CN105861334B - One plant of endogenetic fungus that acacia confusa biomass can be promoted to increase - Google Patents
One plant of endogenetic fungus that acacia confusa biomass can be promoted to increase Download PDFInfo
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- CN105861334B CN105861334B CN201610452938.6A CN201610452938A CN105861334B CN 105861334 B CN105861334 B CN 105861334B CN 201610452938 A CN201610452938 A CN 201610452938A CN 105861334 B CN105861334 B CN 105861334B
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- 241000220480 Acacia confusa Species 0.000 title claims abstract description 36
- 239000002028 Biomass Substances 0.000 title claims abstract description 33
- 241000196324 Embryophyta Species 0.000 title claims abstract description 27
- 241000233866 Fungi Species 0.000 title claims abstract description 23
- 230000001580 bacterial effect Effects 0.000 claims abstract description 21
- 241000221207 Filobasidium Species 0.000 claims abstract description 4
- 241000221566 Ustilago Species 0.000 claims abstract description 4
- 244000005700 microbiome Species 0.000 claims abstract description 4
- 238000004321 preservation Methods 0.000 claims abstract description 3
- 241000894006 Bacteria Species 0.000 claims description 24
- CSNNHWWHGAXBCP-UHFFFAOYSA-L Magnesium sulfate Chemical compound [Mg+2].[O-][S+2]([O-])([O-])[O-] CSNNHWWHGAXBCP-UHFFFAOYSA-L 0.000 claims description 8
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Chemical compound O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 8
- 239000002689 soil Substances 0.000 claims description 6
- 239000002609 medium Substances 0.000 claims description 5
- WQZGKKKJIJFFOK-GASJEMHNSA-N Glucose Natural products OC[C@H]1OC(O)[C@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-GASJEMHNSA-N 0.000 claims description 4
- 239000001888 Peptone Substances 0.000 claims description 4
- 108010080698 Peptones Proteins 0.000 claims description 4
- 229940041514 candida albicans extract Drugs 0.000 claims description 4
- 239000008103 glucose Substances 0.000 claims description 4
- 239000001963 growth medium Substances 0.000 claims description 4
- 238000011534 incubation Methods 0.000 claims description 4
- 229910052943 magnesium sulfate Inorganic materials 0.000 claims description 4
- 235000019341 magnesium sulphate Nutrition 0.000 claims description 4
- 235000019319 peptone Nutrition 0.000 claims description 4
- 239000000843 powder Substances 0.000 claims description 4
- 239000008223 sterile water Substances 0.000 claims description 4
- 239000012138 yeast extract Substances 0.000 claims description 4
- ZPWVASYFFYYZEW-UHFFFAOYSA-L dipotassium hydrogen phosphate Chemical compound [K+].[K+].OP([O-])([O-])=O ZPWVASYFFYYZEW-UHFFFAOYSA-L 0.000 claims description 3
- 239000012530 fluid Substances 0.000 claims description 3
- 235000015097 nutrients Nutrition 0.000 claims description 3
- 238000002360 preparation method Methods 0.000 claims description 3
- 239000008280 blood Substances 0.000 claims description 2
- 210000004369 blood Anatomy 0.000 claims description 2
- 229910021642 ultra pure water Inorganic materials 0.000 claims description 2
- 239000012498 ultrapure water Substances 0.000 claims description 2
- 239000007788 liquid Substances 0.000 claims 1
- 238000011081 inoculation Methods 0.000 abstract description 4
- 238000005259 measurement Methods 0.000 abstract description 4
- 238000000034 method Methods 0.000 description 7
- 241000831652 Salinivibrio sharmensis Species 0.000 description 6
- 238000011160 research Methods 0.000 description 4
- 238000004659 sterilization and disinfection Methods 0.000 description 4
- 238000012545 processing Methods 0.000 description 3
- 230000001737 promoting effect Effects 0.000 description 3
- 239000007787 solid Substances 0.000 description 3
- NBIIXXVUZAFLBC-UHFFFAOYSA-N Phosphoric acid Chemical compound OP(O)(O)=O NBIIXXVUZAFLBC-UHFFFAOYSA-N 0.000 description 2
- ZLMJMSJWJFRBEC-UHFFFAOYSA-N Potassium Chemical compound [K] ZLMJMSJWJFRBEC-UHFFFAOYSA-N 0.000 description 2
- 239000000463 material Substances 0.000 description 2
- 239000011591 potassium Substances 0.000 description 2
- 229910052700 potassium Inorganic materials 0.000 description 2
- 238000000926 separation method Methods 0.000 description 2
- 230000001954 sterilising effect Effects 0.000 description 2
- 238000012360 testing method Methods 0.000 description 2
- 238000005406 washing Methods 0.000 description 2
- 229920001817 Agar Polymers 0.000 description 1
- 108090000790 Enzymes Proteins 0.000 description 1
- 102000004190 Enzymes Human genes 0.000 description 1
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 description 1
