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Classifications
-
- 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
- A01H4/005—Methods for micropropagation; Vegetative plant propagation using cell or tissue culture techniques
- A01H4/006—Encapsulated embryos for plant reproduction, e.g. artificial seeds
Definitions
- the invention pertains to nutritive media, methods for using such media in the production of manufactured seeds, and manufactured seeds comprising same.
- Standard plant breeding techniques are labor intensive and, because they usually involve fusion of gametes, tend to introduce genetic variability.
- the genetic variability is a result of crossing over of chromosomes, which often occurs during meiosis.
- standard breeding techniques require that the breeder wait until a plant is mature enough to breed. In some cases, this means waiting many years. This delay decreases productivity and increases costs. Therefore, it is desirable to produce large numbers of genetically identical plants via culturing of somatic or zygotic plant embryos. Somatic or zygotic plant embryos may be cultured in the form of "manufactured seeds".
- Manufactured seeds are essentially analogs of botanic seed and typically include a nutrient medium (termed here in a "nutritive medium”), and typically include structural features that serve to protect the embryo before, during, and after germination.
- a nutrient medium typically include structural features that serve to protect the embryo before, during, and after germination.
- References that describe manufactured seeds include U.S. Patent Nos. 5,564,224; 5,687,504; and 5,701,699, all to Carlson et al. These patents disclose elements of such seeds, for instance, a manufactured seed coat, methods for using such manufactured seeds, and plant germinants produced from manufactured seeds.
- a problem with manufactured seeds to date is the relatively low numbers of successful germinants from such seeds compared to botanical (natural) seeds, and relatively low numbers of "normal” germinants from manufactured seeds, i.e., viable, uniform, and commercially useful germinants.
- Abnormal germinants possess any of various malformations such as small and/or deformed radicals, hypocotyls, or cotyledons. Although many factors appear to cause abnormal germination, the results generally indicate that manufactured seeds, as currently known in the art, do not provide a correct balance of nutrients and other elements to the seed. Therefore, there is a need for nutritive media that contain a suitable profile of nutrients and other elements that, when incorporated into manufactured seeds, facilitate normal and successful germination and germinant growth with normal organ development.
- a nutritive medium desirably facilitates the germination of manufactured seeds at a rate at least similar to the germination rate of sexually produced seeds.
- Commercial viability can be assessed by taking into account the market size and the volume of product necessary to meet market needs. In the case of plant-related products the market is vast. Therefore, a difference in the germination rate of 10% or 20% can have an immense impact on the commercial viability of manufactured seeds.
- the embryo relies on the megagametophyte for nutrients useful for germination
- the embryo in a manufactured seed relies on the nutritive medium that is provided in the manufactured seed.
- an agar medium used as a surface on which plant embryos are grown appears to have substantially different requirements compared to a nutritive medium for use in manufactured seed. Therefore, there is a need for nutritive media that, when incorporated into manufactured seeds, provides an improved germination rate compared to conventional manufactured seeds.
- manufactured seeds comprise a unit of totipotent plant tissue and a nutritive medium.
- the nutritive medium preferably comprises 0 g/L to 30 g/L gel solute, 5 mM to 200 mM carbon source, 0 mM to 20 mM KN0 3 , 2.5 mM to 5 mM NH 4 N0 3 , 0 mM to 0.01 mM KI, 0.01 mM to 20 mM KH 2 P0 4 , 0.5 mM to 5 mM CaCl 2 , 0.1 mM to 10 mM MgS0 4 , 0 mM to 0.1 mM Na 2 EDTA, 0 mM to 0.1 mM FeS0 4 , 0 mM to 0.1 mM MnS0 4 , 0.01 mM to 1 mM H 3 B0 3 , 0.0001 mM to 0.00
- the nutritive medium can also include any of various additional components such as one or more vitamins, charcoal, smoke suspension, and one or more carbon sources (e.g., a monosaccharide, a disaccharide, or a starch).
