Composition for seed growth and vigour in monocots

Abstract

Coating composition for applying to monocot plant structures from which roots and shoots are capable of growing, wherein the said coating composition comprises one or more organic materials having a melting point of ≧50° Centigrade and one or more additives, methods of making such compositions and coated monocot plant structures such as seeds of monocot plants.

Claims

1. A coating composition for a monocot plant structure, consisting essentially of: particles consisting essentially of: i. at least one organic material selected from the group consisting of waxes having a melting point of ≧50° Centigrade; and ii. at least one additive capable of enhancing at least one of seedling vigour and seedling growth from monocot plant structures, wherein the at least one additive is selected from the group consisting of inorganic additives, live biological agents, and combinations thereof, wherein the particles have a volume mean diameter ≧5 μm.

2. A coating composition according to claim 1, wherein the particles have a volume mean diameter in the range of 10 to 200 μm.

3. The coating composition according to claim 1, wherein the organic material is selected from the group consisting of carnauba wax, beeswax, montan wax, Chinese wax, shellac wax, spermaceti wax, myricyl palmitate, cetyl palmitate, candelilla wax, castor wax, ouricury wax, and rice bran wax or is a mixture of two or more thereof.

4. The method according to claim 1, wherein the monocot plant structure is selected from the group consisting of (1) monocot plant seeds selected from the group consistng of Oryza spp., Triticum spp., Secale spp., Avena spp., Zea spp., Sorghum spp., Hordeum spp., and (2) seeds of hybrid crosses of monotcotyledonous plants.

5. The coating composition according to claim 1, wherein the monocot plant structures are monocot plant seeds are selected from the group consisting of Oryza sativa, T. aestivum, Secale cereale, Avena sativa, Zea mays, Sorghum bicolor, Hordeum vulgare and x Triticosecale.

6. Monocot plant structures comprising a coating composition according to claim 1.

7. Monocot plant structures comprising a coating composition according to claim 1 that are selected from the group consisting of coated seeds and coated bulbs.

8. Monocot plant structures, comprising a coating composition according to claim 1, that are monocot seeds.

9. Monocot plant structures according to claim 7 that are monocot plant seeds selected from the group consisting of Oryza spp., Triticum spp., Secale spp., Avena spp., Zea spp., Sorghum spp., Hordeum spp., and seeds of hybrid crosses of monotcotyledonous plants.

10. Monocot plant structures according to claim 7 that are monocot plant seeds selected from the group consisting of Oryza sativa, T. aestivum, Secale cereale, Avena sativa, Zea mays, Sorghum bicolor, Hordeum vulgare and x Triticosecale.

11. A method of manufacturing a monocot plant structure coating composition, said coating composition being in particulate form for a monocot plant structure and consisting essentially of particles, said particles consisting essentially of at least one organic material selected from the group consisting of waxes, and at least one additive capable of enhancing at least one of seedling vigour and seedling growth from monocot plant structures, wherein the at least one additive is selected from the group consisting of inorganic additives, live biological agents and combinations thereof, the method comprising: 1) selecting a solid mass of said at least one organic material; 2) machining the solid mass of said at least one organic material into particles of a volume mean diameter ≧10 μm; and 3) adding to said particles said at least one additive so as to form said coating composition for enhancing at least one of seedling vigour and seedling growth from monocot plant structure.

12. The method according to claim 11, wherein the organic material is selected from the group consisting of carnauba wax, beeswax, montan wax, Chinese wax, shellac wax, spermaceti wax, candelilla wax, castor wax, ouricury wax, and rice bran wax; or a mixture of two or more thereof.

13. A method of coating monocot plant structures with a coating composition in dry particulate form that is consisting essentially of at least one organic material selected from the group consisting of waxes having a melting point of ≧50° Centigrade and at least one additive for enhancing at least one of seedling vigour and seedling growth from monocot plant structures, the method comprising : i. providing particles consisting essentially of the at least one organic material admixed with the at least one additive capable of enhancing at least one of seedling vigour and seedling growth from monocot plant structures, the at least one additive being selected from the group consisting of an inorganic additive and a live biological agent and the particles being of a predetermined volume mean diameter; and ii. applying the particles consisting essentially of the at least one organic material admixed with the at least one additive to monocot plant structures.

