CN105112340B - One kind having the active bacterium bacterial strain of induction of resistance and its cultural method and application - Google Patents
One kind having the active bacterium bacterial strain of induction of resistance and its cultural method and application Download PDFInfo
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Abstract
It is bacillus thuringiensis (Bacillus thuringiensis) to have the active bacterium bacterial strain of induction of resistance and its cultural method and application, the bacterial strain the invention discloses one kind, and preserving number is CGMCC No.10831.The fluid nutrient medium of above-mentioned bacterium bacterial strain is prepared by the raw material of following mass percent：Yeast extract 0.1% 0.2%, peptone 0.5% 0.6%, beef extract 0.3% 0.4%, sucrose 1% 2%, remainder is distilled water.Bacillus thuringiensis (B.thuringiensise) Sneb1076 or its metabolin of the present invention have the induced activity to soybean, it can be used for the seed treatment of soybean, the growth of soybean plant strain and root can not only be promoted, there is good inhibiting effect to the formation of soybean cyst nematode Heterodera glycines cyst simultaneously, a new way is provided for soybean cyst nematode Heterodera glycines prevention.
Description
Technical field
The present invention relates to bacterial strain fields, and in particular to one kind having the active bacterium bacterial strain of induction of resistance and its cultural method
And application.
Background technology
Many countries and regions occur and cause harm plant nematode in the world, and type is varied.In the world
It is interior, the generation of plant nematode with harm with the development of agriculture and forestry and the variation getting worse of tillage and cultivation system, according to
Esser is counted, and the whole world, which has been reported, until nineteen ninety finds that plant nematode 207 belongs to 4832 kinds of (Liu Weizhi, section imperial jade seals
.2000. Beijing plant pathogeny line insect：Chinese agriculture publishing house, P.1-2).It is estimated that plant nematode causes world master
Take crops year loss late be 12.3%, more than 100,000,000,000 dollars, and it is actual loss considerably beyond estimation, reason be many
Harm caused by nematode does not arouse attention because not having field Visual symptoms or specialization feature, and (Wang Shou China Pomologies is northern
Capital：Chinese agriculture Science Press, 1994.P.1-5), in addition, also constantly increasing to the new discovery that nematode is caused harm in world wide
Add, and some newer agricultural cultivation systems keep nematode problem more prominent, furthermore, nematode and other biological interactions and
The direct or indirect influence that plant generates also is being sharply increased.In this sense, more other lifes are endangered caused by nematode
Object is more hidden！At present to the nematode of plant pest there are about more than 3000 kinds, mainly there is cyst nematode, Bursaphelenchus xylophilus, sweet in China
Potato Ditylenchus dipsaci etc..Cyst nematode is maximum a kind of nematode of agriculturally causing harm, after disease occurs, the general underproduction 10% or so, sternly
Up to 75% or more of weight, or even total crop failure (A.G.Whitehead, 1998.Plant Nematode
Control.ISBN0851991882CAB International).Currently, the prevention of plant nematode is still with chemical prevention
Based on, but in recent years, the application waste of nematocide is big, pollutes environment, and poor selectivity, natural disposition of going out is strong, destroys geobiont
The drawbacks such as fauna are increasingly paid attention to that (Zhang Keqin, He Shichuan, the .1991. such as all common vetch bacteriums and bacterium are in nematode biological and ecological methods to prevent plant disease, pests, and erosion by people
Effect and its progress insecticidal microorganisms .3：55-66), 21 century is known as environmentally friendly century, some effective chemistry kill line
The application of agent is gradually restricted, therefore becomes hot issue naturally to the biological control research of plant nematode.It has reported
The biocontrol bacteria main groups for preventing plant nematode have：Bacillius, Pseudomonas, Acinebacter,
Chromobacterium, Enterobacter, Serratia, Burkholderia etc., wherein with Bacillus and
Most species in Pseudomonas.Past is typically all to utilize BT toxalbumin pair to the Biocontrol Effect of bacillus thuringiensis
The biological control of insect directly has the active research of kill also to have similar report nematode, and processing vegetable seeds is generated
Induction of resistance defends the seldom of disease report, and for utilizing bacillus thuringiensis processed soybeans Seed inducement soybean plant strain root
System generates the research of anti-Plant nematode and invention is not reported.The metabolism induction plant of bacterial strain how is utilized to generate resistance,
The immune system for exciting plant is the key that the present invention is paid close attention to.
