CN105779372A - Potato culture medium suitable for ustilaginoidea virens spore production and application thereof - Google Patents
Potato culture medium suitable for ustilaginoidea virens spore production and application thereof Download PDFInfo
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- CN105779372A CN105779372A CN201610345478.7A CN201610345478A CN105779372A CN 105779372 A CN105779372 A CN 105779372A CN 201610345478 A CN201610345478 A CN 201610345478A CN 105779372 A CN105779372 A CN 105779372A
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- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N3/00—Spore forming or isolating processes
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Abstract
The invention discloses a potato culture medium suitable for ustilaginoidea virens spore production and an application thereof. Potatoes are prepared into a plate culture medium as a spore production culture medium so as to culture and prepare ustilaginoidea virens thin-wall conidiospore. Preparation for the culture medium and an application method for the culture medium comprise the following steps: 1) preparing the culture medium; 2) inverting a culture flat plate; 3) implanting ustilaginoidea virens; 4) carrying out constant-temperature culture; and 5) collecting a new generation of spores. The potato culture medium has the characteristics and the advantages that: the culture medium is simple in composition; the operation is simple, easy and convenient, and shaking culture equipment is not needed; time of obtaining a great number of spores is short; a high spore production state lasts for a long time; and a spore liquid is relatively pure, is less in mixed mycelia, and is generally suitable for being used in a lot of research work without purifying treatment.
Description
Technical field
The present invention relates to plant pathology to learn a skill and microbiological technique.Specifically a kind of suitable ustilaginoidea virens produces potato culture and the application process thereof of spore.
Background technology
In most of fungal disease systems, the brood body of pathogen is except having breeding and expanding the effects such as its population quantity, the more important thing is to have and identify mutually with host, and start to infect the function of host, therefore, in many important research of plant disease processes, it is required for using the brood body of pathogen to make test material, it is clear that, the cultivation preparation method of efficient brood body material easily, will be very beneficial for carrying out smoothly of research work.False smut (is infected and causes) the great disease of one having become China's Rice Production by Ustilaginoideavirens (Cooke) Takahashi, the brood body of ustilaginoidea virens includes the ascospore of condition, the pachypycnidium of Invisible element and thin-walled conidium three types, in laboratory, research work so far mainly uses thin-walled conidium (hereafter by thin-walled conidium referred to as " spore ").
So far, the conidial cultivation technology of preparing of known ustilaginoidea virens thin-walled, mainly liquid cultivating method, the major technique step of the method is, first cultivates the mycelium preparing ustilaginoidea virens, then mycelium is implanted into fluid medium carries out shaken cultivation.Finding in practical work, this cultural method comes with some shortcomings, and as consuming time longer, whole process is it is generally required to more than 15 days；Need shaken cultivation equipment；Cultured products mixed liquor includes substantial amounts of hypha body, bacterial metabolism product and nutrient media components etc..Adopting in the cultured products of this technology, although include substantial amounts of spore, but to obtain comparatively simple spore liquid, generally require and filter the subsequent treatment such as mycelia and precipitation spore, owing to cultured products is viscous pasty state, precipitation spore generally requires high speed centrifugation machine equipment.
Nearest progress has been set up the conidial slat chain conveyor technology of preparing of ustilaginoidea virens thin-walled, and this technical method adopts conventional potato sucrose agar culture medium cultivation to prepare spore.Prepare spore with potato sucrose agar culture medium and can overcome some disadvantages of liquid cultivating method.But there is also with the technology of this medium preparing spore that some are substantially not enough, after producing spore peak, flat-plate bacterial colony quickly grows substantial amounts of non-product spore aerial hyphae, causes that spore quantity sharply declines, it is impossible to the stable offer spore within the long term uses；And collect in the new spore liquid of preparation cultivation results and still suffer from broken section of more mycelia.
Summary of the invention
It is an object of the invention to provide a kind of suitable ustilaginoidea virens and produce potato culture and the application process thereof of spore, utilize Rhizoma Solani tuber osi to make plating medium, as producing spore culture medium, ustilaginoidea virens thin-walled conidium is prepared in cultivation.
The technical scheme is that
A kind of suitable ustilaginoidea virens produces potato culture and the application process thereof of spore, and operating procedure is as follows:
1. medium preparing
In parts by weight: Rhizoma Solani tuber osi 100g, agar 20g, water 1000ml, prepare potato culture, standby after conventional high temperature sterilizing.
2. fall culture plate
In the usual way potato culture prepared by step 1 is melted and pour culture dish into and make culture plate.
3. implant ustilaginoidea virens
Take out with the standby ustilaginoidea virens spore liquid deposited of spore form, the culture plate under aseptic condition, this spore liquid implantation step 2 got ready, with T-shaped glass rod, spore liquid is spread evenly across in platen surface.
4. constant temperature culture
The culture plate of step 3 end of operation is proceeded in incubator, 28 DEG C of constant temperature culture.
5. the collection of a new generation's spore
After step 4 operation is cultivated 5 days, ustilaginoidea virens can form a large amount of a new generations thin-walled conidium on culture plate；Culture plate is forwarded on superclean bench, scrub the bacterium colony on flat board with sterilized water, new pure spore liquid can be obtained.
Advantages of the present invention:
1) culture medium composition is simple.
