CN105325195B - Method for obtaining wheat plant infected with Tilletia controversa Kuhn on ear - Google Patents
Method for obtaining wheat plant infected with Tilletia controversa Kuhn on ear Download PDFInfo
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- 241000209140 Triticum Species 0.000 title claims abstract description 85
- 238000000034 method Methods 0.000 title claims abstract description 34
- 241000722096 Tilletia controversa Species 0.000 title abstract description 13
- 235000021307 Triticum Nutrition 0.000 claims abstract description 60
- 239000000725 suspension Substances 0.000 claims abstract description 21
- 239000002689 soil Substances 0.000 claims abstract description 18
- 241000196324 Embryophyta Species 0.000 claims abstract description 14
- 238000011282 treatment Methods 0.000 claims abstract description 14
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Chemical compound O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims abstract description 14
- 230000001954 sterilising effect Effects 0.000 claims abstract description 12
- 238000005406 washing Methods 0.000 claims abstract description 9
- 229920000136 polysorbate Polymers 0.000 claims abstract description 7
- 238000011160 research Methods 0.000 claims abstract description 7
- 238000002360 preparation method Methods 0.000 claims abstract description 6
- 208000015181 infectious disease Diseases 0.000 claims description 19
- 239000001963 growth medium Substances 0.000 claims description 6
- 238000002347 injection Methods 0.000 claims description 6
- 239000007924 injection Substances 0.000 claims description 6
- 238000009630 liquid culture Methods 0.000 claims description 6
- 238000004659 sterilization and disinfection Methods 0.000 claims description 6
- 229920001817 Agar Polymers 0.000 claims description 5
- 239000008272 agar Substances 0.000 claims description 5
- SUKJFIGYRHOWBL-UHFFFAOYSA-N sodium hypochlorite Chemical compound [Na+].Cl[O-] SUKJFIGYRHOWBL-UHFFFAOYSA-N 0.000 claims description 5
- 239000012528 membrane Substances 0.000 claims description 4
- 238000012545 processing Methods 0.000 claims description 4
- 238000007789 sealing Methods 0.000 claims description 4
- 239000000243 solution Substances 0.000 claims description 4
- 238000009835 boiling Methods 0.000 claims description 2
- 238000001914 filtration Methods 0.000 claims description 2
- 239000000843 powder Substances 0.000 claims description 2
- 238000011081 inoculation Methods 0.000 abstract description 12
- 230000035784 germination Effects 0.000 abstract description 4
- 244000005700 microbiome Species 0.000 abstract description 2
- 239000008223 sterile water Substances 0.000 abstract description 2
- 238000012258 culturing Methods 0.000 abstract 2
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- 241000495841 Oenanthe oenanthe Species 0.000 abstract 1
- 201000010099 disease Diseases 0.000 description 11
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 description 11
- 230000001580 bacterial effect Effects 0.000 description 5
- 241000894006 Bacteria Species 0.000 description 4
- 239000006185 dispersion Substances 0.000 description 4
- 239000000463 material Substances 0.000 description 4
- 238000010899 nucleation Methods 0.000 description 4
- 241000031845 Tilletia laevis Species 0.000 description 3
- 238000005286 illumination Methods 0.000 description 3
- 206010018498 Goitre Diseases 0.000 description 2
- 241000722133 Tilletia Species 0.000 description 2
- 244000052616 bacterial pathogen Species 0.000 description 2
- 239000003153 chemical reaction reagent Substances 0.000 description 2
- 238000005034 decoration Methods 0.000 description 2
- 238000001514 detection method Methods 0.000 description 2
- 230000000694 effects Effects 0.000 description 2
- 239000003292 glue Substances 0.000 description 2
- 201000003872 goiter Diseases 0.000 description 2
- 210000001161 mammalian embryo Anatomy 0.000 description 2
- 208000035404 Autolysis Diseases 0.000 description 1
- 206010057248 Cell death Diseases 0.000 description 1
- 241000233866 Fungi Species 0.000 description 1
- 241000238631 Hexapoda Species 0.000 description 1
- 241001465754 Metazoa Species 0.000 description 1
- 108091092878 Microsatellite Proteins 0.000 description 1
- 241000244206 Nematoda Species 0.000 description 1
- 241000238370 Sepia Species 0.000 description 1
- 241000221563 Ustilaginaceae Species 0.000 description 1
- 238000013459 approach Methods 0.000 description 1
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- 230000015572 biosynthetic process Effects 0.000 description 1
- 210000004556 brain Anatomy 0.000 description 1
- 235000013339 cereals Nutrition 0.000 description 1
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- 238000000576 coating method Methods 0.000 description 1
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- 230000007850 degeneration Effects 0.000 description 1