Invention content
In view of the above-mentioned problems, the present invention obtains a bacillus thuringiensis strain by screening system, the bacterium or the bacterium are utilized
Metabolite processed soybeans seed, can induce soybean and generated during Seed Germination and growth soybean cyst nematode Heterodera glycines are resisted
Property, to provide new approach to solve the problems, such as that soybean cyst nematode is done harm to.
To achieve the above object, the technical solution that the present invention takes is：
It is bacillus thuringiensis Bacillus that one kind, which having the active bacterium bacterial strain of induction of resistance, the bacterial strain,
Thuringiensis Sneb 1076, depositary institution：China Committee for Culture Collection of Microorganisms's common micro-organisms center；
Address：The Chinese Beijing Chaoyang Districts North Star West Road 1 institute 3；Preservation date：On May 21st, 2015；The volume that preservation is registered on the books
Number：CGMCC No.10831.B.thuringiensis Sneb 1076 are one plants filtered out from a large amount of soil bacteria bacterial strains
There is the bacterium bacterial strain of good induction of resistance to soybean cyst nematode Heterodera glycines.
A kind of above-mentioned cultural method with the active bacterium bacterial strain of induction of resistance, includes the following steps：
S1, it weighs by mass percentage：Yeast extract 0.1%-0.2%, peptone 0.5%-0.6%, beef extract
0.3%-0.4%, sucrose 1%-2%, remainder are distilled water, after mixing, obtain fluid nutrient medium；
S2, the fluid nutrient medium obtained by step S1 is sub-packed in the ratio of every bottle of 150mL in the triangular flask of 250mL, point
Strain is inoculated into fluid nutrient medium after the completion of dress, pH6.8-7.0, temperature is 25-28 DEG C, shaking speed 150rmin-1,
Ferment 2-3d, must can be used for the bacterium metabolite that inducing soybean generates anti-soybean cyst nematode resistance.
There is above-mentioned one kind the active bacterium bacterial strain of induction of resistance can be used for preparing plant nematode biological control induction
Resistance preparation.The induction of resistance preparation includes a effective amount of above-mentioned bacillus thuringiensis (B.thuringiensise) or step
Metabolite obtained by rapid S2.
Wherein, the plant nematode is cyst nematode (Soybean cyst nematode).
The present invention passes through the study found that filtering out the more plants of bacteriums with the anti-soybean cyst nematode Heterodera glycines of inducing soybean from soil
Bacterial strain, wherein bacillus thuringiensis (B.thuringiensise) Sneb1076 preventive effects are preferable, and stablize relatively.
The invention has the advantages that：
Bacillus thuringiensis (B.thuringiensise) Sneb1076 or its metabolin of the present invention has to soybean
Induced activity, can be used for the seed treatment of soybean, can not only promote the growth of soybean plant strain and root, while to soybean cyst
The formation of nematode cyst has good inhibiting effect, and a new way is provided for soybean cyst nematode Heterodera glycines prevention.
Specific implementation mode
In order to make objects and advantages of the present invention be more clearly understood, the present invention is carried out with reference to embodiments further
It is described in detail.It should be appreciated that the specific embodiments described herein are merely illustrative of the present invention, it is not used to limit this hair
It is bright.
Embodiment 1
The culture of bacillus thuringiensis (B.thuringiensise) Sneb1076
By in the inoculation to Tube propagation base of bacillus thuringiensis (B.thuringiensise) Sneb1076, train
It is beef-protein medium, i.e. ingredient to support based formulas：Beef extract 3g, yeast extract 1g, peptone 10g, sucrose 10g,
Agar 17-20g, adds distilled water to 1000mL.28 DEG C of culture 3d, obtain test tube species.
Test tube species are inoculated into 250mL triangular flasks (per bottled 150mL) fluid nutrient medium again, culture medium prescription is：With
Weight percent meter, yeast extract 0.1%-0.2%, peptone 0.5%-0.6%, beef extract 0.3%-0.4%, sucrose
1%-2%, remainder are distilled water, and pH 6.8-7.0, fermentation temperature is 25-28 DEG C, shaking speed 150rmin-1, fermentation when
Between 2-3d.The zymotic fluid of formation or the manufactured goods of zymotic fluid can direct or indirect processed soybeans seeds.Processed Soybean Species
Son can mitigate by the harm caused by soybean cyst nematode evil.
