CN106434425A - Rhodopseudomonas-palustris culturing medium and preparing method and application of culturing substance of rhodopseudomonas-palustris culturing medium - Google Patents
Rhodopseudomonas-palustris culturing medium and preparing method and application of culturing substance of rhodopseudomonas-palustris culturing medium Download PDFInfo
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- CN106434425A CN106434425A CN201610758945.9A CN201610758945A CN106434425A CN 106434425 A CN106434425 A CN 106434425A CN 201610758945 A CN201610758945 A CN 201610758945A CN 106434425 A CN106434425 A CN 106434425A
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- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
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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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- 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
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Abstract
The invention discloses a rhodopseudomonas-palustris culturing medium and preparing method and application of a culturing substance of the rhodopseudomonas-palustris culturing medium. The culturing medium is prepared from (NH4)2SO4, CH2COONa, YE and K2HPO4 in a mixed mode. The preparing method of the culturing substance of the rhodopseudomonas-palustris culturing medium includes the steps that target bacterial strains are separated and enriched, then the strains are activated, amplification culturing is carried out, split charging, sealing and storing are carried out after fermented culturing, and the culturing substance is obtained. The invention further discloses the application of the culturing substance of the rhodopseudomonas-palustris culturing medium in the aspects of promoting growth of cabbages and improving the yield of cabbages. The culture medium is low in production cost and convenient to use, and the culturing substance has high fertilizer efficiency, can obviously promote increasing of the yield of cabbage crops, can be effectively applied to safe planting of cabbages in agricultural production, can improve the quality of cabbages, can increase the yield of cabbages, and has the safe and environment-friendly application characteristics.
Description
Technical field
The present invention relates to cultivation and the application of bacterium, specifically a kind of Rhodopseudomonas palustris culture medium and cultivation thereof
The preparation method and application of thing.
Background technology
China arable land in the not enough world 10%, has supported the population of 22%.The behind of this achievement, is to have used in the world
The chemical fertilizer of 35% and agricultural chemicals！After agricultural chemicals, chemical fertilizer, additive etc. become " conventional weapon " of modern agriculture, Chinese arable land acidifying,
Salination, pollution problem are day by day serious.
Due to the Long-Time Service of chemical fertilizer, agricultural chemicals, China's widespread pollution from the overuse of fertilizers and pesticides in rural area problem is also quite prominent, mainly include chemical fertilizer,
The usage amount of agricultural chemicals is higher, and utilization rate is not high.For the prevention and control of agricultural pollution, the Ministry of Agriculture proposes to the year two thousand twenty, and control agricultural is used
Water inventory and Agricultural Water environmental pollution, farmland irrigation water water quality reaching standard, chemical fertilizer, pesticide reduction use.
For protruded above problem, in January, 2015, the Ministry of Agriculture reviews and passes in principle《Fertilizer application amount zero growth rate row
Dynamic scheme》With《Pesticide use amount zero growth rate action scheme》, and propose first the year two thousand twenty Chinese agriculture to realize " one control two
It is basic to subtract three "：Control agricultural water total amount；Reducing chemical fertilizer, Pesticide use amount, chemical fertilizer, pesticide dosage realize zero growth rate；In reply
Reducing amount of chemical fertilizer applied use, fertilization compositions based on earth measurement to be widelyd popularize, mechanized application, organic fertilizer application etc., support novel energetically
Management body and development are managed on a fairly large scale.In August, 2015, the General Office of the State Council issues《Change agricultural development side with regard to quickening
The suggestion of formula》Require, explore and implement fertilizer and chemical fertilizer rational proportion plan, encourage peasant's applying organic manure, support development height
Effect is slow（Control）Release the new-type fertilizers such as fertilizer, improve organic fertilizer application ratio and fertilizer utilization efficiency.
The agricultural problem severe for China and the agricultural development trend preferably adapting to country's future, the present invention designs
The fermentation process of a kind of microorganism photosynthetic bacteria bacterial manure and the application on pakchoi crop thereof, can preferably solve long-term
Use chemical fertilizer bring severe environments, soil issues.Function bacterium Rhodopseudomonas palustris can effectively improve crop pair
The utilization rate of chemical fertilizer simultaneously a certain degree of can prevent and treat crop pest, can reduce the usage amount of crop chemistry fertilizer, increase soil
The content of organic matter enriches soil microbial community, is a kind of ideal microbial manure.There is presently no marsh red false single
Relevant report in terms of born of the same parents bacterium and culture promotion pakchoi volume increase thereof.