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- 238000003745 diagnosis Methods 0.000 description 1
- 239000012153 distilled water Substances 0.000 description 1
- 238000002474 experimental method Methods 0.000 description 1
- 230000036541 health Effects 0.000 description 1
- 238000010438 heat treatment Methods 0.000 description 1
- 239000002054 inoculum Substances 0.000 description 1
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- 230000009885 systemic effect Effects 0.000 description 1
- 230000035899 viability Effects 0.000 description 1
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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
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Abstract
The invention provides a method for obtaining a wheat plant infected with Tilletia controversa Kuhn on ears, belonging to the field of agricultural microorganisms. The method comprises the following steps: (1) preparation of germinated winter spores suspension: culturing the teliospore of Tilletia controversa Kuhn until germination, washing the germinated teliospore with 2 ml/dish of sterile water, and adding 0.02% Tween to obtain teliospore suspension; (2) culturing a wheat plant: sterilizing and accelerating germination of wheat seeds, performing vernalization treatment, and planting the vernalized wheat seeds in sterilized soil until the booting stage; (3) inoculation: and injecting the winter spore suspension into a leaf sheath 2.5cm above the uppermost stem node of the wheat ear. The method can efficiently obtain the wheat pathogenic plants infected with the Tilletia controversa Kuhn on ears, and the wheat pathogenic plants are applied to the research of the Tilletia controversa Kuhn.
Description
Technical field
The invention belongs to field of agricultural microorganism, and in particular to obtain the wheat plant of fringe portion infection T contraversa
Method.
Background technique
T contraversa, i.e. T. contraversa (Tilletia controversa K ü hn, TCK) belong to true
It is a kind of important international quarantine venereal disease evil that Mycota Ustilaginaceae Tilletia, which is caused dwarf bunt of wheat, to small
One of the dominant species that wheat production causes serious harm and China's invasive plants to study.
The morphological feature of T contraversa is as follows: teleutosorus is born in ovary more, forms the spore ball of black powdery,
That is smut goitre, each mycoceicidum contain teleutospore 100,000 to 1,000,000 depending on of different sizes.Winter sorosphere shape or subsphaeroidal, it is yellowish-brown
For color to dark sepia, it is mostly 19-23um that average diameter and standard deviation, which are 20.90+/- 0.72, but occasionally has 17um or 30um
(including glue sheath).The polygonal net eye shape decoration of sclerine, mesh general diameter 3-5um, once in a while in brain line shape or irregularly
Shape, meat ridge average height are 1.425+/- 0.144um, and transparent colloid sheath coating, sterile cell's spherical shape or nearly ball are with outside sporoderm
Shape, colorless and transparent or micro- green have glue sheath sometimes, and diameter is usually less than teleutospore 9-16um, once in a while up to 22um, surface light
Sliding, sporoderm is without decoration.
The biological characteristics of T contraversa are as follows: T. contraversa teliospore germination needs consecutive low temperature,
Generally between 3 DEG C -8 DEG C, and with 4-6 DEG C for optimum temperature, minimum 0 DEG C, up to 10 DEG C.When temperature is 4-6 DEG C,
Under illumination condition, teleutospore usually sprouts after 3-5 weeks, and individual bacterial strains started to sprout at the 16th day, and a small number of bacterial strains are through 7-10
Zhou Houcai starts to sprout.Higher or lower than suitable temperature range, spore germination period accordingly extends, and at 0 DEG C or so, spore is after 8 weeks
Just start to sprout, and generates normal promycelium and spore and secondary microspore.When 10 DEG C, spore starts to sprout after 8 weeks,
Elongated, lopsided promycelium is generated, seldom formation microspore more, and often has autolysis.Cause of disease teleutospore has extremely strong degeneration-resistant
Property, at room temperature, the service life is at least 4 years, and some is up to 7 years, the teleutospore in sick goitre, and the service life in the soil is 3-
7 years, for the teleutospore of dispersion then more than at least a year, germ still had comparable viability in company with after feed feeding domestic animal.The cause of disease winter
Spore heat hardiness is extremely strong, under dry heat condition, need to could inactivate through 130 DEG C of half an hour, and damp and hot, needs 80 DEG C, 20 minutes can cause
Extremely.
