CN103782816A - Quick method for artificial attack of tobacco damping-off - Google Patents
Quick method for artificial attack of tobacco damping-off Download PDFInfo
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- CN103782816A CN103782816A CN201410006422.XA CN201410006422A CN103782816A CN 103782816 A CN103782816 A CN 103782816A CN 201410006422 A CN201410006422 A CN 201410006422A CN 103782816 A CN103782816 A CN 103782816A
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- 235000002637 Nicotiana tabacum Nutrition 0.000 title claims abstract description 37
- 241000208125 Nicotiana Species 0.000 title claims abstract description 36
- 238000000034 method Methods 0.000 title claims abstract description 23
- 241000918585 Pythium aphanidermatum Species 0.000 title claims abstract description 12
- 241000813090 Rhizoctonia solani Species 0.000 claims abstract description 17
- 238000011081 inoculation Methods 0.000 claims description 31
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 19
- 235000019504 cigarettes Nutrition 0.000 claims description 11
- 230000017074 necrotic cell death Effects 0.000 claims description 11
- 239000002689 soil Substances 0.000 claims description 10
- 229920001817 Agar Polymers 0.000 claims description 6
- VVQNEPGJFQJSBK-UHFFFAOYSA-N Methyl methacrylate Chemical compound COC(=O)C(C)=C VVQNEPGJFQJSBK-UHFFFAOYSA-N 0.000 claims description 6
- 229920005372 Plexiglas® Polymers 0.000 claims description 6
- 239000008272 agar Substances 0.000 claims description 6
- 230000001580 bacterial effect Effects 0.000 claims description 6
- 239000002054 inoculum Substances 0.000 claims description 5
- 208000024891 symptom Diseases 0.000 claims description 5
- -1 18 grams Substances 0.000 claims description 4
- 238000007796 conventional method Methods 0.000 claims description 4
- 241001582888 Lobus Species 0.000 claims description 3
- 244000061456 Solanum tuberosum Species 0.000 claims description 3
- 235000002595 Solanum tuberosum Nutrition 0.000 claims description 3
- CZMRCDWAGMRECN-UGDNZRGBSA-N Sucrose Chemical compound O[C@H]1[C@H](O)[C@@H](CO)O[C@@]1(CO)O[C@@H]1[C@H](O)[C@@H](O)[C@H](O)[C@@H](CO)O1 CZMRCDWAGMRECN-UGDNZRGBSA-N 0.000 claims description 3
- 229930006000 Sucrose Natural products 0.000 claims description 3
- 230000035558 fertility Effects 0.000 claims description 3
- 239000001963 growth medium Substances 0.000 claims description 3
- 239000012533 medium component Substances 0.000 claims description 3
- 238000002360 preparation method Methods 0.000 claims description 3
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- 241000233866 Fungi Species 0.000 claims description 2
- 239000003643 water by type Substances 0.000 claims description 2
- 238000005516 engineering process Methods 0.000 abstract description 4
- 230000008901 benefit Effects 0.000 abstract description 3
- 201000010099 disease Diseases 0.000 description 8
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 description 8
- 238000002255 vaccination Methods 0.000 description 5
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- 230000001717 pathogenic effect Effects 0.000 description 3
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- 206010039509 Scab Diseases 0.000 description 2
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- 230000001338 necrotic effect Effects 0.000 description 2
- 230000000474 nursing effect Effects 0.000 description 2
- 238000011160 research Methods 0.000 description 2
- 241000196324 Embryophyta Species 0.000 description 1
- 244000061176 Nicotiana tabacum Species 0.000 description 1
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Abstract
The invention discloses a quick method for an artificial attack of tobacco damping-off. The method includes the following steps: 1, preparing for tobacco seedling cultivation; 2, preparing inoculating mycelia; 3, handling before tobacco seedling inoculating; 4, inoculating rhizoctonia solani; 5, handling after inoculating. The quick method has the advantages that an inoculating technology system is miniaturized, micro seedlings are used, inoculating attack is realized in small space, and short experiment cycle is consumed; the implementation process is simple, and operation is convenient and high in efficiency; the whole process is completed in a common lab, local conditions of the attack are easy to control, interference is small, and technical method standardization is easy to realize.