- 241000238631 Hexapoda Species 0.000 description 1
- UFHFLCQGNIYNRP-UHFFFAOYSA-N Hydrogen Chemical compound [H][H] UFHFLCQGNIYNRP-UHFFFAOYSA-N 0.000 description 1
- 244000048927 Lolium temulentum Species 0.000 description 1
- OAICVXFJPJFONN-UHFFFAOYSA-N Phosphorus Chemical compound [P] OAICVXFJPJFONN-UHFFFAOYSA-N 0.000 description 1
- 241000209504 Poaceae Species 0.000 description 1
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 description 1
- 239000005708 Sodium hypochlorite Substances 0.000 description 1
- 239000008272 agar Substances 0.000 description 1
- 229910000147 aluminium phosphate Inorganic materials 0.000 description 1
- 230000009286 beneficial effect Effects 0.000 description 1
- 230000000443 biocontrol Effects 0.000 description 1
- 238000009395 breeding Methods 0.000 description 1
- 230000001488 breeding effect Effects 0.000 description 1
- 238000001514 detection method Methods 0.000 description 1
- 238000011161 development Methods 0.000 description 1
- 201000010099 disease Diseases 0.000 description 1
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 description 1
- 239000012153 distilled water Substances 0.000 description 1
- 238000009826 distribution Methods 0.000 description 1
- 238000001035 drying Methods 0.000 description 1
- 238000011156 evaluation Methods 0.000 description 1
- 238000002474 experimental method Methods 0.000 description 1
- 238000003958 fumigation Methods 0.000 description 1
- 230000002538 fungal effect Effects 0.000 description 1
- 229910052739 hydrogen Inorganic materials 0.000 description 1
- 239000001257 hydrogen Substances 0.000 description 1
- 238000009630 liquid culture Methods 0.000 description 1
- 238000004519 manufacturing process Methods 0.000 description 1
- 238000002156 mixing Methods 0.000 description 1
- 238000012986 modification Methods 0.000 description 1
- 230000004048 modification Effects 0.000 description 1
- 238000002161 passivation Methods 0.000 description 1
- 239000011574 phosphorus Substances 0.000 description 1
- 229910052698 phosphorus Inorganic materials 0.000 description 1
- 238000007747 plating Methods 0.000 description 1
- 238000004153 renaturation Methods 0.000 description 1
- 230000010076 replication Effects 0.000 description 1
- 239000011780 sodium chloride Substances 0.000 description 1
- SUKJFIGYRHOWBL-UHFFFAOYSA-N sodium hypochlorite Chemical compound [Na+].Cl[O-] SUKJFIGYRHOWBL-UHFFFAOYSA-N 0.000 description 1
- 239000008399 tap water Substances 0.000 description 1
- 235000020679 tap water Nutrition 0.000 description 1
- 238000012549 training Methods 0.000 description 1
- 238000005303 weighing Methods 0.000 description 1
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- C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
- C12N1/14—Fungi; Culture media therefor
- C12N1/145—Fungal isolates
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- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01N—PRESERVATION OF BODIES OF HUMANS OR ANIMALS OR PLANTS OR PARTS THEREOF; BIOCIDES, e.g. AS DISINFECTANTS, AS PESTICIDES OR AS HERBICIDES; PEST REPELLANTS OR ATTRACTANTS; PLANT GROWTH REGULATORS
- A01N63/00—Biocides, pest repellants or attractants, or plant growth regulators containing microorganisms, viruses, microbial fungi, animals or substances produced by, or obtained from, microorganisms, viruses, microbial fungi or animals, e.g. enzymes or fermentates
- A01N63/30—Microbial fungi; Substances produced thereby or obtained therefrom
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- C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
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Abstract
The endogenetic fungus that acacia confusa biomass can be promoted to increase the present invention relates to one plant.The endogenetic fungus be silk Ustilago (Filobasidium) B, preservation is registered in China Committee for Culture Collection of Microorganisms's common micro-organisms center on January 27th, 2016, deposit number is CGMCC No.11910.Acacia confusa endogenetic fungus B is inoculated in acacia confusa earth culture seedling, the biomass of acacia confusa after measurement inoculation 60 days, plant handled by acacia confusa endogenetic fungus B and the biomass difference compareed are extremely significant, biomass compares display, the bacterial strain works well compared with the growth-promoting of control group, and biomass is 1.4 times of control group；Root/shoot ratio compares display, and the root/shoot ratio of the bacterial strain is 1.5 times of control, shows that the bacterial strain has apparent facilitation to the growth of acacia confusa biomass.