- one or more vitamins, charcoal, smoke suspension, and one or more carbon sources e.g., a monosaccharide, a disaccharide, or a starch.
- Any of various different amino acids can be incorporated into the nutritive medium as required. Such amino acids can be those that are commonly found incorporated into proteins (termed "protein amino acids").
- the preferred protein amino acids are one or more of L- asparagine, L-glutamine, L-lysine, L-serine, L-proline, L-arginine, L-valine, L-alanine, L-cysteine, L-leucine, L-tyrosine, L-threonine, L-phenylalanine, L-histidine, L-glycine, L-tryptophan, L- isoleucine, L-methionine, and mixtures thereof.
- the nutritive medium can include one or more amino acids that are not commonly found incorporated into proteins (termed “non-protein amino acids”). There are over 200 such amino acids, some of which are argininosuccinate, citrulline, canavanine, and ornithine. The non- protein amino acids also encompass the D-stereoisomers of the protein amino acids.
- the nutritive medium can comprise one or more compounds involved in the metabolism of nitrogen. Representative examples of such compounds include urea and/or polyamines such as one or more of spermidine, spermine, and putrescine.
- the nutritive medium can include a smoke suspension.
- a suitable smoke suspension contains one or more compounds generated through the process of burning organic matter such as cellulosic material.
- manufactured seeds comprise a modified nutritive medium.
- a medium preferably comprises 0 g/L to 30 g/L gel solute, 0 g/L to 5 g/L charcoal, 5 mM to 200 mM carbon source, 0 mM to 30 mM urea, 0 mM to 20mM KNO 3 , 2.5 mM to 5 mM NH 4 N0 3 , 0 mM to 0.01 mM KI, 0.01 mM to 20 mM KH 2 P0 4 , 0.5 mM to 5 mM CaCl 2 , 0.1 mM to 10 mM MgS0 4 , 0 mM to 0.1 mM Na 2 EDTA, 0 mM to 0.1 mM FeS0 4 , 0 mM to 0.5 mM ferric citrate, 0 mM to 0.1 mM MnS0 4 , 0 mM to
- Such a nutritive medium can also include one or more additional compounds selected from any of various vitamins, hormones, a smoke suspension, and sources of carbon.
- the modified nutritive medium can include 0 to 5 g/L charcoal and a carbon source such as a monosaccharide, a disaccharide, or a starch.
- a modified nutritive medium according to the invention can include one or more compounds involved in the metabolism of nitrogen.
- Representative examples of such compounds are urea and polyamines such as spermidine, spermine, and putrescine.
- the modified nutritive medium according to the invention can include a "smoke suspension” .
- a suitable "smoke suspension” contains one or more compounds that are generated through the process of burning organic matter.
- the modified nutritive medium can include one or more non-protein amino acids.
- a representative embodiment of such a method comprises the steps of: providing a nutritive medium as summarized above, providing a unit of totipotent plant tissue, placing the unit of totipotent plant tissue in "functional contact” with the nutritive medium, and placing the manufactured seed in an environment conducive for plant growth so as to allow the plant tissue to grow and germinate from the manufactured seed.
- a basic nutritive medium according to the invention can have a profile within one of the two ranges provided below in Table 1.
- the column labeled "Concentration Range 1" pertains to preferred concentration ranges of the representative components.
- the column labeled “Concentration Range 2” pertains to more preferred concentration ranges.
- concentrations given in g/L denote grams of the respective component per liter of medium.
- the basic medium preferably contains at least fifteen of the components listed in Table 1 and more preferably at least twenty of the components listed in Table 1.
- the basic nutritive medium can be sterilized by autoclaving, filter sterilizing, or by using another suitable method.
- the nutritive medium can be incorporated into manufactured seed as described in, e.g., U.S. Patent No. 5,701,699 to Carlson, incorporated herein by reference.
- the basic nutritive medium facilitates germination of seedlings from manufactured seed containing such medium.