14. The method according to claim 13, wherein the organic material is selected from the group consisting of carnauba wax, beeswax, montan wax, Chinese wax, shellac wax, spermaceti wax, candelilla wax, castor wax, ouricury wax, and rice bran wax or is a mixture of two or more thereof.

15. The method of coating monocot plant structures according to claim 13 with a coating composition that comprises an organic material selected from the group consisting of waxes having a melting point of ≧50° Centigrade, the method comprising i. providing the organic material; ii. heating the organic material so as to form a liquid phase or a gaseous phase; iii. cooling the liquid phase or gaseous phase of ii) to below the melting point of the organic material so as to form a solid; iv. adding one or more additives to the solid formed in iii); v. machining the solid organic material of iii) into particles of a predetermined volume mean diameter, and vi. applying the particles of v) to monocot plant structures.

16. The method of coating monocot plant structures according to claim 15 with a coating composition that comprises an organic material that is selected from the group consisting of waxes having a melting point of ≧50° Centigrade, the method comprising i. providing the organic material; ii. heating the organic material so as to form a liquid phase or a gaseous phase; iii. adding one or more additives to the liquid phase or gaseous phase of ii); iv. cooling the liquid phase or gaseous phase of iii) to below the melting point of the organic material so as to form a solid; v. machining the solid organic material of iv) into particles of a predetermined volume mean diameter; and vi. applying the particles of v) to monocot plant structures.

Description

EXAMPLE 1

Growth and Vigour in Triticum aestivum

(1) Triticum aestivum seed provided by Herbiseeds (Twyford, UK)

(2) Combination of Carnauba Wax Particles and Inoculant

(3) Rock Phosphate

(4) Rock Phosphate (Garden Direct, UK) with a 30% P.sub.2O.sub.5 content is crushed using a pestle and mortar and then passed through a 32 micron mesh sieve.

(5) Carnauba Wax Sizing

(6) Steps in Air Milling in Boyes Micronisation Process (for carnauba wax particles with a VMD of approx. 75 μm)

(7) 1. 2 kg carnauba wax blocks are first kibbled into approximately 4 to 6 mm pieces in a KT Handling Ltd Model 04 kibbler (serial no. 729/C) following the manufacturer's instructions.

(8) 2. The kibbled pieces are then passed through an Apex Construction Ltd Model 314.2 Comminuting Mill (serial no. A21306) and reduced further in size to a range of 250 to 300 um.

(9) 3. The comminuted particles are then passed through a Hosokawa Micron Ltd Alpine 100AFG jet mill (serial no. 168092) following the manufacturer's instructions, setting the mill at a speed of 2500 rpm for particles having a VMD of approx. 75 μm), with a positive system pressure of 0.03 bar.

(10) 4. The grinding air is to be kept to 6 bar, the system rinsing air flow and Classifying Wheel gap rinsing air are both to be set at a minimum of 0.5 bar and no more than 0.75 bar, the cleaning air filter is to register a delta of no more than 5 bar to achieve a final particle size with a VMD of approx. 75 μm.

(11) Rock Phosphate is combined with Carnauba wax particles (VMD 75 μm) at a ratio of 1:3 (Rock Phosphate:Carnauba wax particles). A homogeneous mix of is attained through tumbling seed and carnauba wax formulation in a cylinder, adapted to produce lateral mixing/tumbling through the inclusion of angled interior vanes, placed on a Wheaton roller for 5 minutes.

(12) Dry Powder Formulation of Mycorrhizae International Culture Collection of VA Mycorrhizal Fungi (INVAM).

(13) Mycorrhizae concentration is measured by diluting 1 gm in 1 l of water, before further diluting by taking 1 ml of the suspension and making it up to 1000 ml. A 20 μl sample is then added to an Improved Neubauer Counting Slide and a count made of 4 large squares (0.1 mm^3) in both of the grids. The mean for each square is calculated and the mean of the two grids used to produce a measurement of spores per 100 nl of water. The dilution factor is then applied to produce an approximation of the number of spores per gram.

(14) Carnauba wax particles produced according to the method described above with the exception that the milling speed was set at 12,500 rpm providing particles having a VMD of 16 μm are combined with at a ratio of 1:3 (Mycorrhizae:Carnauba wax particles) in a 50 ml tube using a Stuart roller mixer set at 25 rpm for 5 minutes. This can then be used to calculate the quantity of spore/Carnauba wax particles powder mix required for the seed coating based on a standard of 1×10.sup.4 spores gram.sup.−1 of seed.