Description
Technical field
The endogenetic fungus that acacia confusa biomass can be promoted to increase the present invention relates to one plant.
Background technique
The research of endophyte of plant starts from the end of the 19th century, and Vogl divides from rye grass Lolium temulentum L. seed
Separate out first plant of endophyte.But the endophyte really started in numerous studies plant originates in last century the eighties, mainly
It conducts a research in the vegetation of Temperate Region in China, subtropical zone and torrid areas.
The distribution of plant endogenesis epiphyte is wide, type is more, and forefathers out of most plants research shows that can separate
To endogenetic fungus, it can be seen that endogenetic fungus is prevalent in plant.At least belong to 290 at more than 80 in worldwide
Endophyte is had found in a variety of gramineae farm crops.The unique function that the endo-mycorrhiza separated from plant at present has according to it
Can be divided into: nitrogen-fixing bacteria consolidate plant growth-promoting bacterias, the biocontrol microorganisms with disease and insect resistance such as potassium bacterium, solid phosphorus bacterium and have promotion plant
To the resistance bacterium of the repair ability of poor environment.
In relation to endogenetic fungus, to the influence research that acacia confusa is grown, there is also blank.
Summary of the invention
The purpose of the present invention is to provide the endogenetic fungus that one plant of bacterial strain can promote acacia confusa biomass to increase, the fungies
For silk Ustilago (Filobasidium) B, it is general in China Committee for Culture Collection of Microorganisms on January 27th, 2016
Preservation is registered at logical microorganism center, and address is Yard 1, BeiChen xi Road, Chaoyang District, Beijing City 3, and deposit number is CGMCC No.
11910。
To achieve the above object, the present invention adopts the following technical scheme:
The separation of the bacterium, purifying include:
A, material: being divided into three parts for the root for collecting different year, branch stem, leaf, cleaned up with tap water, point
It is attached in valve bag, is saved in 4 DEG C of refrigerators；
B, sterilize: 75% alcohol rinses 30s → aseptic water washing 3-4 times → 15% sodium hypochlorite and rinses (root 10min, stem
5min, leaf 2min) → aseptic water washing 4-5 times.In the small beaker that water after sample last time is embathed is contained in sterilizing, flat
Whether lining out is used as control, thorough with the disinfection of sample survey, generates behind several days if any bacterium colony, then shows sample surfaces
Disinfection is not thorough, and need to continue to adjust sterilization method；
C, the selection of inoculation position: blade: leaf tip, leaf central part and leaf base 3 processing；Branch stem: upper, middle and lower is divided into 3
Position, wherein the 3rd position is branch stem junction；Root: it is divided into 2 parts of the tip of a root and foundation；
D, it is inoculated with: the explant after disinfection being cut with vaccinating lancet, is laid in media surface, in 28 DEG C of constant temperature incubations
Bacterium colony upgrowth situation is observed after culture 5-7 days in case.Rapidly, the bacterial strain that can be first generated is separated for some bacterial strain breedings
Purifying；The bacterial strain of slow growth and negligible amounts can extend observing time, purify after its growth is stablized；
Solid culture: using improvement Martin's solid medium, be formulated for peptone 5.0g/L, yeast extract powder 2.0g/L,
Glucose 20.0g/L, dipotassium hydrogen phosphate 1.0g/L, magnesium sulfate 0.5g/L, agar 14.0g/L, it is other be sterile water, pH value 6.4
± 0.2, cultivation temperature is 28 DEG C, and training method is plate culture, incubation time 48h；
E, different types of endogenetic fungus access plating medium that separation is bred is purified, by 2-3 point
The above-mentioned of single culture is respectively obtained after connecing purifying, switchingFilobasidiumFunction stem；
A kind of preparation method of above-mentioned endogenetic fungus application bacterium solution that acacia confusa biomass can be promoted to increase, being will be upper
The bacterial strain access fluid nutrient medium stated, shaking table shaken cultivation, cultivation temperature is 28 DEG C, and 48~72h of incubation time utilizes hemocytometer
Number plate calculates bacterial concentration, and bacterium solution is diluted to 1.0-9.0 × 10 with ultrapure water6Cfu/ml is that finished product is spare；Liquid Culture
Base: to improve Martin's culture medium, wherein peptone 5.0g/L, yeast extract powder 2.0g/L, glucose 20.0g/L, phosphoric acid hydrogen two
Potassium 1.0g/L, magnesium sulfate 0.5g/L, it is other be sterile water, pH value 6.4 ± 0.2.