- certain components e.g., thiamine
- the basic nutritive medium can be formulated using such components that do not contain HCl. For example, it is possible to use thiamine without the HCl instead of thiamine-HCl.
- the present invention also encompasses nutritive media, derived from the basic nutritive medium described above, having an altered profile of components, including one or more added components.
- Such additional components include one or more protein amino acids, one or more non-protein amino acids, charcoal, smoke suspension, one or more carbon sources, and any of various components of the metabolic pathways pertaining to nitrogen utilization by plants. Components of these pathways include, but are not limited to, urea, putrescine, spermidine, and spermine. ⁇ . Additional Components a. Amino Acids
- Representative protein amino acids that can be added to the basic nutritive medium include one or more of the following: L-glutamine, L-arginine, L-proline, L-asparagine, L-lysine, L-serine, L-valine, L-alanine, L-cysteine, L-leucine, L-tyrosine, L-threonine, L-phenylalanine, L- histidine, L-tryptophan, L- glycine, and L-isoleucine.
- a preferred concentration of protein amino acids in the nutritive medium is 0 mM to 8 mM, more preferably 0.01 mM to 4 mM.
- the nutritive medium may comprise one or more amino acids that are not usually constituents of proteins.
- Such "non-protein” amino acids are commonly found in certain plants as storage proteins, and over 200 such non-protein amino acids are known.
- the "non-protein” amino acids include any of the D-stereoisomers of the protem amino acids.
- omithine, canavanine, or other non-protein amino acid (or mixture thereof) can be added to the nutritive medium at a concentration of 0 mM to 3 mM, more preferably 0. ImM to 1.25 mM per non-protein amino acid.
- charcoal can favorably influence germination when added to a nutritive medium.
- the charcoal is in the form of a powder and is activated by pretreatment with HCl.
- Charcoal appears to facilitate increased hypocotyl length and increased radical length of germinating plant embryos of certain species of plants.
- the preferred concentration of charcoal is 0 g/L to 5 g/L, more preferably 2 g/L to 3 g/L.
- the by-products of burning organic matter can contribute to increased germination.
- Solutions containing these by-products can be generated by burning organic matter (e.g. , wood or other cellulosic material), washing the charred material with water, and collecting the water. Solutions can also be obtained by heating the organic matter (e.g., wood or other cellulosic material), and condensing and diluting volatile substances released from such heating.
- a nutritive medium according to the invention comprising a smoke suspension can increase organ length and improve germination of certain plant species.
- Smoke suspension can be incorporated into the nutritive medium in any of various forms. For instance, smoke suspension can be incorporated as an aerosol, a powder, or as activated clay. The preferred concentration of
- the carbon source in a nutritive medium according to the invention can be, for example, one or more monosaccharides (as representative "simple” carbon sources).
- the nutritive medium can include one or more "complex" carbon sources such as any of various disaccharides (e.g.
- a preferred concentration of the carbon source (either as a single compound or as a mixture of compounds) in the medium is 5 mM to 200 mM, more preferably 25 mM to 175 mM.
- a growing plant needs to be able to take in nitrogen and metabolically utilize it.
- a representative source of nitrogen is arginine. Therefore, certain compounds involved in the synthesis and degradation of arginine can be especially useful as additives to a nutritive medium according to the invention.
- One such compound is urea.
- a preferred concentration of urea is 0 mM to 30 mM.
- a more preferred concentration of urea, if used, is 5 mM to 20 mM.
- any of various polyamines involved in the degradation of arginine can be added to a nutritive medium according to the invention.
- Suitable polyamines can be selected from the breakdown products of certain amino acids.
- a preferred concentration of spermidine, putrescine, and/or spermidine is 0 mM to 0.5 mM per compound.
- a more preferred concentration is 0.01 mM to 0.5 mM per such compound present in the medium.
- the nutritive medium can comprise any of various additional components.