Content of the invention
It is an object of the invention to provide a kind of Rhodopseudomonas palustris culture medium and culture thereof and application, can be effective
The problems such as the fertilizer efficiency utilization rate solving to bring because of Long-Time Service chemical fertilizer is low, agricultural environment environment, soil degradation, to solve
The problem proposing in above-mentioned background technology.
Rhodopseudomonas palustris involved in the present invention（Rhodopseudomonas palustris）By present patent application
People is isolated and purified to be obtained, and bacterial strain patent applied for is protected（The patent No.：CN201410750375.X）.Bacterial strain is stored in Chinese Typical Representative
Culture collection center（It is called for short CCTCC）, deposit number CCTCC No：M 2014524, depositary institution address is that Wuhan, China is big
Learning, preservation date is on October 29th, 2014.
For achieving the above object, the present invention provides following technical scheme：
A kind of Rhodopseudomonas palustris culture medium, is called for short PSBM culture medium, and the raw material by following content is formulated：(NH4)2SO4
0.5～4g/L, CH2COONa 0.5～4g/L, YE（Yeast extract）0.5～1.5g/L, K2HPO40.1～0.5g/
L.
The preparation method of the culture of a kind of described Rhodopseudomonas palustris culture medium, comprises the following steps that：
（1）Aimed strain separation and concentration：Take after the water sample natural sedimentation that will take and mix with the solid medium after thawing in right amount
After be down flat plate, illumination cultivation occurs to single bacterium colony, carries out illumination in single bacterium colony of random picking different shape to fluid nutrient medium
Cultivate 72～96h, be accredited as purpose bacterial strain；
（2）Actication of culture：The bacterial classification just separating is cultivated in described solid medium single bacterium colony, and rear picking list bacterium colony is to liquid
Illumination cultivation in body culture medium, condition of culture 25～35 DEG C, intensity of illumination 1500～2000 Lx；
（3）Expand and cultivate：The bacterial classification of activation is transferred in fluid nutrient medium with the inoculum concentration of 2～10%, continues training under the same terms
Support 72～96h, till culture becomes peony；
（4）Fermented and cultured：Will be enlarged by the bacterial classification after cultivating uses fluid nutrient medium to cultivate by the inoculum concentration of 1～5%, temperature
20～35 DEG C, fermented and cultured 5～7d under the conditions of intensity of illumination 500～1500 Lx；
（5）Zymotic fluid direct packaging seals and preserves, and makes the culture of Rhodopseudomonas palustris culture medium, i.e. photosynthetic bacteria bacterium
Agent, the microbial inoculum product viable count made can reach 500,000,000/more than mL.
A kind of application of the culture of described Rhodopseudomonas palustris culture medium, the training of Rhodopseudomonas palustris culture medium
Supporting application in terms of promoting Growth of Cabbage, raising yield of pakchoi for the thing, concrete applying step is as follows：The live final singling of pakchoi
Rear 5～10d starts to spray, and Rhodopseudomonas palustris culture is pressed 1:The thinner ratio of 200～300 is diluted, straight after dilution
Connect and spray in pakchoi blade face, spray once every 7～10d, spray continuously 2～3 times depending on growth cycle situation,.
Compared with prior art, the invention has the beneficial effects as follows：
Rhodopseudomonas palustris provided by the present invention is widely present in natural environment, and conventional method separates, and easily obtains；Cultivate
Method is simple, and incubation is not necessarily to the high-temperature and high-pressure conditions such as temperature control and oxygen supply, operates safety；Use bacterium prepared by bacterial strain of the present invention
Agent production low cost is easy to use, can significantly promote pakchoi crop yield, can be effectively applied to agricultural
The safety plantation of pakchoi in production；The bacterial strain of the present invention and culture thereof have higher fertilizer efficiency, can reduce planting process
The usage amount of middle chemical fertilizer；Nutriment contained by the bacterial strain of present invention offer and culture thereof and growth of microorganism are secreted
Active material, can effectively improve the resistance of pakchoi crop, and improve its quality and yield, has safety, green, ring
The application characteristic protected.
Brief description
Fig. 1 is the Electronic Speculum figure in the culture of Rhodopseudomonas palustris.
Fig. 2 be Rhodopseudomonas palustris culture medium culture dilution after design sketch.
Detailed description of the invention
Below in conjunction with the accompanying drawing in the embodiment of the present invention, the technical scheme in the embodiment of the present invention is carried out clear, complete
Describe, it is clear that described embodiment is only a part of embodiment of the present invention, rather than whole embodiments wholely.Based on
Embodiment in the present invention, it is every other that those of ordinary skill in the art are obtained under the premise of not making creative work
Embodiment, broadly falls into the scope of protection of the invention.