In the research of the infection mechanism to wheat light Tilletia foetida, it usually needs be seeded to it by manual method small
Wheat plant makes the latter infect to obtain and catch an illness wheat plant to observe and analyze, and some artificial infections exist in the prior art
Method, for example, the plumule wound monospore inocalation method of the easily creations such as Jianping, i.e., scratch the embryo of germination seed with aseptic operation pocket knife
At the top of bud scale, about 4~5mm is cut2It is counter to be affixed on plumule wound with the single agar fritter for sprouting TCK teleutospore, every plant
Wheat seeding is inoculated with 1 sprouting spore.Inoculation wheat seeding is cultivated under the conditions of being placed in 5 DEG C of illumination (12h/d).And plumule monospore inoculation
Method cuts about 4~5mm with aseptic operation pocket knife2It is counter to be affixed on plumule with the single agar fritter for sprouting TCK spore,
Every plant of wheat seeding is inoculated with 1 sprouting spore.Inoculation wheat seeding is cultivated under the conditions of being placed in 5 DEG C of illumination (12h/d).Through both artificial infections
The wheat plant disease incidence that method obtains is respectively 4.44% and 0.44%.2014 Nian Yaozhuo et al. are respectively with 4 kinds of different people
Work inoculation method carries out systemic infection to wheat seed to wheat light Tilletia foetida, and only the first is obtained with the third method
3.33% and 6.66% morbidity disease fringe.
Lack a kind of Inoculation Method that the part seed only for part booting is infected on this field.
Summary of the invention
The present invention provides a kind of short by artificial infection acquisition fringe portion infection wheat for blank existing for above-mentioned field
The wheat plant method of Tilletia foetida.
Technical scheme is as follows:
A method of obtaining the wheat plant of fringe portion infection T contraversa, which is characterized in that including walking as follows
It is rapid:
(1) the teleutospore suspension that preparation is sprouted: the teleutospore of T contraversa is cultivated to sprouting, after sprouting
The aseptic water washing of teleutospore 2ml/ ware, adds 0.02% tween, and teleutospore suspension is made；
(2) it cultivates wheat plant: vernalization treatment will be carried out after wheat seed sterilizing vernalization, the wheat seed after vernalization is planted
Kind is in sterilized soil to boot stage；
(3) it is inoculated with: the teleutospore suspension is injected into the leaf sheath above the most upper stipes of wheat tassel at 2.5cm
It is interior.
Culture described in step (1) refers to, with 2% soil extract liquid culture medium, trains in 5 DEG C of artificial climate grown cultures casees
Teleutospore 15 days of culture T contraversa are inverted in full exposure in supporting.
The concentration of the teleutospore suspension is 100 × 104~105 × 104A/mL, the injection volume of every fringe wheat are 1ML.
The wheat seed sterilizing vernalization, with 30%NaClO solution to wheat seed surface sterilization 5min, aseptic water washing
After 5 times, by wheat seed presoaking and germinating, up to coleoptile length to 1~3mm.
The vernalization treatment refers to that the wheat seed dispersion after vernalization is laid on the wet filter paper in culture ware lid, sealed membrane
Sealing, is placed in 5 DEG C of incubator, 4 weeks low temperature vernalization treatments, until seedling is long to 3~5cm.
The condition of culture described in step (2) is 25 DEG C of whole day, relative humidity 50%, full exposure processing, watering one overnight
It is secondary.
The boot stage refers to, wheat plant grows 30 days to 25~35cm of plant height under conditions of step (2) described culture,
Blade is long to 7 leaves.
Application of any method in terms of T contraversa research.