- modified nutritive media according to the invention can have a profile within one of the two ranges provided in Table 2, below.
- Table 2 the column labeled
- Consration Range 1 pertains to preferred concentration ranges of the respective components.
- the column labeled “Concentration Range 2” pertains to more preferred concentration ranges.
- concentrations given in g/L denote grams of the respective component per liter of medium.
- the modified medium preferably contains at least one of the protein amino acids listed in Table 2, and more preferably at least five of the protein amino acids listed in Table 2.
- the modified medium contains at least 15 of the other components (that are not protein amino acids) listed in Table 2, and more preferably at least 20 of the other components listed in Table 2.
- a "polyamine” is any of various compounds comprising molecules having two or more amine groups, such as, but not limited to, putrescine, spermine, and spermidine.
- concentrations of individual components can be varied depending upon the species of plant embryo contained in the manufactured seed and upon the environment in which the seed is to be germinated.
- Plant Tissue Plant tissue incorporated into a manufactured seed comprising a nutritive medium according to the invention is preferably totipotent.
- totipotent refers to a capacity to grow and develop into a normal plant.
- Totipotent plant tissue has both the complete genetic information of a plant and the ready capacity to develop into a complete plant if cultured under favorable conditions.
- Totipotent plant tissue is obtainable from several areas of a plant, such as meristematic tissue and plant embryonic tissue.
- Meristematic tissue is comprised of undifferentiated plant cells that divide to yield other meristematic cells as well as differentiated cells that elongate and further specialize to form structural tissues and organs of the plant.
- Meristematic tissue is located, for example, at the extreme tips of growing shoots or roots, in buds, and in the cambium layer of woody plants.
- Plant embryonic tissue can be found (in the form of a "zygotic" embryo) inside a botanic seed produced by sexual reproduction.
- plant "somatic" embryos can be produced by culturing totipotent plant cells such as meristematic tissue under laboratory conditions in which the cells comprising the tissue are separated from one another and urged to develop into minute complete embryos.
- cleavage polyembryony a process termed "cleavage polyembryony" known in the art can be induced during natural embryo development in seed.
- totipotent plant tissue is referred to herein simply as the “embryo” , unless stated otherwise.
- a nutritive medium according to the invention can include a substance that causes the medium to be a semisolid or have a congealed consistency under normal environmental condition (see Definitions below).
- a detailed description of various types of gel solutes can be found in U.S. Patent No. 5,564,224 to Carlson, incorporated herein by reference.
- U.S. Patent No. 5,564,224 discloses various oxygen-absorbing substances (also termed "oxygen-carrying" substances). Such substances can be added to a nutritive medium according to the present invention to enhance both the absorption of oxygen and the retention of oxygen by the nutritive medium, thereby allowing the medium to maintain a concentration of oxygen that is higher than would otherwise be present in the medium solely from the absorption of oxygen from the atmosphere.
- the amount and type of oxygen-carrying substance that is added to the medium will vary depending on the species being germinated from the manufactured seed. In some cases a nutritive medium including an oxygen-carrying substance does not facilitate improved germination compared to an otherwise identical nutritive medium lacking an oxygen-carrying substance.
- Nutritive media according to the invention are useful for manufacturing and germinating manufactured seeds in a variety of different contexts. As previously mentioned, the optimal concentrations of the various components can be adjusted within the respective stated ranges depending upon the species of embryo and the conditions under which the manufactured seed will be sown. Representative methods used for making manufactured seeds including a nutritive medium are described in U.S. Patent No. 5,701,699 to Carlson, incorporated herein by reference.
- Unit of totipotent plant tissue refers to the minimum mass of plant tissue that can give rise to a mature plant. For example, a single cell can mature into a plant, and is therefore, a unit of totipotent plant tissue. However, in a practical sense, a suitable unit of totipotent plant tissue comprises multiple cells (hence the term "tissue") such as in a plant embryo.