A kind of application method of above-mentioned endogenetic fungus application bacterium solution that acacia confusa biomass can be promoted to increase, is application
The bacterium solution of the bacterial strain, which is poured, imposes on acacia confusa Seedling rhizosphere soil or nursery stock is directly inoculated with.
A kind of application method of above-mentioned endogenetic fungus application bacterium solution that acacia confusa biomass can be promoted to increase, is to use
5.5×106Cfu/ml bacterium solution is poured in forest land in the rhizosphere of every plant of acacia confusa by every plant of 100ml.
Bacterial strain of the present invention is isolated and purified from acacia confusa plant, and acacia confusa earth culture is inoculated in
Seedling further confirms that it has the work for promoting acacia confusa biomass to increase to plant strain growth according to indices Comprehensive Evaluation
With search out can be applied to produce to the beneficial function endogenetic fungus for promoting biomass to increase.
The method applied is poured by bacterium solution and is inoculated in acacia confusa seedling, is measured when being just inoculated with April and when the 60 day June of inoculation
Biomass, the root/shoot ratio of acacia confusa.Plant handled by acacia confusa endogenetic fungus B and the biomass, root/shoot ratio that compare are poor
It is different significant.Biomass compares display, and bacterial strain B processing works well compared with the growth-promoting of control group, and biomass is the 1.4 of control group
Times.Root/shoot ratio compares display, and the root/shoot ratio of bacterial strain B is 1.5 times of control.Show growth of the bacterial strain B to acacia confusa biomass
With apparent facilitation.
Detailed description of the invention
Fig. 1 is influence of the different disposal to acacia confusa Seedling Biomass；
Fig. 2 is influence of the different disposal to acacia confusa seedling root/shoot ratio.
Specific embodiment
Rhizosphere soil, which pours, grants nursery stock and is directly inoculated with application
This test uses earth culture pot experiment, and test material therefor is the Taiwan phase that Municipal Forestry Bureau, Zhangzhou City, Fujian Province provides
Think one year seedling.Select the consistent acacia confusa seedling planting of growing way in diameter 15cm, the plastic tub of high 10cm.Soil used
It is the stringent yellow soil Jing Guo fumigation.It is weighed by mixing, every basin is put into the 3kg soil of equivalent.It is extensive by one month
After renaturation growth, it is 5.5 × 10 that continuous three days, which apply concentration in acacia confusa rhizosphere,6The 100mL bacterium solution of cfu/ml, uses distilled water
Solution is as blank control.
Bacterium solution preparation: by the fluid nutrient medium of strains tested access 40mL, culture medium is improvement Martin's culture medium, wherein egg
It is white peptone 5.0g/L, yeast extract powder 2.0g/L, glucose 20.0g/L, dipotassium hydrogen phosphate 1.0g/L, magnesium sulfate 0.5g/L, other
For sterile water, pH value 6.4 ± 0.2, the culture of 72h is passed through in constant-temperature shaking incubator.It is dilute by ten times with sterile saline
Interpretation of the law dilutes bacterium solution, calculates bacterial concentration using blood counting chamber and is configured to 5.5 × 106cfu/mL.Inoculation carries out after 60 days
The measurement of the indexs such as height of seedling, ground diameter, detection method are as follows:
The measurement of biomass
The different parts of plant are respectively put into envelope and are put signs on, 105 DEG C of drying half an hour (passivation of baking oven are put into
Enzyme), it dries, after 12h, is placed into baking oven after weighing, replication is until constant weight at 60 DEG C.Claim to obtain sample
Weight (W), as sample dry weight, the biomass of plant is indicated with this, is accurate to 0.01g.