Description
Technical field
The present invention relates to agrotechnique and biotechnology.A specifically quick method that grows tobacco damping off artificial onset.
Background technology
Infecting by disease fungus Rhizoctonia solani the tobacco damping off causing is one of important disease of tobacco production, mainly fall ill in seedling stage, cause dead seedling, when serious, cause some fields to be short of seedling in a large number, in recent years in the tobacco strain phase also common generation, particularly some high-yield culturing districts, often there is the higher incidence of disease, owing to mainly causing the basal part of stem necrosis of tobacco plant in this disease of strain phase, thereby cause whole strain to be injured, cause larger loss, this has become to produce the major issue facing, address this problem, still depend on the basic research of strengthening tobacco damping off, and artificial onset is this disease about one of basic means of research.To a little less than the Study of Thin of this disease, also do not form so far standard and ripe artificial onset's method due to for a long time.Although this disease main harm cigarette strain basal part of stem, but inventor finds, rhizoctonia solani also can infect tobacco upper blade and cause downright bad scab clearly, this pathogenic property can be used in artificial infection morbidity, and do not find so far tobacco rhizoctonia solani can infect the pathogenic advantageous feature of blade, in artificial onset's practice, be applied and set up supporting technology.
Summary of the invention
The object of this invention is to provide a quick method that grows tobacco damping off artificial onset, complete in a short time artificial onset's work.
The technical scheme that the present invention solves the problems of the technologies described above is as follows:
One quick method that grows tobacco damping off artificial onset is the characteristic of utilizing rhizoctonia solani can infect tobacco leaf to cause necrosis, inoculates immature cigarette seedling in culture dish, causes cigarette seedling leaf to be fallen ill fast, shows clear symptom.The step of artificial onset's method is as follows:
1. tobacco seedling is cultivated and is prepared:
With the common better and loose soil of tray splendid attire fertility, at indoor culture cigarette seedling, grow to after 3～4 vanelets for subsequent use by conventional method.
2. the mycelial preparation of inoculation:
Rhizoctonia solani is transplanted to the dull and stereotyped center of the common fungi culture medium of PSA, PSA medium component is: 200 grams of potatos, 20 grams of sucrose, 18 grams, agar, water 1000mL, under 28 ℃ of conditions, be cultured to and grow compared with macrocolony, cut-off footpath is that 6mm card punch is beaten and got mycelia agar nahlock of the same size in flat-plate bacterial colony periphery, makes inoculum with this mycelia piece.
3. before tobacco seedling inoculation, dispose:
Get the applicable seedling that step 1 cultivates, the moistening softening soil of growing seedlings that first waters before inoculation, so that lift seedlings.Take out clean culture dish and be placed in common shallow square plate, and add thin layer clear water in culture dish; Choose the seedling that has 3～4 vanelets, gently pull up and forward in the clear water of other container splendid attire with have gentle hands, rock gently the soil in wash-out seedlings root, seedling is forwarded in the above-mentioned culture dish of getting ready, leaflet horse is put on the ware edge of culture dish, on root system leans against at the bottom of ware, make seedling be upright state, and make clear water submergence root system.
4. inoculation rhizoctonia solani:
The mycelia piece that picking step 2 is prepared, is placed with gently in step 3 and disposes on the lobus cardiacus of complete seedling or on heart bud.
5. after inoculation, dispose:
Step 4 operate complete after, the culture dish of splendid attire being inoculated to seedling is placed in shallow square plate, and plexiglass tent is covered on shallow square plate, toward injecting thin layer clear water in shallow square plate, makes plexiglass tent enclose inside and forms stable high wet condition in cover; Then package unit is put in to the general room cultivation that possesses daily scattered light, cultivation temperature maintains 25～30 ℃.Note avoiding direct sunlight according to shining.
Inoculation result causes seedling leaves to occur water stain shape necrosis, and last whole seedling total necrosis is rotted.Under normal circumstances, visible obviously necrosis afterwards shows to inoculate 1 day, and after 3～4 days, complete stool is rotted.