(15) A homogeneous mix of is attained through tumbling seed and carnauba wax formulation in a cylinder, adapted to produce lateral mixing/tumbling through the inclusion of angled interior vanes, placed on a Wheaton roller for 5 minutes.

(16) Chitosan

(17) Chitosan (≧75% Deacetylated chitin, Poly(D-glucosamine)) (Sigma Aldrich, UK) is crushed using a pestle and mortar and then passed through a 32 micron mesh sieve.

(18) This is combined with Carnauba wax particles (VMD 75 μm) at a ratio of 1:19 (Chitosan:Carnauba wax particles). A homogeneous mix of is attained through tumbling seed and carnauba wax formulation in a cylinder, adapted to produce lateral mixing/tumbling through the inclusion of angled interior vanes, placed on a Wheaton roller for 5 minutes.

(19) Treatments:

(20) 1. Carnauba wax particles and Mycorrhizae 2. Carnauba wax particles and Rock Phosphate 3. Carnauba wax particles and Chitosan 4. Mycorrhizae control 5. Rock Phosphate control 6. Chitosan control 7. Carnauba wax particle control (vehicle control) 8. Untreated Control

(21) Seeds are planted in two 84 well plug trays using moist seed potting compost (John Innes No. 2). The trays are placed in a Vitopod propagator (Greenhouse Sensations, UK) at 18° C. Moisture content (Brannan Soil Moisture Meter, Fisher Scientific, UK) and pH levels (Brannan Soil pH meter, Fisher Scientific, UK) are checked to ensure that the conditions are consistent across the tray. The order of the treatments is randomised (by row units) to reduce any unforeseen bias.

(22) At the true leaf stage the plants are carefully transplanted from the plugs to 7 cm square pots filled with a sterilised top soil. The macro-nutrient (nitrates, phosphates and potassium) content of the top soil is measured using a La Motte Model STH-4 soil testing kit and recorded. Six replicates for each treatment (48 plants) are randomly assigned to each of three propagators, and further randomised within the propagator (total=144 plants). The propagators are set at 13.5° C., 18° C. and 22.5° C. Light is provided on a 16:8 Light:Dark cycle using a twin bulb T5 lighting array suspended 150 mm above the propagator (Lightwave T5, 48 w, 3300 lumens). T5 tubes (6500 Kelvin) deliver the bright blue/white light required by the plant for growth without emitting much heat which may scorch tender seedlings

(23) Moisture content and pH levels are checked to ensure that the conditions are consistent across the propagator by measuring six random plants along a pathway (alternating between a W and Z). This is repeated for each propagator.

(24) Plants are watered as required based on conditions to maintain consistent soil moisture content of 15% throughout all plants.

(25) The lids of the propagators are removed at such time as required due to the plant height.

(26) After 21 days the plants are removed from the propagators and the following measurements recorded:

(27) Root weight (fresh)

(28) Shoot weight (fresh)

(29) % Mycorrhyzal root colonisation (by microscopic examination)

(30) Plant tissue is measured for macro-nutrient content using the instructions provided with a La Motte Model PT-3R Plant Tissue Test kit.

(31) Analysis

(32) The percentage data (root colonisation data) were arcsine transformed. The influence of the factors and their interactions are tested with a two-way analysis of variance. Where the ANOVA reveals significant effects by the factors, the differences between treatments are separated using a post hoc least significant difference (LSD), multiple comparison test (p≦0.05).

(33) The influence of the factors and their interactions are tested with a 2-way ANOVA. The analysis was done for each temperature separately and with temperature as a factor. For the ANOVA with temperature as a factor, treatments were used as a sub-plot factor. Fisher's Least Significance Differences were calculated at the 5% significance level to compare treatment means. Shapiro-Wilks's test was performed to test for non-normality

(34) The above procedures are followed to apply rock phosphate, chitosan and Glomus sp. to seeds of Corn (Zea mays).

(35) The above procedures are followed to apply rock phosphate, chitosan and Glomus sp. to seeds of Rice (Oryza sativa).

(36) The above procedures are followed to apply rock phosphate, chitosan and Glomus sp. to seeds of Sorghum (Sorghum bicolor).

(37) Delivery of Macronutrients Using Carnauba Wax Particles as a Seed Coating on Wheat Aim:

(38) to assess the potential for formulating essential macronutrients into carnauba wax particles and using this as a seed coating to provide the germinating seed and seedling with supplementary nutrients to aid in early stage growth.