Embodiment 1
The culture medium of the present invention and strain isolation comprise the following steps：
（1）Culture medium is prepared：PSBM culture medium contains (NH4)2SO42g/L、CH2COONa2g/L、YE （Yeast extract, ferment
Female extract or yeast extract） 1.5g/L、K2HPO40.4g/L, is dissolved in 1L distilled water, natural ph, high pressure steam sterilization
30min, this is fluid nutrient medium；Sterilizing after the agar powder of solid PSBM culture medium interpolation 2%；
（2）Strain isolation, enrichment：The PSBM solid medium of thawing is cooled to 30～40 DEG C be advisable, adds in culture medium
The pond sludge top layer water sample that 1ml gathers, is down flat plate, is inverted in 25～30 DEG C after mixing, preferable temperature 28 DEG C, illumination is strong
7～10d is cultivated under the conditions of degree 1500～2000Lx；
（3）The internal red single colony inoculation of picking plating medium in the vial equipped with 100ml PSBM fluid nutrient medium,
The same terms continues to cultivate 84h, identifies and obtains purpose bacterial strain.
Embodiment 2
Use what Rhodopseudomonas palustris prepared microbial inoculum to specifically include following steps：
（1）Actication of culture：Mixing the bacterial strain of separation preservation with solid PSBM culture medium and being down flat plate, be inverted in 28 DEG C, illumination is strong
96h is cultivated, occurring to red colonies under the conditions of degree 1500～2000Lx；
（2）Spawn incubation：By step（1）Red single bacterium colony picking of middle cultivation is inoculated into equipped with 100mL PSBM fluid nutrient medium
Vial in, under the same terms cultivate 96h, reddening look to culture medium；
（3）Productive culture：By step（2）The bacterial classification of middle cultivation is transferred to equipped with 10L PSBM fluid nutrient medium with the inoculum concentration of 3%
Vial in, temperature 28 DEG C, fermented and cultured 6d under the conditions of intensity of illumination 1000 Lx；
（4）After cultivation terminates, culture seals packing and makes fermenting agent product.
Preparing fermenting agent, single used time in cycle 15d by this case method, prepared microbial inoculum living bacteria count is more than
Equal to 500,000,000/mL.
Embodiment 3
Rhodopseudomonas palustris and application in terms of improving yield of pakchoi for the fermenting agent thereof, specifically include following steps：
（1）Experimental field for local conventional arable land, physical features is smooth, and test area amounts to 60 m2, during test, before sowing, just execute the end
Outside Fei, no longer applying any other any fertilizer, other field management manage routinely and carry out；
（2）Test sets four process altogether：Ck：Clear water, M：PSBM culture medium dilutes 200 times, A：Microbial inoculum dilutes 200 times, B：Microbial inoculum
Dilute 300 times, every cell 6m2, each processes 2 times and repeats；
（3）Pakchoi sowing on September 22, grows about about 5cm and starts to spray dilution microbial inoculum, within every 7 days, spray once, spray 2 altogether
Secondary；October 10 sprayed for the first time, and October 17, second time was sprayed, and October 24 measured yield；Experiment overall process for cloudy day or
Fine day, without rainfall, strong wind weather；
（4）Gathering custom according to pakchoi, each cell plucks the maximum plant of naked eyes identification, and each cell randomly draws three
Point, totally three ten strain, weigh respectively, calculates equivalent yield and rate of growth；
As shown in Figure 2, microbial inoculum sprays pakchoi after diluting 200 times and 300 times to result, increases production respectively relative to clear water comparison
40.63% and 35.94%, under the conditions of extension rate 200 times, microbial inoculum process group yield is higher than medium treatment group, therefore incite somebody to action this
There is provided Rhodopseudomonas palustris is provided and culture is applied to pakchoi plantation, there is great using value.
It is obvious to a person skilled in the art that the invention is not restricted to the details of above-mentioned one exemplary embodiment, Er Qie
In the case of without departing substantially from the spirit or essential attributes of the present invention, the present invention can be realized in other specific forms.Therefore, no matter
From the point of view of which point, all should regard embodiment as exemplary, and be nonrestrictive, the scope of the present invention is by appended power
Profit requires rather than described above limits, it is intended that all by fall in the implication of equivalency and scope of claim
Change is included in the present invention.Any reference in claim should be considered as limiting involved claim.
Claims (3)
1. a Rhodopseudomonas palustris culture medium, it is characterised in that the raw material by following content is formulated：(NH4)2SO4
0.5～4g/L, CH2COONa 0.5～4g/L, YE 0.5～1.5g/L, K2HPO40.1～0.5g/L.