- “Functional contact” is any contact of the plant tissue with the nutritive medium that allows for the flow of water and components from the nutritive medium to the unit of totipotent plant tissue. Therefore, functional contact can include actual physical contact or indirect contact resulting from, e g , enclosing the plant tissue with a cotyledon restraint such as described in U S Patent No 5,687,504, incorporated herein by reference
- Non-protein amino acid is any amino acid that is not normally found as a constituent of proteins, for example, but not limited to, argimnosuccinate, citrulline, homose ⁇ ne, and ormthme
- Gel solute is a solute that, when added to the medium, causes the medium to become a semisohd or otherwise congeal
- a preferred gel solute is neither cytotoxic nor substantially phytotoxic
- Candidate gel solutes include, but are not limited to, the following sodium alginate, agar, agarose, amylose, pectin, dextran, gelatin, starch, amylopectin, modified celluloses such as methylcellulose and hydroxyethylcellulose and polyacrylamide
- Somatic embryo is a plant embryo produced via the laboratory cultu ⁇ ng of totipotent plant cells or by induced cleavage polyembryony
- Zygotic embryo is a plant embryo removed from a natural botanic seed of the correspondmg plant "Germinant” is a plant embryo that has undergone sufficient growth and development to protrude from a manufactured seed, analogous to a plant embryo protruding from a natural botanic seed
- “Radicle” is that part of a plant embryo that develops into the primary root of the resulting plant “Cotyledon” refers generally to the first, first pair, or first whorl (depending on the plant type) of leaf-like structures on the plant embryo that function primarily to make food compounds m the seed available to the developing embryo, but m some cases act as food storage or photosynthetic structures
- Hypocotyl is that portion of a plant embryo or seedling located below the cotyledons but above the radicle
- Ring length pertains to the length of the radicle at the time the radicle was measured
- Normalcy denotes the presence of all expected parts of a plant (e g , radicle, hypocotyl, cotyledon(s), epicotyl) at time of evaluation In the case of gymnosperms, normalcy is characterized by the radicle having a length greater than 3 mm and no visibly discernable malformations compared to the appearance of control bare embryos grown on the surface of nutrient agar or the like
- the basic nutritive medium and the modified nutritive medium are preferably prepared from pre-made stocks.
- the required amount of each stock solution (that is not heat-labile) is added to water.
- Non-stock chemicals such as sucrose, charcoal, and agar
- the medium is brought up to an appropriate volume and the pH is adjusted.
- the medium is then sterilized, e.g. , by autoclaving.
- Pre-sterilized (e.g. , by filtration) heat-labile components are added after the medium is removed from the autoclave and has cooled.
- the medium can additionally contain one or more antibiotics and/or antimycotics.
- the medium can contain 0.1 mg/L GA 4/7 (available from Abbot Laboratories, North Chicago, IL) and/or 1 mL/L media Sigma product # A7292. Both of these products contain an antibiotic and an antimycotic.
- both products are heat-labile, they are preferably added after the medium has cooled to below 40 °C.
- the sterilization of different components can be performed using any of various suitable techniques that can be performed at various stages in preparation.
- a stock solution can be made that contains one or more of the components in the media. This stock solution can be sterilized before addition to the medium, or after addition to the medium.
- the solutions can be sterilized by filtration, exposure to UV light, autoclaving, or any other suitable means.
- the sterilization principles described above can also be applied to other media additives.
- amino acids, carbon sources, smoke suspension, or polyamines can be made into a stock solution that is filter sterilized.
- the resulting sterilized stock solutions can then be added to the medium after an intermediate solution containing other components has been autoclaved.
- these additives might be added directly to the medium and the medium as a whole subsequently sterilized.
- the manufactured seeds are then sown in the sand and allowed to germinate.
- the manufactured seeds are cultured under continuous light at room temperature (23 °C) for four to five weeks.
- This example is directed to the effect of adding charcoal to the basic nutritive medium.