As a result with analysis
Influence of the different disposal to acacia confusa biomass and root/shoot ratio
Biomass is the important indicator for reflecting nursery stock production power level, and there is very close for the growth and development of it and nursery stock
Relationship.As shown in Figure 1, the biomass of the seedling of inoculating strain B is bigger than control group by 37.12%, and in extremely significant between control group
Otherness (sig=0.008, the level of signifiance 0.01), thus, growth of the bacterial strain to the biomass of acacia confusa seedling
With extremely significant facilitation.Shown in Fig. 2, the root/shoot ratio control group of the seedling of inoculating strain B is big by 46.89%, and and control group
Between extremely significant otherness (sig=0.005, the level of signifiance 0.01), illustrate increasing of the bacterial strain B to acacia confusa seedling root/shoot ratio
There is greatly extremely significant facilitation.
Present invention discover that the endogenetic fungus that two plants of hybrid bacterial strains can promote acacia confusa seedling biomass to increase, by this plant of Taiwan
Yearning between lovers endogenetic fungal bacterial strain is inoculated in acacia confusa seedling using the method for pouring root.Pass through biomass to plant, root/shoot ratio
Measurement show that the biomass for being inoculated with the processing of the bacterial strain, root/shoot ratio increase and is largely higher than control, shows it for promoting
The growth of biomass has very big function, to further confirm that obtaining this plant of endogenetic fungus can promote acacia confusa biological
The growth of amount.
The foregoing is merely presently preferred embodiments of the present invention, all equivalent changes done according to scope of the present invention patent with
Modification, is all covered by the present invention.
Claims (3)
1. one plant of endogenetic fungus that acacia confusa biomass can be promoted to increase, it is characterised in that: the fungi is silk Ustilago
(Filobasidium) B, on January 27th, 2016 in China Committee for Culture Collection of Microorganisms's common micro-organisms center
Preservation is registered, deposit number is CGMCC No. 11910.
2. a kind of application bacterium solution of the one plant as described in claim 1 endogenetic fungus that acacia confusa biomass can be promoted to increase,
It is characterized by: the preparation method using bacterium solution, by the silk Ustilago (Filobasidium) B bacterial strain access liquid
Body culture medium, shaking table shaken cultivation, cultivation temperature are 28 DEG C, 48~72h of incubation time, and it is dense to calculate bacterium solution using blood counting chamber
Degree, is diluted to 1.0 × 10 with ultrapure water for bacterium solution6-9.0×106Cfu/ml, it is spare；Fluid nutrient medium: for improvement Martin's culture
Base, wherein peptone 5.0g/L, yeast extract powder 2.0g/L, glucose 20.0g/L, dipotassium hydrogen phosphate 1.0g/L, magnesium sulfate
0.5g/L, it is other be sterile water, pH value 6.4 ± 0.2.
3. a kind of purposes for applying bacterium solution as claimed in claim 2, it is characterised in that: the application bacterium solution is used for acacia confusa
Seedling rhizosphere soil, which pours, to be applied or nursery stock is directly inoculated with.
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Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2014165174A2 (en) * | 2013-03-12 | 2014-10-09 | Strobel Gary A | Microorganisms for producing volatile organic compounds and methods using same |
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| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2014165174A2 (en) * | 2013-03-12 | 2014-10-09 | Strobel Gary A | Microorganisms for producing volatile organic compounds and methods using same |
Non-Patent Citations (2)
| Title |
|---|
| Identification and bioactive properties of endophytic fungi isolated from phyllodes of Acacia species;H.B.Q. Tran,et al;《Resrarchgate》;20141010;全文 * |
| 喀斯特生境台湾相思根瘤菌的生物学特性研究;吕成群等;《微生物学通报》;20080220(第02期);全文 * |
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