2. the preparation method of the culture of a Rhodopseudomonas palustris culture medium as claimed in claim 1, it is characterised in that
Comprise the following steps that：
（1）Aimed strain separation and concentration：Take after the water sample natural sedimentation that will take and mix with the solid medium after thawing in right amount
After be down flat plate, illumination cultivation occurs to single bacterium colony, carries out illumination in single bacterium colony of random picking different shape to fluid nutrient medium
Cultivate 72～96h, be accredited as purpose bacterial strain；
（2）Actication of culture：The bacterial classification just separating is cultivated in described solid medium single bacterium colony, and rear picking list bacterium colony is to liquid
Illumination cultivation in body culture medium, condition of culture 25～35 DEG C, intensity of illumination 1500～2000 Lx；
（3）Expand and cultivate：The bacterial classification of activation is transferred in fluid nutrient medium with the inoculum concentration of 2～10%, continues training under the same terms
Support 72～96h, till culture becomes peony；
（4）Fermented and cultured：Will be enlarged by the bacterial classification after cultivating uses fluid nutrient medium to cultivate by the inoculum concentration of 1～5%, temperature
20～35 DEG C, fermented and cultured 5～7d under the conditions of intensity of illumination 500～1500 Lx；
（5）Zymotic fluid direct packaging seals and preserves, and makes the culture of Rhodopseudomonas palustris culture medium, i.e. photosynthetic bacteria bacterium
Agent, the microbial inoculum product viable count made can reach 500,000,000/more than mL.
3. the application of the culture of a Rhodopseudomonas palustris culture medium as claimed in claim 2, it is characterised in that marsh
Application in terms of promoting Growth of Cabbage, raising yield of pakchoi for the culture of red pseudomonas culture medium, concrete application step
Rapid as follows：After the live final singling of pakchoi, 5～10d starts to spray, and Rhodopseudomonas palustris culture is pressed 1:200～300 dilute
Release ratio to be diluted, directly spray after dilution in pakchoi blade face, spray once every 7～10d, continuous depending on growth cycle situation
Spray 2～3 times,.
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| CN201610758945.9A CN106434425A (en) | 2016-08-30 | 2016-08-30 | Rhodopseudomonas-palustris culturing medium and preparing method and application of culturing substance of rhodopseudomonas-palustris culturing medium |
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Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN108587975A (en) * | 2018-05-16 | 2018-09-28 | 广西青又青生物肥业有限公司 | A kind of Rhodopseudomonas palustris culture medium and cultural method |
| CN113913450A (en) * | 2021-10-29 | 2022-01-11 | 湖南省植物保护研究所 | Method for expressing chitosanase by using rhodopseudomonas palustris, chitosanase, recombinant plasmid, recombinant bacterium, fermentation inoculant and application |
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| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102061279A (en) * | 2010-11-22 | 2011-05-18 | 河北省微生物研究所 | Method for producing rhodopseudomonas palustris fermentation liquor by high-density fermentation |
| CN102628024A (en) * | 2012-03-31 | 2012-08-08 | 广东绿百多生物科技有限公司 | Co-culture method of phycomycetes for promoting plant growth |
| CN104498399A (en) * | 2014-12-10 | 2015-04-08 | 湖南省植物保护研究所 | Rhodopseudomonas palustris strain, biological agent and preparation method and application of biological agent |
-
2016
- 2016-08-30 CN CN201610758945.9A patent/CN106434425A/en active Pending
Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102061279A (en) * | 2010-11-22 | 2011-05-18 | 河北省微生物研究所 | Method for producing rhodopseudomonas palustris fermentation liquor by high-density fermentation |
| CN102628024A (en) * | 2012-03-31 | 2012-08-08 | 广东绿百多生物科技有限公司 | Co-culture method of phycomycetes for promoting plant growth |
| CN104498399A (en) * | 2014-12-10 | 2015-04-08 | 湖南省植物保护研究所 | Rhodopseudomonas palustris strain, biological agent and preparation method and application of biological agent |
Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN108587975A (en) * | 2018-05-16 | 2018-09-28 | 广西青又青生物肥业有限公司 | A kind of Rhodopseudomonas palustris culture medium and cultural method |
| CN113913450A (en) * | 2021-10-29 | 2022-01-11 | 湖南省植物保护研究所 | Method for expressing chitosanase by using rhodopseudomonas palustris, chitosanase, recombinant plasmid, recombinant bacterium, fermentation inoculant and application |
| CN113913450B (en) * | 2021-10-29 | 2024-02-02 | 湖南省植物保护研究所 | Method for expressing chitosanase by rhodopseudomonas palustris, chitosanase, recombinant plasmid, recombinant bacteria, fermentation bacteria and application |
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Application publication date: 20170222 |