The present invention provides a kind of method that artificial infection obtains the wheat plant of fringe portion infection T contraversa, packets
Include following steps:
(1) the teleutospore suspension that preparation is sprouted: the teleutospore of T contraversa is cultivated to sprouting, after sprouting
The aseptic water washing of teleutospore 2ml/ ware, adds 0.02% tween, and teleutospore suspension is made；The winter spore being prepared
Sub- suspension has the suspended concentration for meeting inoculation condition, and the tween for being added 0.02% is to play a protective role to teleutospore.
(2) it cultivates wheat plant: vernalization treatment will be carried out after wheat seed sterilizing vernalization, the wheat seed after vernalization is planted
Kind is in sterilized soil to boot stage；The effect that sterilized soil cultivation is sterilized and used to wheat seed is to prevent doing for other germs
It disturbs to guarantee for wheat plant to be infected to be sterile before inoculation；The effect of vernalization treatment is to guarantee wheat plant
Later period can smoothly breed solid.
(3) it is inoculated with: the teleutospore suspension is injected into the leaf sheath above the most upper stipes of wheat tassel at 2.5cm
It is interior.Inoculation can achieve the purpose for only infecting part fringe grain in this way.
Culture described in step (1) refers to, with 2% soil extract liquid culture medium, trains in 5 DEG C of artificial climate grown cultures casees
Teleutospore 15 days of culture T contraversa are inverted in full exposure in supporting.What is obtained under this condition of culture and culture environment sprouts
The silk that infects that the teleutospore of hair can produce infection ability help to obtain the teleutospore suspension for meeting inoculation condition.
The concentration of the teleutospore suspension is 100 × 104~105 × 104A/mL.Teleutospore suspension under the concentration is used
It is more excellent in being inoculated with.
The wheat seed sterilizing vernalization refers to, with 30%NaClO solution to wheat seed surface sterilization 5min, sterile water punching
After washing 5 times, by wheat seed presoaking and germinating, up to coleoptile length to 1~3mm.30% liquor natrii hypochloritis disappears to wheat seed
The purpose of poison is to kill the various germs that wheat seed itself carries, to ensure that the wheat seed before being inoculated with is sterile.Embryo
The long plantation for being more suitable for the later period to the seed of 1~3mm of bud scale.
The vernalization treatment refers to that the wheat seed dispersion after vernalization is laid on the wet filter paper in culture ware lid, sealed membrane
Sealing, is placed in 5 DEG C of incubator, 4 weeks low temperature vernalization treatments, until seedling is long to 3~5cm.By this vernalization treatment step
Wheat seedling can ensure that the later period has developed into and breed the active wheat head.
The condition of culture described in step (2) is 25 DEG C of whole day, relative humidity 50%, full exposure processing, watering one overnight
It is secondary.Such condition of culture is more suitable for the growth of the quick health of wheat, and the strain of boot stage wheat can be obtained in the short time.
The boot stage refers to, wheat plant grows 30 days to 25~35cm of plant height under conditions of step (2) described culture,
Blade is long to 7 leaves.Inoculation of the invention is carried out in long be more advantageous to this period of wheat plant to obtain part disease fringe and portion
Divide infected seed.
Application of the method in terms of T contraversa research is also claimed in the present invention.
Verified statistics infects wheat booting period using method artificial infection T contraversa of the present invention and plants
Strain, can obtain the wheat plant with the susceptible wheat head of the susceptible wheat in part or part, disease incidence is up to 92%.
Detailed description of the invention
Fig. 1 shows tri-leaf period is grown to after wheat cultivation, growing way is almost the same；
Injection connects bacterium when after A being indicated wheat growth 30 days in Fig. 2 to boot stage；B indicates same time plant height 33cm, 7 leaves
Whole strain wheat；C indicates the small fringe portion isolated by B；
A indicates that white arrow shows the single infected seed after injection connects bacterium 25 days on sick fringe in Fig. 3；B indicates that white arrow shows note
Penetrate the single infected seed after connecing bacterium 35 days on sick fringe.
Specific embodiment
Product described in present invention be described in more detail combined with specific embodiments below, but the present invention is not limited with this
Range.Unless otherwise specified, following used reagents and material are commercially available, and used method is conventional behaviour
Make.
Biomaterial
Wheat breed: the winter selects No. 3, is known kind, commercially available.