(39) Macronutrients Selected:

(40) Phosphorus (P)

(41) Phosphorus (P) is an essential part of the process of photosynthesis. Involved in the formation of all oils, sugars, starches, etc. Helps with the transformation of solar energy into chemical energy; proper plant maturation; withstanding stress. Effects rapid growth. Encourages blooming and root growth.
Potassium (K) Potassium is absorbed by plants in larger amounts than any other mineral element except nitrogen and, in some cases, calcium. Helps in the building of protein, photosynthesis, fruit quality and reduction of diseases. Potassium is supplied to plants by soil minerals, organic materials, and fertilizer.

(42) Both Potassium and Phosphorus can be found in soluble form in Monobasic Potassium Phosphate or MKP (KH.sub.2PO.sub.4), a soluble salt commonly used as a fertiliser and plant growth supplement.

(43) Formulation Method

(44) Carnauba is heated on a hotplate at 100° C. to a molten state. Monopotassium phosphate (MKP) is dissolved in deionised water to the required concentration. The MKP solution is slowly added to the molten wax under stirring at 1500 rpm. Stirring continues for 5 minutes before the water/wax emulsion is poured onto a metal sheet to cool. The resulting solid wax including micro-droplets of MKP is then micronized in an air mill to a VMD of approx. 10.3.

(45) Carnauba Wax Sizing Method

(46) Steps in Air Milling in Boyes Micronisation Process (for carnauba wax particles with a VMD of approx. 10.3 μm)

(47) 1. 2 kg carnauba wax blocks are first kibbled into approximately 4 to 6 mm pieces in a KT Handling Ltd Model 04 kibbler (serial no. 729/C) following the manufacturer's instructions.

(48) 2. The kibbled pieces are then passed through an Apex Construction Ltd Model 314.2 Comminuting Mill (serial no. A21306) and reduced further in size to a range of 250 to 300 um.

(49) 3. The comminuted particles are then passed through a Hosokawa Micron Ltd Alpine 100AFG jet mill (serial no. 168092) following the manufacturer's instructions, setting the mill at a speed of 12,500 rpm with a positive system pressure of 0.03 bar.

(50) 4. The grinding air is to be kept to 6 bar, the system rinsing air flow and Classifying Wheel gap rinsing air are both to be set at a minimum of 0.5 bar and no more than 0.75 bar, the cleaning air filter is to register a delta of no more than 5 bar to achieve a final particle size with a VMD of approx. 10.3 um.

(51) Experimental Method

(52) Wax particles containing 10% MKP are added to 10 g of Wheat seed, cv. Hereward (Herbiseeds, UK) at loadings of 0.1% and 1% by mass. Seed is well mixed to ensure a homogenous distribution across the seed. A third batch of seed is combined with unformulated carnauba wax particles as a control.

(53) 10 seeds for each treatment are sown in 20 cell modular seed trays with an individual cell size: Length 37 mm×Width 37 mm×Depth 65 mm, with each tray representing a single sample. Each treatment is replicated four times.

(54) The pots are filled with a sieved, heat-sterilised seed mix (Levingtons F1 Seed and Modular Compost—Low Nutrient)) to level with the top of the cell. Low Conductivity: 250-280 μS, Standard pH: 5.3-5.7, Mg/liter added: N—100, P—200, K—200.

(55) They are then lightly tamped and 30 ml of deionised water added to each cell through a course filter. A single seed is then placed on the surface of the soil and covered with a thin layer of vermiculite to a depth of 2-3× the diameter of the seed, as per supplier recommendation. The trays are placed within plastic gravel trays (two per tray) which are lined with capillary matting to aid watering.

(56) The gravel trays are then placed in a thermostatically controlled plant growth chamber (Fitotron SGC120, Weiss Gallenkamp, Loughborough, UK). Temperature cycling is set at 20° C./10° C. on a 16/8 hr schedule. Lighting at 150 μmol m.sup.−2 .sup.s.sup.−1 on a 16/8 hr photoperiod is introduced at first emergence.

(57) Plants are watered daily from the bottom in order to maintain a compost moisture level of approximately 40% in the cells

(58) After 10 days the plants are removed from the individual cells and the compost mix separated from the root structure. Plants from each 10 cell tray are combined and separated into shoots, made up of the first true leaves and growing tip, and roots.