- Douglas-fir zygotic embryos were used in manufactured seed containing the basic medium.
- the control group included 36 manufactured seeds, and the test group included 36 manufactured seeds. Seeds of the test group contained the basic nutritive medium with the addition of 2.5 g/L charcoal. Seeds of the control group contained the basic nutritive medium without charcoal.
- the addition of amino acids to the basic nutritive medium described in Table 3 was evaluated to determine the effect of the added amino acids on the germination of manufactured seeds containing such nutritive medium.
- the manufactured seeds were constructed by the methods described in Example 2.
- the control group consisted of manufactured seeds containing the basic medium.
- Test groups consisted of manufactured seeds containing the basic nutritive medium additionally comprising 0.50 mM L-glutamine, 0.125 mM L-proline, and 0.125 mM L-asparagine plus one additional test component (a different test component for each test group).
- test components were 0.50 mM L-arginine, 0.50 mM ornithine, or a combination of polyamines (0.10 mM putrescine, 0.10 mM spermidine, and 0.10 mM spermine).
- the manufactured seeds of all treatments contained Douglas fir embryos. There were six replicates with six manufactured seeds from each treatment in each replicate. The seeds were germinated in sterile sand as described in Example 2. The seeds were allowed to germinate for 25 days after which organ length and normalcy of germinants were assessed. These results are shown in Table 5.
- the basic medium, without pluronic F-68 and DC-200 silicone oil, described in Table 3 was supplemented to bring the sucrose concentration to 50 g/L. This supplemented basic medium was then compared to the modified medium described in Table 3.
- the manufactured seeds were constructed containing loblolly pine embryos according to Example 1.
- Manufactured seeds containing the modified nutritive medium exhibited significantly better germination, normalcy, epicotyl presence, and radical elongation than manufactured seeds containing the basic nutritive medium.
- the two media formulations listed in Table 3 were incorporated into respective populations of manufactured seeds containing Douglas fir zygotic embryos. This was done to determine which of the two media was more effective at producing normal germination and organ elongation of such embryos from manufactured seeds. A third treatment was included to determine the effect of sucrose concentration on germination of such embryos.
- Manufactured seeds were constructed as described in Example 1. The study consisted of six replicate germination boxes with ten manufactured seeds/treatment/box. The seeds were allowed to germinate in sterile sand as described above for 27 days. Measurements were then taken and recorded. The data are presented in Table 7.
- modified medium with either concentration of sucrose significantly increased organ elongation of Douglas fir germinants from manufactured seed.
- the modified medium with 50 g/L sucrose produced the longest radicals and hypocotyls and increased cotyledon length.
- Example 7 The Addition of Liquid Smoke to Nutritive Medium This example is directed to the effect of a nutritive medium including "liquid smoke” (as a representative smoke suspension) on the germination of manufactured seeds containing such a medium.
- the starting medium used was the modified medium listed in Table 3 without Pluronic F-
- Hickory Seasoning Liquid Smoke were added. (Wright's Concentrated Hickory Seasoning Liquid Smoke is available from B&G foods, Inc. Roseland, NJ 07068). Treatment groups contained either 0.01 ⁇ L, 0.1 ⁇ L, 1 ⁇ L, 10 ⁇ L, 100 ⁇ L, 1 mL, or 10 mL of liquid smoke per 100 mL medium.
- the manufactured seeds were prepared as described in Example 2, using Loblolly pine somatic embryos. The seeds were sown in germination boxes containing sterile sand as described above. The study consisted of six germination boxes and eight (30 seeds each) treatments with five seeds per treatment per box (a total of 240 manufactured seeds). After 34 days the seeds were scored for normalcy and organ elongation. Results are shown in Table 8.
- Example 8 Effects of a Carbon Source on Germination of Manufactured Seeds
- Manufactured seeds were prepared as described in Example 1.
