CN110178579A - A kind of Cruciferae clubroot identification method - Google Patents
A kind of Cruciferae clubroot identification method Download PDFInfo
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- 241000219193 Brassicaceae Species 0.000 title claims abstract description 23
- 241001503436 Plasmodiophora brassicae Species 0.000 claims abstract description 85
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- 241000894006 Bacteria Species 0.000 claims abstract description 41
- 201000010099 disease Diseases 0.000 claims abstract description 40
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- 239000000243 solution Substances 0.000 claims abstract description 24
- 235000015097 nutrients Nutrition 0.000 claims abstract description 15
- 238000002347 injection Methods 0.000 claims abstract description 10
- 239000007924 injection Substances 0.000 claims abstract description 10
- 208000035240 Disease Resistance Diseases 0.000 claims abstract description 6
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Classifications
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- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01G—HORTICULTURE; CULTIVATION OF VEGETABLES, FLOWERS, RICE, FRUIT, VINES, HOPS OR SEAWEED; FORESTRY; WATERING
- A01G13/00—Protecting plants
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- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01G—HORTICULTURE; CULTIVATION OF VEGETABLES, FLOWERS, RICE, FRUIT, VINES, HOPS OR SEAWEED; FORESTRY; WATERING
- A01G24/00—Growth substrates; Culture media; Apparatus or methods therefor
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- A01G—HORTICULTURE; CULTIVATION OF VEGETABLES, FLOWERS, RICE, FRUIT, VINES, HOPS OR SEAWEED; FORESTRY; WATERING
- A01G24/00—Growth substrates; Culture media; Apparatus or methods therefor
- A01G24/10—Growth substrates; Culture media; Apparatus or methods therefor based on or containing inorganic material
- A01G24/12—Growth substrates; Culture media; Apparatus or methods therefor based on or containing inorganic material containing soil minerals
- A01G24/15—Calcined rock, e.g. perlite, vermiculite or clay aggregates
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- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01G—HORTICULTURE; CULTIVATION OF VEGETABLES, FLOWERS, RICE, FRUIT, VINES, HOPS OR SEAWEED; FORESTRY; WATERING
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- A01G—HORTICULTURE; CULTIVATION OF VEGETABLES, FLOWERS, RICE, FRUIT, VINES, HOPS OR SEAWEED; FORESTRY; WATERING
- A01G31/00—Soilless cultivation, e.g. hydroponics
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- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01G—HORTICULTURE; CULTIVATION OF VEGETABLES, FLOWERS, RICE, FRUIT, VINES, HOPS OR SEAWEED; FORESTRY; WATERING
- A01G7/00—Botany in general
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- A01G9/02—Receptacles, e.g. flower-pots or boxes; Glasses for cultivating flowers
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Abstract
The invention discloses a kind of Cruciferae clubroot identification methods, belong to clubroot identification method technical field, specifically includes the following steps: Cruciferae plant is carried out nursery using Seedling bag cultural method；Plasmodiophora brassicae bacterium solution is injected to by plant root using injection method after plant grows a piece of true leaf；During plant strain growth, nutrient solution is sprayed to plant；It falls ill to plant root, carries out clubroot Disease Resistance Identification；Identification method of the present invention is cultivated using Seedling bag method, injection method connects bacterium, easy to operate and plant disease incidence, disease index highest, and identification method is accurate.
Description
Technical field
The invention belongs to clubroot identification method technical fields, and in particular to a kind of Cruciferae clubroot identification method.
Background technique
Clubroot is to threaten a kind of worldwide soil-borne disease of crucifer, which is by rape plasmodiophora brassicae
(Plasmodiophora brassicae Woronin) infects Cruciferae root, and parenchyma cell is caused largely to divide increasing
It grows and increases, and then root gradually forms tumour.Clubroot obligatory parasitism Cruciferae, the Cruciferae that China is mainly planted are made
Object is Chinese cabbage, and pakchoi, cabbage, radish, cabbage heart etc., the generation of clubroot causes crop in cruciferae very serious
Harm.