Embodiment 2
Bacillus thuringiensis (B.thuringiensise) Sneb1076 fermented products prevent soybean cyst nematode Heterodera glycines field
Effect test
This experiment respectively in Shenyang City, Liaoning Province Agricultural University Of Shenyang experimental plot, Shenyang City, Liaoning Province Kangping city experimental plot and
Heilongjiang Province Daqing academy of agricultural sciences experimental plot carries out field efficacy experiment.The zymotic fluid stoste that will be prepared, using liquid kind 1: 70
Volume ratio carries out seed pelleting processing, dries in the shade, processed soya seeds is normally sowed, soybean plant strain exists under the conditions of cool place
Seedling stage can generate the resistance to soybean cyst nematode Heterodera glycines, and Induced resistant effect is more stable, obtain good control effect.
1. test preparation
Prepare bacillus thuringiensis (B.thuringiensise) Sneb1076's by aforementioned liquids fermentation culture method
Zymotic fluid waits for spare.
2. experiment soybean varieties
Distant beans 15, Liaoning Academy of Agricultural Sciences's soybean research are provided.
3. method for treating seeds
By soya seeds first with 5.0% liquor natrii hypochloritis surface sterilization 5min, then with aseptic water washing 3-5 times.It will system
The zymotic fluid got ready carries out seed pelleting processing using 1: 70 volume ratio of liquid kind, cloudy dry under the conditions of cool place, spare.
4. test method
This experiment is respectively in Shenyang City, Liaoning Province Agricultural University Of Shenyang experimental plot, Shenyang City, Liaoning Province Kangping city and Hei Long
The Daqings Jiang Sheng academy of agricultural sciences experimental plot carries out field efficacy experiment, and experimental plot soybean cyst nematode Heterodera glycines are very serious at three, and main cultivation is made
Object soybean and corn etc..It is control with the soya seeds without the soya seeds of coating and the coating of NA liquid mediums, is planting
45d is investigated afterwards, and soybean plant strain, which is connected root, to be dug out, and keeps root system complete, investigates soybean root system cyst number and root-rot state of an illness grade
Number.
5. test result
The soybean handled using Sneb1076 fermented products, soybean root system cyst number, which is substantially less than, to be compareed, in Shenyang Agriculture
The field efficacy of university, Kangping and grand celebration is respectively 68.1%, 79.4% and 51.5%, the soybean after Cotton seeds with it is untreated
Soybean there are significant differences.
The field efficacy of 1 Sneb1076 fermented product Cotton seeds soya seeds of table
Embodiment 3
Bacillus thuringiensis (B.thuringiensise) Sneb1076 fermented products prevent soybean cyst nematode Heterodera glycines potting
Effect test
Anti- stability is lured in order to verify it, has carried out the control effect research of potting hereby.This experiment is in Shenyang Agriculture
University greenhouse greenhouse carries out pot test effect experiment, with without the Soybean Species of the soya seeds of coating and the coating of NA liquid mediums
Son is control, and experiment is in triplicate.
1. test preparation
Prepare bacillus thuringiensis (B.thuringiensise) Sneb1076's by aforementioned liquids fermentation culture method
Zymotic fluid waits for spare.
2. experiment soybean varieties
Distant beans 15, Liaoning Academy of Agricultural Sciences's soybean research are provided.
3. method for treating seeds
By soya seeds first with 5.0% liquor natrii hypochloritis surface sterilization 5min, then with aseptic water washing 3-5 times.It will system
The zymotic fluid got ready carries out seed pelleting processing using 1: 70 volume ratio of liquid kind, aseptically dries, spare.
4. for trying nematode
The rhizosphere cyst soil for picking up from Shenyang City, Liaoning Province Kangping city experimental plot, is identified as soybean cyst nematode Heterodera glycines (Soybean
Cyst nematode), by the cyst of the elutriation full maturation of crossing sieve method picking, it is put into the sterilizes culture dish for filling sterile water,
Then 3min, and aseptic water washing 3 times are sterilized with 0.5%NaCLO, therefrom the complete and full cyst of picking, merging fills suitable
In the sterilizes culture dish for measuring sterile water, puts it into 28 DEG C of insulating box and cultivate, meet the second instar larvae J2 hatched after 3d
Kind to soybean root easily susceptible in greenhouse expand numerous.The cyst for expanding numerous acquisition is hatched into J2 using tray method, is prepared into
200/mL of nematode suspension.