T contraversa (Tilletia controversa K ü hn) bacterial strain: all TCK bacterial strains of the present invention are existing
There is document L.GAO et al.Development of a SCAR Maker by inter-simple sequence repeat
for dignoisis of Dwarf Bunt of Wheat and Detection of Tilletia controversa Kü
Hn.Folia Microbiol.55 (3), 258-264 (2010) is recorded, referring to the record of wherein materials and methods part in beauty
The separation strains that state is separated to by professor B.Goates.In addition also it is documented in L.GAO et al.AnISSR-based Approach
for the Molecular Detection and Diagnosis of Dwarf Bunt of Wheat,Caused by
Tilletia controversaKu ¨ hn.J Phytopathol159:155-158 (2011), for wherein materials and methods part
TCK bacterial strain provided by the Dr Blair Goates of record.Also there is preservation in this laboratory, can from the applying date in 20 years to
The public, which provides, is used for confirmatory experiment.
Reagent and consumptive material
Agar, distilled water, NaClO, tween are commercially available.
2% soil extract liquid culture medium, the preparation method is as follows: weigh soil 75g, 500ml boiling water, with 8 layers of filtered through gauze,
Agar powder 20g is added after filtering, water is added to be settled to 1L, 121 DEG C, 20min, moist heat sterilization.
Embodiment 1, the method for the invention artificial infection T contraversa and infect result observation statistics
Present embodiments provide a kind of wheat plant that fringe portion infection T contraversa is obtained through artificial infection
Method, concrete operations are as follows:
The sterilizing and vernalization of wheat seed: 5min is sterilized with 30% NaClO solution surface, aseptic water washing 5 times, is removed
The T contraversa that wheat seed surface may carry.By wheat seed presoaking and germinating, up to coleoptile length to 1~3mm.
Soil disinfection: Nutrition Soil is put into high-pressure sterilizing pot, 60~100 DEG C of heating continue 30~60min, kill soil
Interior most bacterium, fungi, nematode and insect.
Vernalization treatment: the seeded dispersion after vernalization is laid on the wet filter paper in culture ware lid, and sealed membrane sealing is placed in 5
DEG C incubator in, 4 weeks low temperature vernalization treatments, until seedling is long to 3~5cm.
Seed: being planted in sterilized soil by wheat condition of culture after 4 weeks, routine observation wheat growth situation, until mature.
Greenhouse temperature is arranged 25 DEG C of whole day, and relative humidity 50%, full exposure processing, watering overnight is once.
Tri-leaf period is grown to after wheat cultivation, growing way is almost the same, as shown in Figure 1.After normal plants 30 days, wheat
25~35cm of plant height, blade is long to 7 leaves, and no tiller phenomenon (Fig. 2 B), removing whole strain discovery has small ear (Fig. 2 C) occur, at this time for
Boot stage starts injection inoculation.
It is prepared by the teleutospore suspension of sprouting: using the sprouting teleutospore of the 2% soil extract liquid culture medium culture germ,
Its suspension concentration be 100 × 104~105 × 104/mL, be placed in 5 DEG C of artificial climate grown cultures casees (LT-36VLC8,
PERCIVAL, USA) full exposure be inverted culture, after 15 days automatically be inverted research grade microscope (IX83, OLYMPUS,
Japan the sprouting situation of teleutospore is observed under).By the aseptic water washing of 2ml/ ware of the teleutospore in sprouting, add
0.02% tween.
Injecting method: being injected into inoculum in booting with a syringe, and the injection volume of every fringe wheat is 1ML.It will
Suspension is injected into the leaf sheath above the most upper stipes that tassel is being developed at 2.5cm, and about boot leaf is just born at this time
Stage (Fig. 2A), plant plant in the greenhouse to maturation.This method generates the part disease fringe of tool part infected seed after 3 weeks
(Fig. 3).
Through counting, 92% is up to using the artificial infection wheat plant disease incidence that the present embodiment the method obtains.Thus
As it can be seen that method provided by the present embodiment can efficiently obtain the wheat disease plant of fringe portion infection T contraversa,
And these wheat disease plants are applied to the research of T contraversa.