Advantage of the present invention is:
1) inoculation technique system microminiaturization, is mainly manifested in small seedling, and in very little space, inoculation morbidity, consumes the very short test period.
2) implementation process is simple, and easy to operate efficiency is high.
3) omnidistancely in common lab, complete, morbidity local condition easily controls, and is disturbed littlely, easily realizes technical method standardization.
Accompanying drawing explanation
Fig. 1 is the tobacco seedling schematic diagram that the present invention cultivates in common lab.
In figure, A cultivates with common enamel square tray; B cultivates with culture dish.
Fig. 2 is of the present invention the schematic diagram after seedling and the inoculation mycelia piece thereof being placed on culture dish ware edge.
Fig. 3 is that postvaccinal seedling is covered on the schematic diagram in shallow square plate by the present invention's plexiglass tent.
Fig. 4 is that the present invention inoculates the morbidity performance after 2 days.
In figure, A～E is the performance of different seedling inoculation germs; F portion is blank inoculation contrast.
Fig. 5 is that the present invention inoculates the morbidity performance after 3 days.
In figure, A～E is the morbidity performance of inoculation germ, and wherein A illustrates the state of an illness of many strains seedling, the performance of B～E signal individual plant seedling; F is blank inoculation contrast.
Embodiment
Below in conjunction with accompanying drawing and embodiment, the invention will be further described.
Cigarette seedling of the present invention is cultivated and inoculation morbidity whole process is all implemented in common lab, and it is to the south that laboratory only need to possess window, and printing opacity is good, and is equipped with common air-conditioning at once.At the other cigarette seedling of cultivating of indoor nearly window, it is mainly to adopt larger seedling-cultivating tray unification to grow seedlings in a large number that the present invention designs according to a conventional method, and as shown in the A of Fig. 1, this nursing that can make to grow seedlings (being mainly the nursing of watering) is relatively simple; After seedling size is applicable to using, seedling is passed to enforcement inoculation operation on other utensil; Adopting this mode to grow seedlings and also have benefit, is exactly that the general tobacco seedling growth of cultivating is normally big or small uneven, transplants seedlings to inoculate elsewhere operation and can select seedling of the same size to test; In addition, the operation of transplanting seedlings can obtain the seedling without earthy state, and the test that need to get rid of soil interference for some is favourable.Practical application also can adopt the utensils such as culture dish to grow seedlings, as shown in the B of Fig. 1;
What inoculation adopted is the small cigarette seedling that grows to 3～4 vanelets, as shown in the B portion of Fig. 1, conventionally only need to cultivate 3～4 weeks.Inoculation operation is to implement in common culture dish, the vanelets center of the verticillate shape of seedling inoculation germ, as shown in Figure 2.
The key condition that tobacco rhizoctonia solani infects tobacco morbidity is to need environment in high wet condition, and the present invention adopts self-control cloche to cover the mode of seedling, as shown in Figure 3, provides stable super-humid conditions, is also convenient to pathogenic process simultaneously and observes.
Morbidity starts in vaccination conventionally, shows as water stain shape necrosis, and as shown in Figure 4, with spreading of mycelia, necrotic plaque is expansion towards periphery fast, and last whole seedling is downright bad rotting all, as shown in Figure 5.Under normal circumstances, inoculate 1 day afterwards visible necrotic plaque occur, after 3～4 days, complete stool is downright bad rots.
Inoculation causes speed or the weight of PD, relevant with rhizoctonia solani inoculation mycelium cell age state, also relevant with postvaccinal cultivation temperature, state of an illness weight can be weighed by downright bad forefront point and the index such as distance or downright bad lesion area or the strain incidence of disease of vaccination.
Embodiment 1
One quick method that grows tobacco damping off artificial onset is inoculated tobacco rhizoctonia solani bacterial strain Rs-1 on the small seedling of tobacco bred cloud and mist 87, as follows operation:
1. tobacco seedling is cultivated and is prepared: with the better and loose soil of splendid attire fertility in common enamel square tray, use conventional method is at indoor culture cigarette seedling, grows to after 3～4 vanelets as shown in the B of Fig. 1 for subsequent use.