5. test method
The zymotic fluid stoste of Sneb1076 is subjected to seed pelleting processing using 1: 70 volume ratio of liquid kind, will be dried after coating
Soya seeds be seeded into the seedling alms bowl for educating 12 × 12cm, per basin be inoculated with 2000 soybean cyst nematode Heterodera glycines.With without coating
Soya seeds and the soya seeds of NA liquid mediums coating are control, and 45d is investigated after transplanting, measurement soybean plant height,
Root long, root fresh weight and root interior lines borer population.
6. test result
The soybean handled using Sneb1076 fermented products, root knot number is significantly lower than control, soybean plant height, root long, stalk
With untreated soybean there are notable difference, potting preventive effect is 40.7% for weight, root fresh weight.
The potting preventive effect of 2 Snef1076 fermented product Cotton seeds soya seeds of table
The above is only a preferred embodiment of the present invention, it is noted that for the ordinary skill people of the art
For member, without departing from the principle of the present invention, it can also make several improvements and retouch, these improvements and modifications are also answered
It is considered as protection scope of the present invention.
Claims (5)
1. one kind having the active bacterium bacterial strain of induction of resistance, which is characterized in that the bacterial strain is bacillus thuringiensis
(Bacillus thuringiensis), preserving number are CGMCC No.10831.
2. a kind of cultural method with the active bacterium bacterial strain of induction of resistance as described in claim 1, which is characterized in that packet
Include following steps：
S1, it weighs by mass percentage：Yeast extract 0.1%-0.2%, peptone 0.5%-0.6%, beef extract 0.3%-
0.4%, sucrose 1%-2%, remainder are distilled water, after mixing, obtain fluid nutrient medium；
S2, the fluid nutrient medium obtained by step S1 is sub-packed in the ratio of every bottle of 150mL in the triangular flask of 250mL, has been dispensed
Strain is inoculated into fluid nutrient medium after, pH6.8-7.0, temperature is 25-28 DEG C, shaking speed 150rmin-1, fermentation
2-3d obtains metabolite.
3. a kind of application with the active bacterium bacterial strain of induction of resistance as described in claim 1, which is characterized in that for making
Standby plant nematode biological control induction of resistance preparation.
4. a kind of application with the active bacterium bacterial strain of induction of resistance as claimed in claim 3, which is characterized in that described to lure
Impedance preparation includes the metabolism obtained by a effective amount of above-mentioned bacillus thuringiensis (B.thuringiensise) or step S2
Product.
5. a kind of application with the active bacterium bacterial strain of induction of resistance as claimed in claim 3, which is characterized in that described
Plant nematode is cyst nematode (Soybean cyst nematode).
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Citations (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| EP0303426A2 (en) * | 1987-08-12 | 1989-02-15 | Mycogen Corporation | Novel isolates of bacillus thuringiensis having activity against nematodes |
| AU656726B2 (en) * | 1990-06-11 | 1995-02-16 | Mycogen Corporation | Novel bacillus thuringiensis microbes active against nematodes, and genes encoding novel nematode-active toxins cloned from bacillus thuringiensis isolates |
| CN1849890A (en) * | 2006-04-06 | 2006-10-25 | 湖北省生物农药工程研究中心 | Agritol suspension seedling-coating agent |
| CN102108339A (en) * | 2010-11-26 | 2011-06-29 | 沈阳农业大学 | Bacillus megaterium with capability of inducing stress resistance of soybeans and application thereof |
| CN102925387A (en) * | 2012-10-26 | 2013-02-13 | 沈阳农业大学 | Bacillus simplex for inducing soybean to generate soybean cyst nematode resistance and application |
-
2015
- 2015-09-15 CN CN201510607225.8A patent/CN105112340B/en active Active
Patent Citations (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| EP0303426A2 (en) * | 1987-08-12 | 1989-02-15 | Mycogen Corporation | Novel isolates of bacillus thuringiensis having activity against nematodes |
| AU656726B2 (en) * | 1990-06-11 | 1995-02-16 | Mycogen Corporation | Novel bacillus thuringiensis microbes active against nematodes, and genes encoding novel nematode-active toxins cloned from bacillus thuringiensis isolates |
| CN1849890A (en) * | 2006-04-06 | 2006-10-25 | 湖北省生物农药工程研究中心 | Agritol suspension seedling-coating agent |
| CN102108339A (en) * | 2010-11-26 | 2011-06-29 | 沈阳农业大学 | Bacillus megaterium with capability of inducing stress resistance of soybeans and application thereof |
| CN102925387A (en) * | 2012-10-26 | 2013-02-13 | 沈阳农业大学 | Bacillus simplex for inducing soybean to generate soybean cyst nematode resistance and application |
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