Claims (4)
1. a kind of method for the wheat plant for obtaining fringe portion infection T contraversa, which comprises the steps of:
(1) the teleutospore suspension that preparation is sprouted: the teleutospore of T contraversa is cultivated to sprouting, by the winter spore after sprouting
The son aseptic water washing of 2ml/ ware, adds 0.02% tween, and teleutospore suspension is made；
(2) cultivate wheat plant: will wheat seed sterilize vernalization after carry out vernalization treatment, by after vernalization wheat seed plantation in
To boot stage in sterilized soil；
(3) it is inoculated with: the teleutospore suspension is injected into the leaf sheath above the most upper stipes of wheat tassel at 2.5cm,
The concentration of the teleutospore suspension is 100 × 104~105 × 104A/mL, the injection volume of every fringe wheat are 1ML；
The boot stage refers to that wheat plant grows 30 days to 25~35cm of plant height under conditions of step (2), and blade is long to 7 leaves；
Culture described in step (1) refers to, with 2% soil extract liquid culture medium, in 5 DEG C of artificial climate grown cultures case cultures
Teleutospore 15 days of culture T contraversa are inverted in full exposure；The preparation method of the 2% soil extract liquid culture medium is such as
Under: weigh soil 75g, agar powder 20g is added with 8 layers of filtered through gauze in 500ml boiling water after filtering, and add water to be settled to 1L, 121
DEG C, 20min, moist heat sterilization；
The condition of culture described in step (2) is 25 DEG C of whole day, relative humidity 50%, full exposure processing, and watering overnight is primary.
2. the vernalization the method according to claim 1, wherein the wheat seed sterilizes, with 30%NaClO solution
To wheat seed surface sterilization 5min, after aseptic water washing 5 times, by wheat seed presoaking and germinating, until coleoptile length to 1~
3mm。
3. the wheat seed after vernalization disperses the method according to claim 1, wherein the vernalization treatment refers to
It being laid on the wet filter paper in culture ware lid, sealed membrane sealing is placed in 5 DEG C of incubator, 4 weeks low temperature vernalization treatments, until
3~5cm of Miao Changzhi.
4. application of any method of claim 1-3 in terms of T contraversa research.
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Citations (6)
| Publication number | Priority date | Publication date | Assignee | Title |
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| CN101326885A (en) * | 2008-08-04 | 2008-12-24 | 翟庆军 | Method for preventing and treating wheat fungal diseases using biological preparation, plant stress-resistance chemical inducer |
| CN102399753A (en) * | 2011-12-16 | 2012-04-04 | 中国农业科学院植物保护研究所 | Specific monoclonal antibody of tilletia controversa kuhn and immunofluorescent detection method |
| RU2474101C2 (en) * | 2011-03-15 | 2013-02-10 | Михаил Васильевич Шаравара | Method of increase in productivity of grain crops |
| CN104221648A (en) * | 2013-08-13 | 2014-12-24 | 中国农业科学院植物保护研究所 | Method for cultivating tilletia controversa kuhn disease plants in indoor environments |
| CN104593332A (en) * | 2014-10-22 | 2015-05-06 | 中国农业科学院植物保护研究所 | Kit and method for detecting Tilletia controversa Kuhn |
| CN105039491A (en) * | 2015-07-10 | 2015-11-11 | 中国农业科学院植物保护研究所 | Method for identifying teliospores of Tilletia controversa Kuhn and Tilletia foetida |
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Patent Citations (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101326885A (en) * | 2008-08-04 | 2008-12-24 | 翟庆军 | Method for preventing and treating wheat fungal diseases using biological preparation, plant stress-resistance chemical inducer |
| RU2474101C2 (en) * | 2011-03-15 | 2013-02-10 | Михаил Васильевич Шаравара | Method of increase in productivity of grain crops |
| CN102399753A (en) * | 2011-12-16 | 2012-04-04 | 中国农业科学院植物保护研究所 | Specific monoclonal antibody of tilletia controversa kuhn and immunofluorescent detection method |
| CN104221648A (en) * | 2013-08-13 | 2014-12-24 | 中国农业科学院植物保护研究所 | Method for cultivating tilletia controversa kuhn disease plants in indoor environments |
| CN104593332A (en) * | 2014-10-22 | 2015-05-06 | 中国农业科学院植物保护研究所 | Kit and method for detecting Tilletia controversa Kuhn |
| CN105039491A (en) * | 2015-07-10 | 2015-11-11 | 中国农业科学院植物保护研究所 | Method for identifying teliospores of Tilletia controversa Kuhn and Tilletia foetida |
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