Currently, more and more to the research of clubroot both at home and abroad, either breeding for disease resistance or plasmodiophora brassicae biological strain
Identification requires to do plant the investigation of clubroot character.And all it is both at home and abroad at present traditional substrate culture method that uses, it is this
Method is sampling, in the presence of there are not matrix residual in easy cleaning, root, need the problems such as taking a substantial amount of time when character is investigated.Especially
It is that the sewage for cleaning root in crop field nowhere discharges, and the resting spore time-to-live of plasmodiophora brassicae is up to 20 years again, can make in this way
It is more and more at plasmodiophora brassicae contaminated land area, it is also more serious to the harm of crop in cruciferae.
Bacterium method is effectively connect in clubroot qualification process has the method much reported both at home and abroad at present to have, insertion
(Strelkov et al, 2007), and bacterium local method (fourth cloud is spent, and 2005；Hu Jingfeng etc., 2010), injection method (Donald et
Al, 2009), soak root method (Li Qian etc., 2012), leaching bud method etc. (Cao et al, 2009), these connect bacterium method have it is respective
Up to the present advantage and disadvantage not yet form and unified stable connect bacterium method.
As the onset area of clubroot at home and abroad is increasing, the state of an illness is also increasingly heavier, and the research to plasmodiophora brassicae is
Extremely urgent task, and most of these researchs require to do the investigation of clubroot character.For this purpose, it is fast to establish a clubroot
Fast, accurate identification method is very important, and can be mitigated pollution caused by test significantly in this way, be reduced the wave of manpower and material resources
Take, so that the research for plasmodiophora brassicae mitigates more work loads.
Summary of the invention
The present invention provides a kind of Cruciferae clubroot identification methods, solve conventional method not easy cleaning, root has
Matrix residual, time-consuming and the problem of result accuracy difference, the present invention are achieved by the following technical solution.
It is an object of the present invention to provide a kind of Cruciferae clubroot identification methods, comprising the following steps:
Cruciferae plant is subjected to nursery using Seedling bag cultural method；Using injection after plant grows a piece of true leaf
Plasmodiophora brassicae bacterium solution is injected to plant root by method；During plant strain growth, nutrient solution is sprayed to plant；It falls ill to plant root, into
Row clubroot Disease Resistance Identification；
The Seedling bag cultural method is Cruciferae plant to be used haydite, sand and perlite mixed-matrix, and match
To spray nutrient solution, nursery is carried out.
Preferably, the Seedling bag cultural method specifically includes the following steps:
The sterilized matrix A being made of the mixing of peat, vermiculite and perlite is attached in Seedling bag, matrix is sprayed water slow
Seed is multicast in matrix by slow wetting after water supply in media absorption, after germination, plant field planting into flowerpot, and flowerpot
It is provided with haydite, sand and perlite 0.9~1.1:0.9 in mass ratio~1.1:1 that sterilized partial size is respectively 8~15mm
The matrix B of composition sprays nutrient solution during plant strain growth, withered to prevent seedling.
Preferably, the injection method specifically: bacterium solution is connect into bacterium to plant root with pipettor, every plant of plasmodiophora brassicae suspends
It is 1 × 10 that liquid, which connects bacterium amount,5~1 × 108A spore.
It is highly preferred that the knee bacterium suspension is made by following methods:
It takes morbidity root nodule clear water to clean, is activated at room temperature；Sterile water is added in activated morbidity root nodule, institute
State morbidity root nodule: sterile water amount ratio is 2~4g:10mL, and is crushed 2~3 times with broken wall mill bacterium machine, and filtering, merging filtrate fills
Divide and shakes up；Plasmodiophora brassicae spore concentration in rear filtrate is shaken up with blood cell counting plate detection, and according to the concentration detected with sterile
Water is diluted to 1 × 105~1 × 108Knee bacterium suspension is made in a spore/mL.
Preferably, the nutrient solution is A Fudaoning mill water culture nutrient solution.
Preferably, the clubroot Disease Resistance Identification comprising the following specific steps
Disease index is calculated, clubroot identification is carried out according to disease index calculated value；Wherein, the disease index calculating side
Shown in method such as formula (I):
Disease index=∑ [diseased plant numbers at different levels × relative disease series] × 100/ (investigation total strain number × 3) (I)
It is as follows that clubroot standard of perfection is carried out according to disease index calculated value: when disease index is less than or equal to 25, being determined
It is disease-resistant；Disease index is susceptible when being greater than 25；
The relative disease series are as follows:
0 grade: root system normal growth is not fallen ill；1 grade: lateral root morbidity has tubercle, and main root is not fallen ill；2 grades: main root has tubercle,
Or lateral root has big tumor or main root and lateral root to have big tumor；3 grades: main root, lateral root have big tumor.