2. the mycelial preparation of inoculation: rhizoctonia solani is transplanted to PSA culture medium flat plate center, PSA medium component is: 200 grams of potatos, 20 grams of sucrose, 18 grams, agar, water 1000mL, at 28 ℃, cultivate and grow compared with macrocolony for 48 hours, cut-off footpath is that 6mm card punch is beaten and got mycelia agar nahlock of the same size in flat-plate bacterial colony periphery, makes inoculum with this mycelia piece.
3. before tobacco seedling inoculation, dispose: get the applicable seedling that step 1 cultivates, inoculate first 0.5～1 hour and first water the moistening soil of growing seedlings that softens, so that lift seedlings.Take out clean 9cm culture dish and be placed in common shallow square plate, and add 20ml clear water in culture dish; Choose the seedling that has 3～4 vanelets, gently pull up and forward in the clear water of other container splendid attire with have gentle hands, rock gently the soil in wash-out seedlings root, seedling is forwarded in the above-mentioned 9cm culture dish of getting ready, leaflet horse is put on the ware edge of culture dish, on root system leans against at the bottom of ware, make seedling be upright state, and clear water submergence root system in ware.
4. inoculation rhizoctonia solani: the mycelia piece that picking step 2 is prepared, be placed with gently in step 3 and dispose on the lobus cardiacus of complete seedling or on heart bud, complete after as shown in Figure 2.
5. after inoculation, dispose: step 4 operate complete after, take from the culture dish that splendid attire inoculated seedling by plexiglass tent processed and cover on shallow square plate, in shallow square plate, inject thin layer clear water, make cloche enclose inside and form the interior stable high wet condition of cover, as shown in Figure 3; Then package unit is put in to the general room cultivation that possesses daily scattered light, cultivation temperature maintains 25～27 ℃.Note avoiding direct sunlight according to shining.
Bearing performance goes out the downright bad symptom of blade clearly as the same with Fig. 4 in Fig. 3; Inoculate forefront point downright bad after 48 hours and the distance 2.1cm of vaccination.
Embodiment 2
One quick method that grows tobacco damping off artificial onset is inoculated tobacco rhizoctonia solani bacterial strain Rs-1 on the small seedling of tobacco bred cloud and mist 87, implements by the step 1 of embodiment 1 to step 5 operation.Wherein change in the operation with embodiment 1 step 5 of the operation of step 5, after the step 5) inoculation of embodiment 1, cultivation temperature maintains 25～27 ℃, and after the inoculation of the step 5) of the present embodiment, cultivation temperature maintains 19～20 ℃.Result also shows the downright bad symptom of blade clearly as the same with Fig. 4 in Fig. 3; Inoculate forefront point downright bad after 48 hours and the distance 0.6cm of vaccination.
Embodiment 3
One quick method that grows tobacco damping off artificial onset is inoculated tobacco rhizoctonia solani bacterial strain Rs-1 on the small seedling of tobacco bred cloud and mist 87, implements by the step 1 of embodiment 1 to step 5 operation.Wherein in step 2) operation with embodiment 1 step 2) operation change, the step 2 of embodiment 1) the cultivation duration of inoculum mycelia is 48 hours, the step 2 of the present embodiment) the cultivation duration of inoculum mycelia is 96 hours.Result shows the symptom of blade necrosis clearly as Fig. 3～Fig. 5 equally, but scab expansion speed is slower than the speed of embodiment 1, inoculates forefront point downright bad after 48 hours and the distance 0.8cm of vaccination.