- Media formulations were based on the basic medium except that the following carbon sources and concentrations were used: Treatment 1 - basic medium containing 20 g/L sucrose; Treatment 2 - basic medium containing 20 g/L maltose; Treatment 3 - basic medium containing 20 g/L glucose; Treatment 4 - basic medium containing 10 g/L maltose; Treatment 5 - basic medium containing 10 g/L glucose, and Treatment 6 - basic medium containing 20 g/L honey.
- Stock solutions of the glucose and maltose used to prepare media used in Treatments 2-5 were filter sterilized instead of autoclaved. In this study there were six replicate germination boxes containing sterile sand with five manufactured seeds per Treatment in each box.
- Treatment 3 produced 0.85 cm radicals; Treatment 1 produced 0.68 cm radicals; Treatment 6 produced 0.61 cm radicals; Treatment 5 produced 0.43 cm radicals; Treatment 2 produced 0.38 cm radicals; and Treatment 5 produced 0.34 cm radicals.
- Example 1 various concentrations of sucrose were incorporated into the basic medium used in manufactured seeds containing loblolly pine zygotic embryos.
- Manufactured seeds were produced as described in Example 1.
- Treatment groups consisted of manufactured seeds containing the basic medium as described in Table 3 with either 40 g/L filter-sterilized sucrose or 50 g/L filter-sterilized sucrose.
- the manufactured seeds were sown in sterile sand in germination boxes as described in Example 1 with 10 seeds/treatment/box (total of 60 seeds/treatment). Data was collected 31 days after sowing. The results are shown in Table 9.
Abstract
Description
Claims
Priority Applications (4)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
AU15314/99A AU731857B2 (en) | 1997-11-20 | 1998-11-20 | Nutritive media and manufactured seeds comprising same |
NZ505099A NZ505099A (en) | 1997-11-20 | 1998-11-20 | Nutritive media and manufactured seeds comprising same |
CA2309258A CA2309258C (en) | 1997-11-20 | 1998-11-20 | Nutritive media and manufactured seeds comprising same |
US10/371,612 US8466086B2 (en) | 1997-11-20 | 2003-02-20 | Nutritive media and manufactured seeds comprising same |
Applications Claiming Priority (2)
Application Number | Priority Date | Filing Date | Title |
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US6623297P | 1997-11-20 | 1997-11-20 | |
US60/066,232 | 1997-11-20 |
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US09529933 A-371-Of-International | 1998-11-20 | ||
US10/371,612 Division US8466086B2 (en) | 1997-11-20 | 2003-02-20 | Nutritive media and manufactured seeds comprising same |
Publications (1)
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WO1999026470A1 true WO1999026470A1 (en) | 1999-06-03 |
Family
ID=22068160
Family Applications (1)
Application Number | Title | Priority Date | Filing Date |
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PCT/US1998/024820 WO1999026470A1 (en) | 1997-11-20 | 1998-11-20 | Nutritive media and manufactured seeds comprising same |
Country Status (4)
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AU (1) | AU731857B2 (en) |
CA (1) | CA2309258C (en) |
NZ (1) | NZ505099A (en) |
WO (1) | WO1999026470A1 (en) |
Cited By (19)
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DE10011998A1 (en) * | 2000-03-11 | 2001-09-20 | Forschungszentrum Juelich Gmbh | A new growth promoting active substance combination contains L-serine, L-asparagine and/or L-glutamine provides a growth medium for cells, particularly human or mammalian cells |
US7016792B2 (en) | 2001-06-22 | 2006-03-21 | Hottinger Baldwin Messtechnik Gmbh | Transducer comprising a connected data memory |