Compared with prior art, the present invention has the following advantages:
(1) root is easy to clean, quick；The present invention cultivation medium kind easy to clean using haydite, sand, perlite
It plants, provides source of nutrition with A Fudaoning nutrient solution for plant, the cultivation medium around such root is in loose condition (of surface), in root
When portion's sampling or the investigation of clubroot character need to wash root, root can be cleaned up quickly, and not need too many water；
(2) root damage is reduced, error is reduced；Cultivation medium that the present invention uses and matrix have a very big difference, perlite,
Sand, haydite will not closely be attached to plant root as the substrate culture method that traditional identification method uses, substrate block,
A large amount of fibrous root can be torn off when cleaning, cause test result inaccurate；Identification method of the present invention is easy to clean, and speed is fast, keeps away
Tubercle on fibrous root is exempted to wash off in the process of cleaning, has also greatly reduced root in the damage of cleaning process, improve the standard of test
Exactness；
(3) pollution to soil is reduced；The present invention reduces the use to matrix, reduces water consumption when washing root, makes to take
The discharge amount of sewage and matrix with plasmodiophora brassicae is reduced, and can be greatly reduced the pollution to soil and water resource in this way, be made knee
The widened speed of bacterium contaminated area reduces；
(4) waste of manpower and material resources is reduced；The matrix price that traditional cultivation method uses is high and cannot reuse, and makes pottery
Grain, perlite, sand cultivation medium can use the sultry sterilizing methods of high temperature after use, these can be reused, in this way
The waste of resource is greatly reduced, and in the investigation of large batch of clubroot character, biggish root nodule completely dispenses with cleaning, directly
The sand, haydite, perlite of root, which are gently shaked off, can be clearly distinguished morbidity grade, only do not fall ill with it is small on fibrous root
Tumor needs simply to be cleaned with water, does not need that tune can be greatly improved in this way as cleaning three or four times when conventional method is investigated
Speed is looked into, mitigates workload when investigation, to save manpower；
(5) identification method of the present invention connects bacterium, easy to operate and plant disease incidence, disease using the cultivation of Seedling bag method, injection method
Feelings index highest, identification method are accurate.
Specific embodiment
In order to enable those skilled in the art to more fully understand, technical solution of the present invention is practiced, below with reference to specific
The invention will be further described for embodiment and data, but illustrated embodiment is not as a limitation of the invention.
Experimental method and detection method described in following each embodiments are unless otherwise specified conventional method；The examination
Agent and material can be commercially available on the market unless otherwise specified.
In order to absolutely prove clubroot identification method of the present invention, we have chosen the plasmodiophora brassicae material of domestic different regions,
And the plasmodiophora brassicae of each material is inoculated on four different hosts, plant carries out the disease-resistant sex investigation of clubroot after connecing bacterium 6 weeks.
The plasmodiophora brassicae material that the present invention uses collected from domestic each department and expand it is numerous obtain, and 12 kinds of plasmodiophora brassicaes have been
The identification that biological strain is completed by conventional method chooses 12 kinds of plasmodiophora brassicaes, using identification method of the present invention, by comparing
The qualification result of the two verifies the accuracy of identification method of the present invention, and 12 kinds of plasmodiophora brassicae materials are specific, and see Table 1 for details:
1, table test uses plasmodiophora brassicae source
12 kinds of plasmodiophora brassicae materials in table 1 are prepared into knee bacteria suspension, specific preparation method is as follows.
Prepare embodiment 1
(1) it takes the root nodule material 30g that respectively falls ill in table 1 to be cleaned with clear water, activates at room temperature for 24 hours；
(2) by activated root nodule, 100ml sterile water is added, broken wall grinds bacterium machine 10s and crushes 2 times, and 8 layers of 1 stratus of gauze are female
Cloth filtering；
(3) filtrate will put together, sufficiently shake up twice, with blood cell counting plate detection filtrate in plasmodiophora brassicae filtrate it is dense
Degree, and 1 × 10 is diluted to sterile water according to the concentration of the plasmodiophora brassicae filtrate detected7A spore mL-1, plasmodiophora brassicae is made and suspends
Liquid.