(59) Plant Tissue Analysis: (Conducted to the Following Method by NRM Laboratories (Bracknell, UK)

(60) Total Phosphorus (P), Potassium (K), Magnesium (Mg), Calcium (Ca), Sodium (Na), Manganese (Mn), Copper (Cu), Iron (Fe), Zinc (Zn), Boron (B) determination using Inductively Coupled Plasma-Optical Emission Spectroscopy (ICP-OES) method

(61) Equipment

(62) 1. ICP Emission Spectrograph 2. Autosampler 3. Digital Dilutor
Reagents 1. 3N Hydrochloric Acid: Dilute 250 ml. of concentrated hydrochloric acid to 1 liter using deionized water and mix well. 2. Nitric Acid: HNO.sub.3
Standards 1. Stock Solutions: Use 1000 PPM certified, NIST traceable, plasma grade standards for the 16 listed elements. 2. Instrument Calibration Standards: a. WAT 1—deionized water b. WAT 2—Mn, Fe, Al, B, Cu, Zn, Na, Pb, Cd, Ni, Cr, Mo—Pipet 10 ml. of stock solution of each element into a 1 liter volumetric flask. Add 5 ml. of nitric acid. Dilute to volume with deionized water and mix well. c. PLN2—P, K, Ca, Mg—Pipet the designated ml. of stock solution into a 1 liter volumetric flask. Add 5 ml. of nitric acid. Dilute to volume with deionized water and mix well.

(63) TABLE-US-00001 Stock Final Solution Concentration Instrument Element ml. ppm. Readout % P 10 10 1.00 K 50 50 5.00 Ca 20 20 2.00 Mg 10 10 1.00 3. Instrument Calibration Verification Standards: a. A second set of calibration standards obtained from a different manufacturer.
Sample Preparation

(64) Samples are dried and ground to pass through a 1 mm screen.

(65) The elements in the residue remaining after the destruction of the organic matter by ashing at 550° C. are dissolved in hydrochloric acid: 1. Dry Ash a. Weigh 1 g sample into a 10 ml. glazed, high-form porcelain crucible. b. Ash in a muffle furnace for 4 hours at 500 C. c. Let cool and add 5 ml. of 3N HCl. d. Place on a hot plate and boil gently for 5 minutes. e. Let cool and transfer to a 100 ml. volumetric flask. Dilute to volume with deionized water and mix well. Use this solution for the analysis of Mn, Fe, Al, B, Cu, Zn, Na, Pb, Cd, Ni, Cr and Mo. f. Dilute the solution obtained in 1e. one to ten with deionized water using a digital dilutor. Use this solution for the analysis of P, K, Ca and Mg.
ICP Procedure 1. Set up and operate the ICP Emission Spectrograph in accordance with manufacturer's specifications. 2. Mn, Fe, B, Cu, Zn, Na, Ni analysis. a. Choose PLANT from the method menu. b. Calibrate the instrument using WAT1 and WAT2 instrument calibration standards. c. Analyze the sample digests obtained in 1e. of the sample preparation section. 3. P, K, Ca, Mg analysis. a. Choose PLANTDIL from the method menu. b. Calibrate the instrument using WAT1 and PLN2 instrument calibration standards. c. Analyze the digests obtained in 1f. of the sample preparation section.
Quality Control 1. Following calibration, analyze one high instrument calibration standard, one instrument calibration verification standard and one quality control sample. a. Instrument Calibration Standard: Values must be within 3% of the known value for K and Mo. All other elements must be within 2% of the known value. b. Instrument Calibration Verification Standard: Values must be within 10% of the certified values. c. Quality Control Sample: Values for all elements must be within limits established by the Extension chemist. 2. Analyze a high instrument calibration standard after each tenth sample and at the end of the set of samples. a. Values must be within 8% of the known values. b. If any of the values are greater than 8% from the known values, recalibrate the instrument and begin sample analysis from the last “good” instrument calibration standard. 3. Prepare one duplicate sample for each 10 samples. If the set contains less than 10 samples, prepare one duplicate per set. a. Results on the duplicate sample should agree within 20% of the average value of the two samples.

REFERENCES

(66) 1. Isaac, R. A. and W. C. Johnson, 1985, Elemental Analysis of Plant Tissue by Plasma Emission Spectroscopy Collaborative Study. JAOAC. 68(3), pp 499-505.