EP1699288A1 (en) * | 2003-12-04 | 2006-09-13 | Cellfor, Inc. | Method of ex vitro sowing, germination, growth and conversion of plant somatic embryos or germinants, and nutrient medium used therefor |
US7131234B2 (en) | 2003-11-25 | 2006-11-07 | Weyerhaeuser Co. | Combination end seal and restraint |
US7228658B2 (en) | 2003-08-27 | 2007-06-12 | Weyerhaeuser Company | Method of attaching an end seal to manufactured seeds |
AT503565B1 (en) * | 2006-06-16 | 2007-11-15 | Jhs Privatstiftung | PROCESS FOR THE PRODUCTION OF ORGANICALLY BOUND VITAMIN B |
US7356965B2 (en) | 2003-12-11 | 2008-04-15 | Weyerhaeuser Co. | Multi-embryo manufactured seed |
US7547488B2 (en) | 2004-12-15 | 2009-06-16 | Weyerhaeuser Nr Company | Oriented strand board panel having improved strand alignment and a method for making the same |
US7555865B2 (en) | 2003-11-25 | 2009-07-07 | Weyerhaeuser Nr Company | Method and system of manufacturing artificial seed coats |
US7568309B2 (en) | 2004-06-30 | 2009-08-04 | Weyerhaeuser Nr Company | Method and system for producing manufactured seeds |
US7591287B2 (en) | 2003-12-18 | 2009-09-22 | Weyerhaeuser Nr Company | System and method for filling a seedcoat with a liquid to a selected level |
US7654037B2 (en) | 2005-06-30 | 2010-02-02 | Weyerhaeuser Nr Company | Method to improve plant somatic embryo germination from manufactured seed |
RU2449525C2 (en) * | 2010-04-15 | 2012-05-10 | Анатолий Александрович Астахов | Agent for presowing seeds treatment, mostly sunflower (versions) |
US8252718B2 (en) | 2009-09-30 | 2012-08-28 | Weyerhaeuser Nr Company | Nutritive media for use in manufactured seeds |
US8375630B2 (en) | 2009-09-30 | 2013-02-19 | Weyerhaeuser Nr Company | Method to improve germination of embryos from manufactured seed |
US8466086B2 (en) | 1997-11-20 | 2013-06-18 | Weyerhaeuser Nr Company | Nutritive media and manufactured seeds comprising same |
US8871514B2 (en) | 2009-09-30 | 2014-10-28 | Weyerhaeuser Nr Company | Methods of preparing nutritive media for growth and/or germination of plant embryos |
EP1974014B1 (en) | 2006-01-04 | 2017-04-19 | Baxalta Incorporated | Oligopeptide-free cell culture media |
CN115380662A (en) * | 2022-10-27 | 2022-11-25 | 中国医学科学院药用植物研究所 | Seed coating method for improving drought resistance of isatis roots in seedling stage |
Families Citing this family (1)
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WO2006094400A1 (en) * | 2005-03-10 | 2006-09-14 | Cellfor Inc. | Aerated liquid priming of conifer somatic germinants |
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- 1998-11-20 WO PCT/US1998/024820 patent/WO1999026470A1/en active IP Right Grant
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- 1998-11-20 CA CA2309258A patent/CA2309258C/en not_active Expired - Fee Related
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US7016792B2 (en) | 2001-06-22 | 2006-03-21 | Hottinger Baldwin Messtechnik Gmbh | Transducer comprising a connected data memory |
US7228658B2 (en) | 2003-08-27 | 2007-06-12 | Weyerhaeuser Company | Method of attaching an end seal to manufactured seeds |
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EP1699288A4 (en) * | 2003-12-04 | 2008-04-02 | Cellfor Inc | Method of ex vitro sowing, germination, growth and conversion of plant somatic embryos or germinants, and nutrient medium used therefor |
US7356965B2 (en) | 2003-12-11 | 2008-04-15 | Weyerhaeuser Co. | Multi-embryo manufactured seed |
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Also Published As
Publication number | Publication date |
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AU731857B2 (en) | 2001-04-05 |
CA2309258C (en) | 2012-05-01 |
AU1531499A (en) | 1999-06-15 |
NZ505099A (en) | 2002-03-01 |
CA2309258A1 (en) | 1999-06-03 |
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