Prepare embodiment 2
(1) it takes root nodule material 20 g clear water of respectively falling ill in table 1 to clean, activates at room temperature for 24 hours；
(2) by activated root nodule, 100ml sterile water is added, broken wall grinds bacterium machine 10s and crushes 2 times, and 8 layers of 1 stratus of gauze are female
Cloth filtering；
(3) filtrate will put together, sufficiently shake up twice, with blood cell counting plate detection filtrate in plasmodiophora brassicae filtrate it is dense
Degree, and 1 × 10 is diluted to sterile water according to the concentration of the plasmodiophora brassicae filtrate detected5A spore mL-1, plasmodiophora brassicae is made and suspends
Liquid.
Prepare embodiment 3
(1) it takes the root nodule material 40g that respectively falls ill in table 1 to be cleaned with clear water, activates at room temperature for 24 hours；
(2) by activated root nodule, 100ml sterile water is added, broken wall grinds bacterium machine 15s and crushes 3 times, and 8 layers of 1 stratus of gauze are female
Cloth filtering；
(3) it filtrate will put together, sufficiently shake up twice, the concentration of plasmodiophora brassicae in filtrate is detected with blood cell counting plate,
And 1 × 10 is diluted to sterile water according to the concentration of the plasmodiophora brassicae filtrate detected8A spore mL-1, knee bacterium suspension is made.
The above-mentioned measuring method with knee bacteria concentration in blood cell counting plate detection filtrate:
Above-mentioned each group plasmodiophora brassicae filtrate is sufficiently shaken up, takes 10 μ l plasmodiophora brassicae filtrates that the aniline blue solution of 100 μ l is added, adds
Distilled water mixes, 5min to be dyed to 1ml, takes 10 μ l to mix liquid and be added to covered on blood counting chamber, in optical microscopy
The number of lower observation spore, is computed the concentration for obtaining bacterium solution.
The clubroot method of resistance identification that the present invention uses are as follows: disease index is to divide Chinese cabbage cultivar to a certain knee
The whether disease-resistant standard of bacterium, disease index be less than or equal to 25 when, be determined as it is disease-resistant, disease index be greater than 25 when be susceptible, institute
It states shown in disease index calculation method such as following formula (I):
Disease index=∑ [diseased plant numbers at different levels × relative disease series] × 100/ (investigation total strain number × 3)
(Ⅰ)
The relative disease series are as follows:
0 grade: root system normal growth is not fallen ill；
1 grade: lateral root morbidity has tubercle, and main root is not fallen ill；
2 grades: main root has tubercle or lateral root to have big tumor or main root and lateral root to have big tumor；
3 grades: main root, lateral root have big tumor.
Four kinds of hosts are selected, are inoculated with above-mentioned 12 kinds of rhizobium respectively, specific embodiment is as follows.
Embodiment 1
Cruciferae clubroot identification method, comprising the following steps:
(1) will in Seedling bag be packed into matrix (matrix, that is, market sale horticultural gardening matrix, ingredient be peat, vermiculite and
Perlite is mixed to prepare with arbitrary proportion), moisture is sprayed with watering can, matrix is soaked, by host cabbage (Badger
Shipper it) is seeded into Seedling bag for 18 plants, setting Chinese cabbage susceptible variety 91-12 is control, and the quantity of sowing is also 18 plants；
(2) be transplanted in flowerpot after Badger Shipper germination, flowerpot be provided with sterilized partial size be respectively 8~
The homomixture of haydite, sand and perlite the 1:1:1 in mass ratio composition of 15mm, growth period spray A Fudaoning water to plant
Cultivation nutrient fluid, to prevent withered；Transplanting 1 week (plant grows a piece of true leaf) carries out connecing bacterium using injection method afterwards, and classification in table 1 is compiled
It number being seeded in Badger Shipper and control group for 1 plasmodiophora brassicae, detection plasmodiophora brassicae spore concentration compound concentration is 1 ×
107Bacterium solution is connect bacterium to plant root by the spore suspension of a spore/ml, pipettor, and every plant of knee bacterium suspension connects bacterium amount
For 1ml, pay attention to before and after connecing bacterium in for 24 hours should not nutrient solution, be diluted to prevent bacterium solution, reduce plant disease incidence；
(3) the disease-resistant sex investigation of clubroot is carried out after plant connects bacterium 6 weeks, the plant root infected at this time has been fallen ill,
Root nodule is formed, plant root is gently extracted, the sand perlite etc. of root attachment is gently shaked off, is slowly cleaned in clear water
Root influences grade classification of falling ill in order to avoid making root nodule is impaired to fall.