(67) 2. AOAC Official Method 985.01, in Official Methods of Analysis of AOAC International, 16th edition, Volume I Chapter 3, p. 4.

(68) 3. AOAC Official Method 968.08 D(a), in Official Methods of Analysis of AOAC International, 16th edition, Volume I Chapter 4, p. 23.

(69) Phosphate Solubilisation Using Beneficial Microbes

(70) Several bacterial species are able to impart a beneficial effect upon plant growth. Mostly they are associated with the plant rhizosphere, so they are called as rhizobacteria. This group of bacteria has been termed plant growth promoting rhizobacteria, and among them are strains from genera such as Alcaligenes, Acinetobacter, Arthrobacter, Azospirillum, Bacillus, Burkholderia, Enterobacter, Erwinia, Flavobacterium, Paenibacillus, Pseudomonas, Rhizobium, and Serratia.

(71) The production of organic acids by phosphate solubilizing bacteria has been well documented and identified as the main mechanism for phosphate solubilisation. Gluconic acid seems to be the most frequent agent of phosphate solubilisation (Pseudomonas sp.), and 2-ketogluconic acid is also identified in strains with phosphate solubilizing ability (Rhizobium sp.).

(72) Saprophytic fungi are also known to solubilise both organic and inorganic phosphates. Several genus, including Trichoderma, Penicillium, and Gliocladium have exhibited potential as biofertilisers. Morales et al (2011) demonstrated that Penicillium albidum was able to solubilise 64 mg of organic/inorganic phosphate per gram of fungi.

(73) Experiment to Assess the Potential for Delivery of Phosphate Solubilising Organisms as a Seed Costing Using Carnauba Wax Particles

(74) Using a dry spore powder of a phosphate solubilising organism, such as Penicillium bilaii.

(75) Spores are combined with carnauba wax particles with a VMD of approximately 10 μm at a ratio of 1:3. The powders are agitated to create a homogenous mix and applied to sterilised wheat seed at a loading of 0.1% (by mass). Additional batches of seed are treated with spores only (0.1%), Entostat only (0.1%) and untreated seed.

(76) Phosphate Solubilising Activity Screening

(77) Plate screening using Pikovskays' medium (see below) is used to demonstrate phosphate solubilising activity of the treated seed. 9 cm petri dishes are divided into quadrants and a seed is placed in the centre of each quadrant. Plates are incubated at 20° C. for 4 days.

(78) Active phosphate solubilising agents produce clear zones around the seed as they solubilise the insoluble mineral phosphates within the media. The radius of the clear zones is measured and compared to the mean results achieved for each treatment. Differences are analysed using one-way ANOVA and Tukey Post-Hoc diagnostic test where significance is found.

(79) Phosphate Uptake by Plant

(80) Seeds are treated as described above.

(81) Ca.sub.3(PO.sub.4).sub.2 is used as a source of insoluble phosphate.

(82) Sure to Grow PET grow cubes (25×25×38 mm) are soaked in deionised water containing 1% Ca.sub.3(PO.sub.4).sub.2 in suspension until saturated. Cubes are placed in free draining plant trays on a level surface to prevent nutrient run-off and migration whilst taking care to avoid pooling of water at the root zone. 10 cubes are used per tray and the mean of these represents one replicate. Each treatment is replicated 8 times.

(83) A single wheat seed is placed in the cross-cut X in the top of each cube. Seed trays are then covered to maintain a humid environment and regularly top watered with the 1% Ca.sub.3(PO.sub.4).sub.2 suspension to maintain a moist cube. Trays are incubated at 20° C. and 10° C. on a 16/8 hr thermal cycle. On germination the cover is removed and the seedling exposed to lighting on a 16/8 hr photoperiod.

(84) After 15 days the plants are removed from the grow cube and nutrient content of the plant tissue is analysed using the ICP method described above.

(85) Differences in the Phosphate content between treatments are assessed statistically using one-way ANOVA.

(86) Pikovskays' Medium

(87) TABLE-US-00002 Components Quantities (g l.sup.−1) Glucose 10 Ca.sub.3(PO.sub.4).sub.2 5 (NH.sub.4).sub.2SO.sub.4 0.5 NaCl 0.2 MgSO.sub.4•7H.sub.2O 0.1 KCI 0.2 Yeast Extract 0.5 MnSO.sub.4•H.sub.2O 0.002 FeSO.sub.4•7H.sub.2O 0.002 pH 7.0