Claims (1)
1. a quick method that grows tobacco damping off artificial onset, is characterized in that, utilizes rhizoctonia solani can infect tobacco leaf to cause the characteristic of necrosis, inoculates immature cigarette seedling in culture dish, causes cigarette seedling leaf to be fallen ill fast, shows clear symptom;
The step of method is as follows:
1) tobacco seedling is cultivated and is prepared:
With the common better and loose soil of tray splendid attire fertility, at indoor culture cigarette seedling, grow to after 3～4 vanelets for subsequent use by conventional method;
2) inoculate mycelial preparation:
Rhizoctonia solani is transplanted to the dull and stereotyped center of the common fungi culture medium of PSA, PSA medium component is: 200 grams of potatos, 20 grams of sucrose, 18 grams, agar, water 1000mL, under 28 ℃ of conditions, be cultured to and grow compared with macrocolony, cut-off footpath is that 6mm card punch is beaten and got mycelia agar nahlock of the same size in flat-plate bacterial colony periphery, makes inoculum with this mycelia piece;
3) before tobacco seedling inoculation, dispose:
Get the applicable seedling that step 1) cultivates, the moistening softening soil of growing seedlings that first waters before inoculation, so that lift seedlings, takes out clean culture dish and is placed in common shallow square plate, and add thin layer clear water in culture dish; Choose the seedling that has 3～4 vanelets, gently pull up and forward in the clear water of other container splendid attire with have gentle hands, rock gently the soil in wash-out seedlings root, seedling is forwarded in the above-mentioned culture dish of getting ready, leaflet horse is put on the ware edge of culture dish, on root system leans against at the bottom of ware, make seedling be upright state, and make clear water submergence root system;
4) inoculation rhizoctonia solani:
Picking step 2) the mycelia piece prepared, be placed with gently in step 3) and dispose on the lobus cardiacus of complete seedling or on heart bud;
5) after inoculation, dispose:
After step 4) operation is complete, the culture dish of splendid attire being inoculated to seedling is placed in shallow square plate, and plexiglass tent is covered on shallow square plate, in shallow square plate, injects thin layer clear water, makes plexiglass tent enclose inside and forms stable high wet condition in cover; Then package unit is put in to the general room cultivation that possesses daily scattered light, cultivation temperature maintains 25～30 ℃, notes avoiding direct sunlight according to shining;
Inoculation result causes seedling leaves to occur water stain shape necrosis, and last whole seedling total necrosis is rotted.Under normal circumstances, visible obviously necrosis afterwards shows to inoculate 1 day, and after 3～4 days, complete stool is rotted.
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Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN105284450A (en) * | 2015-09-21 | 2016-02-03 | 广西大学 | Method for quickly inducing rice blast via box type seedling nursing |
Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1287175A (en) * | 2000-09-08 | 2001-03-14 | 广东省农业科学院蔬菜研究所 | Water culture bacterination process to determine bacterial wilt resistance of plant |
| CN1709030A (en) * | 2005-07-27 | 2005-12-21 | 中国农业科学院蔬菜花卉研究所 | Method for identifying bacterial wilt resistivity of solanaceous crops |
| CN101496481A (en) * | 2009-03-09 | 2009-08-05 | 云南省烟草科学研究所 | Method for identifying resistance to tobacco bacterial wilt during seedling stage |
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Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1287175A (en) * | 2000-09-08 | 2001-03-14 | 广东省农业科学院蔬菜研究所 | Water culture bacterination process to determine bacterial wilt resistance of plant |
| CN1709030A (en) * | 2005-07-27 | 2005-12-21 | 中国农业科学院蔬菜花卉研究所 | Method for identifying bacterial wilt resistivity of solanaceous crops |
| CN101496481A (en) * | 2009-03-09 | 2009-08-05 | 云南省烟草科学研究所 | Method for identifying resistance to tobacco bacterial wilt during seedling stage |
Non-Patent Citations (2)
| Title |
|---|
| MARLENY GONZALEZ等: "Tobacco leaf spot and root rot caused by Rhizoctonia solani Kühn", 《MOLECULAR PLANT PATHOLOGY》, vol. 12, no. 3, 30 June 2011 (2011-06-30) * |
| 吴元华等: "烟草靶斑病原鉴定及生物学特性研究", 《沈阳农业大学学报》, vol. 43, no. 5, 31 October 2012 (2012-10-31) * |
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN105284450A (en) * | 2015-09-21 | 2016-02-03 | 广西大学 | Method for quickly inducing rice blast via box type seedling nursing |
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