Embodiment 2
Step with embodiment 1, the difference is that, plasmodiophora brassicae spore concentration be 1 × 105A spore/ml, haydite, sand
It is 0.9:0.9:1, the plasmodiophora brassicae that inoculation class number is 2 with perlite mass ratio.
Embodiment 3
Step with embodiment 1, the difference is that, plasmodiophora brassicae spore concentration be 1 × 108A spore/ml, haydite, sand
It is 1.1:1.1:1, the plasmodiophora brassicae that inoculation class number is 3 with perlite mass ratio.
Embodiment 4
Step with embodiment 1, the difference is that, inoculation class number be 4 plasmodiophora brassicae.
Embodiment 5
Step with embodiment 1, the difference is that, inoculation class number be 5 plasmodiophora brassicae.
Embodiment 6
Step with embodiment 1, the difference is that, inoculation class number be 6 plasmodiophora brassicae.
Embodiment 7
Step with embodiment 1, the difference is that, inoculation class number be 7 plasmodiophora brassicae.
Embodiment 8
Step with embodiment 1, the difference is that, inoculation class number be 8 plasmodiophora brassicae.
Embodiment 9
Step with embodiment 1, the difference is that, inoculation class number be 9 plasmodiophora brassicae.
Embodiment 10
Step with embodiment 1, the difference is that, inoculation class number be 10 plasmodiophora brassicae.
Embodiment 11
Step with embodiment 1, the difference is that, inoculation class number be 11 plasmodiophora brassicae.
Embodiment 12
Step with embodiment 1, the difference is that, inoculation class number be 12 plasmodiophora brassicae.
Embodiment 13
Step with embodiment 1, the difference is that, host be cabbage (Jersey Queen).
Embodiment 14
Step with embodiment 13, the difference is that, inoculation class number be 2 plasmodiophora brassicae.
Embodiment 15
Step with embodiment 13, the difference is that, inoculation class number be 3 plasmodiophora brassicae.
Embodiment 16
Step with embodiment 13, the difference is that, inoculation class number be 4 plasmodiophora brassicae.
Embodiment 17
Step with embodiment 13, the difference is that, inoculation class number be 5 plasmodiophora brassicae.
Embodiment 18
Step with embodiment 13, the difference is that, inoculation class number be 6 plasmodiophora brassicae.
Embodiment 19
Step with embodiment 13, the difference is that, inoculation class number be 7 plasmodiophora brassicae.
Embodiment 20
Step with embodiment 13, the difference is that, inoculation class number be 8 plasmodiophora brassicae.
Embodiment 21
Step with embodiment 13, the difference is that, inoculation class number be 9 plasmodiophora brassicae.
Embodiment 22
Step with embodiment 13, the difference is that, inoculation class number be 10 plasmodiophora brassicae.
Embodiment 23
Step with embodiment 13, the difference is that, inoculation class number be 11 plasmodiophora brassicae.
Embodiment 24
Step with embodiment 13, the difference is that, inoculation class number be 12 plasmodiophora brassicae.
Embodiment 25
Step with embodiment 1, the difference is that, host be turnip (Wilhelmsburger).
Embodiment 26
Step with embodiment 25, the difference is that, inoculation class number be 2 plasmodiophora brassicae.
Embodiment 27
Step with embodiment 25, the difference is that, inoculation class number be 3 plasmodiophora brassicae.
Embodiment 28
Step with embodiment 25, the difference is that, inoculation class number be 4 plasmodiophora brassicae.
Embodiment 29
Step with embodiment 25, the difference is that, inoculation class number be 5 plasmodiophora brassicae.
Embodiment 30
Step with embodiment 25, the difference is that, inoculation class number be 6 plasmodiophora brassicae.
Embodiment 31
Step with embodiment 25, the difference is that, inoculation class number be 7 plasmodiophora brassicae.
Embodiment 32
Step with embodiment 25, the difference is that, inoculation class number be 8 plasmodiophora brassicae.
Embodiment 33
Step with embodiment 25, the difference is that, inoculation class number be 9 plasmodiophora brassicae.
Embodiment 34
Step with embodiment 25, the difference is that, inoculation class number be 10 plasmodiophora brassicae.
Embodiment 35
Step with embodiment 25, the difference is that, inoculation class number be 11 plasmodiophora brassicae.
Embodiment 36
Step with embodiment 25, the difference is that, inoculation class number be 12 plasmodiophora brassicae.
Embodiment 37
Step with embodiment 1, the difference is that, host be turnip (Laurentian).
Embodiment 38
Step with embodiment 37, the difference is that, inoculation class number be 2 plasmodiophora brassicae.
Embodiment 39
Step with embodiment 37, the difference is that, inoculation class number be 3 plasmodiophora brassicae.
Embodiment 40
Step with embodiment 37, the difference is that, inoculation class number be 4 plasmodiophora brassicae.
Embodiment 41
Step with embodiment 37, the difference is that, inoculation class number be 5 plasmodiophora brassicae.
Embodiment 42
Step with embodiment 37, the difference is that, inoculation class number be 6 plasmodiophora brassicae.
Embodiment 43
Step with embodiment 37, the difference is that, inoculation class number be 7 plasmodiophora brassicae.
Embodiment 44
Step with embodiment 37, the difference is that, inoculation class number be 8 plasmodiophora brassicae.
Embodiment 45
Step with embodiment 37, the difference is that, inoculation class number be 9 plasmodiophora brassicae.
Embodiment 46
Step with embodiment 37, the difference is that, inoculation class number be 10 plasmodiophora brassicae.
Embodiment 47
Step with embodiment 37, the difference is that, inoculation class number be 11 plasmodiophora brassicae.
Embodiment 48
Step with embodiment 37, the difference is that, inoculation class number be 12 plasmodiophora brassicae.
1~embodiment of embodiment 48 be respectively four hosts (Badger Shipper, Jersey Queen,
Wilhelmsburger and Laurentian) it is inoculated with 12 kinds of plasmodiophora brassicaes in table 1 respectively, it is grown 6 weeks after plant connects bacterium, to plant
Strain carries out clubroot Characters Identification, and plant root is cleaned up, and by root nodule size divided rank, specific incidence is shown in Table 2,
Biological strain division is carried out by Williams biological strain identification system, by this qualification result and forefathers' qualification result (i.e. table 1
In the identification of biological strain has been completed by conventional method) comparison, comparing result is shown in Table 3.
2 1~embodiment of embodiment 48 of table is inoculated with host's morbidity result after 12 parts of plasmodiophora brassicaes
3 above-mentioned 12 parts of plasmodiophora brassicae Race Identification results of table
Note :+indicate that host is susceptible ,-indicate that host is disease-resistant.
By table 2 and 3 result of table it is found that identification method qualification result of the present invention identifies that qualification result is consistent with forefathers, it was demonstrated that
The accuracy of identification method of the present invention, in addition, the present invention is using cultivation mediums easy to clean such as haydite, sand, perlites
Plantation, provides source of nutrition with A Fudaoning nutrient solution for plant, and the cultivation medium around such root is in loose condition (of surface),
When root sampling or clubroot Characters Identification need to wash root, root can be cleaned up quickly, and not need too many water；
The cultivation medium and matrix that the present invention uses have very big difference, and perlite, sand, haydite will not be used as traditional identification method
Substrate culture method, substrate block is closely attached to plant root, can tear off a large amount of fibrous root in cleaning, test is caused to tie
Fruit inaccuracy；Identification method of the present invention is easy to clean, and speed is fast, avoids tubercle on fibrous root and washes off in the process of cleaning,
Root is greatly reduced in the damage of cleaning process, improves the accuracy of test.
The present invention reduces the use to matrix, water consumption when washing root is reduced, makes the sewage and base that carry plasmodiophora brassicae
The discharge amount of matter is reduced, and can be greatly reduced the pollution to soil and water resource in this way, be made the widened of plasmodiophora brassicae contaminated area
Speed reduces；The matrix price that traditional cultivation method uses is high and cannot reuse, and haydite, perlite, the cultivation such as sand
Medium can use the sultry sterilizing methods of high temperature after use, these can be reused, and greatly reduce the wave of resource in this way
Take, and in large batch of clubroot Characters Identification, biggish root nodule completely dispenses with cleaning, directly by the sand of root, haydite,
Perlite, which is gently shaked off, can be clearly distinguished morbidity grade, and only not falling ill, it is simply clear with water to need with the tubercle on fibrous root
It washes, does not need to greatly improve identification speed in this way as original mirror fixed time cleaning three or four times, mitigate work when identification
Amount, to save manpower.
In conclusion identification method of the present invention connects bacterium using the cultivation of Seedling bag method, injection method, easy to operate and plant falls ill
Rate, disease index highest, identification method are accurate.
Obviously, various changes and modifications can be made to the invention without departing from essence of the invention by those skilled in the art
Mind and range.In this way, if these modifications and changes of the present invention belongs to the range of the claims in the present invention and its equivalent technologies
Within be also intended to include these modifications and variations.
Claims (6)
1. a kind of Cruciferae clubroot identification method, which comprises the following steps:
Cruciferae plant is subjected to nursery using Seedling bag cultural method；Use injection method will after plant grows a piece of true leaf
Plasmodiophora brassicae bacterium solution is injected to plant root；During plant strain growth, nutrient solution is sprayed to plant；It falls ill to plant root, carries out root
Swollen disease Disease Resistance Identification；
The Seedling bag cultural method is Cruciferae plant to be used haydite, sand and perlite mixed-matrix, and be equipped with spray
Nutrient solution is spilt, nursery is carried out.
2. Cruciferae clubroot identification method according to claim 1, which is characterized in that the Seedling bag cultural method
Specifically includes the following steps:
The sterilized matrix A being made of the mixing of peat, vermiculite and perlite is attached in Seedling bag, water spray wetting, to matrix
Seed is multicast in matrix after moisture absorption, after germination, plant field planting into flowerpot, flowerpot is provided with sterilized
Partial size is respectively that the haydite, sand and perlite of 8~15mm is 0.9~1.1:0.9~1.1:1 composition matrix B in mass ratio,
Nutrient solution is sprayed during plant strain growth, it is withered to prevent seedling.
3. Cruciferae clubroot identification method according to claim 1, which is characterized in that the injection method specifically:
Knee bacterium suspension is connect into bacterium to plant root, it is 1 × 10 that every plant of knee bacterium suspension, which connects bacterium amount,5~1 × 108A spore.
4. Cruciferae clubroot identification method according to claim 3, which is characterized in that the knee bacterium suspension by
Following methods are made:
It takes morbidity root nodule clear water to clean, is activated at room temperature；Sterile water, the hair is added in activated morbidity root nodule
Old complaint tumor: sterile water amount ratio is 2~4g:10mL, and is crushed 2~3 times with broken wall mill bacterium machine, and filtering, merging filtrate sufficiently shakes
It is even；Detection shakes up plasmodiophora brassicae spore concentration in rear filtrate, and is diluted to 1 × 10 with sterile water according to the concentration detected5~1 ×
108Knee bacterium suspension is made in a spore/mL.
5. Cruciferae clubroot identification method according to claim 1, which is characterized in that the nutrient solution is A Fudao
Peaceful mill water culture nutrient solution.
6. Cruciferae clubroot identification method according to claim 1, which is characterized in that the clubroot disease resistance mirror
It is fixed comprising the following specific steps
Disease index is calculated, clubroot identification is carried out according to disease index calculated value；Wherein, the disease index calculation method is such as
Shown in formula (I):
Disease index=∑ [diseased plant numbers at different levels × relative disease series] × 100/ (investigation total strain number × 3) (I)
It is as follows that clubroot standard of perfection is carried out according to disease index calculated value: when disease index is less than or equal to 25, being judged to resisting
Disease；Disease index is susceptible when being greater than 25；
The relative disease series are as follows:
0 grade: root system normal growth is not fallen ill；1 grade: lateral root morbidity has tubercle, and main root is not fallen ill；2 grades: main root has tubercle or side
Root has big tumor or main root and lateral root to have big tumor；3 grades: main root, lateral root have big tumor.
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