2) obtain a large amount of spore required time short, normally only need 4～6 days；Persistent period is long, can maintain the big volume production spore time more than 15 days.
3) operate simple and easy convenient, it is not necessary to shaken cultivation equipment；
4) spore liquid is comparatively pure, and the mycelia mixed is broken less, also is adapted for the use of many research work typically without purified treatment.
Accompanying drawing explanation
Fig. 1 utilizes the inventive method to cultivate ustilaginoidea virens after 5 days, and the minute colony grown on potato culture flat board forms a large amount of a new generations spore on bacterium colony.
Fig. 2 utilizes the inventive method to cultivate ustilaginoidea virens after 20 days, and the minute colony on potato culture flat board still keeps a large amount of a new generations spore on bacterium colony.
Detailed description of the invention
Below in conjunction with accompanying drawing, the invention will be further described with embodiment.
Potato sucrose culture medium is a kind of culture medium that laboratory is conventional, its carbon nutrition is mainly sucrose, so far find, the growth promoter optimum carbon element of ustilaginoidea virens is also precisely sucrose, thus the cultivation of current ustilaginoidea virens, especially the conidial preparation of thin-walled is cultivated, and mainly uses potato sucrose culture medium.Simultaneously because the understanding of ustilaginoidea virens sporulating character is limited, have no always and set up flat board product spore technology, cause the conidial preparation practice of thin-walled, use liquid culture technology for a long time always.
Slat chain conveyor technology prepared by the ustilaginoidea virens spore of nearest first public foundation, the potato sucrose culture medium that its culture medium used is also is carbon nutrition with sucrose.This technology utilizes the characteristic of the big volume production spore of minute colony of ustilaginoidea virens, collects spore during cultivating formation minute colony.Inventor, utilizing above-mentioned slat chain conveyor technology to prepare discovery in the practice of ustilaginoidea virens spore, generally in cultivating 5～7 days, can collect a large amount of spore.After cultivating 8 days, spore quantity sharply declines, and reason is that after producing spore peak, bacterium colony continued growth forms vigorous non-sporogenous hyphae, particularly aerial hyphae, is hidden by originally small product spore bacterium colony.Visible, the actual product spore time of this culture technique is very short.In order to dock in requisition for the work of spore with downstream in practice, generally require and work in every process is carried out stricter Synchronization Design, otherwise easily miss product spore peak period.
Owing to ustilaginoidea virens nourishing and growing in potato sucrose culture medium is all compared vigorous with reproductive growth, thus for ustilaginoidea virens growth promoter on single potato culture, it does not have people pays close attention and tests.It is more weak that inventor once tested discovery ustilaginoidea virens growth promoter in single potato culture, and therefore this potato culture does not cause the attention of inventor for a long time yet.Work recently chances on, Rhizoma Solani tuber osi plating medium is implanted ustilaginoidea virens spore, also formation petite can be grown after spore germination growth, bacterium colony also produces the new spore of substantial amounts of offspring, and bacterium colony is not easy grow non-sporogenous hyphae, the big volume production spore state of bacterium colony can keep more than 2 weeks, thus, Rhizoma Solani tuber osi plating medium is actually ustilaginoidea virens and desirably produces spore culture medium；The composition adding potato culture is simply and readily prepared, considerably advantageous application in practice；Obviously, utilize the spore technology of preparing of Rhizoma Solani tuber osi plating medium, very simple and efficient, can guarantee that again within one period, the spore output that preparation obtains is relatively stable.
The quantity of the spore of new generation that preparation cultivation results obtains is relevant with the quantity forming petite on flat board, petite quantity is then female with transplanting relevant for spore quantity, when being not resulted in petite overlap, female more many for spore, cultivate the spore of new generation obtained more many；Generally the mother being implanted into every ware flat board (culture dish diameter is 9cm) is controlled 10 for spore count4The order of magnitude is advisable；When the mother stocked is 10 for spore concentration6Individual/ml time, every ware flat board can be coated with that 10～100 μ l are female makes transplant, preparation a new generation spore for spore liquid.
Final step eluting is collected preparation and is cultivated the spore of new generation obtained, and in superclean bench sterile working, can obtain the spore liquid not having to pollute.If without considering pollution problem, then on routine experimentation table, spore can be collected by eluting.
Embodiment 1
Adopting a kind of suitable ustilaginoidea virens of the present invention to produce potato culture and the application process thereof of spore, the thin-walled conidium of ustilaginoidea virens bacterial strain Uv-108 is prepared in cultivation, implements operation as follows:
1. medium preparing
In parts by weight: Rhizoma Solani tuber osi 100g, agar 20g, water 1000ml, prepare potato culture, standby after conventional high temperature sterilizing.
2. fall culture plate
In the usual way potato culture prepared by step 1 is melted and pour 9cm culture dish into and make culture plate.
3. implant ustilaginoidea virens
With the standby ustilaginoidea virens bacterial strain Uv-108 spore liquid deposited of spore form, (concentration is for 10 in taking-up6Individual/ml), the culture plate under aseptic condition, this spore liquid 50 μ l implantation step 2 got ready, with T-shaped glass rod, spore liquid is spread evenly across in platen surface.
4. constant temperature culture
The culture plate of step 3 end of operation is proceeded in incubator, 28 DEG C of constant temperature culture.
5. the collection of a new generation's spore
After step 4 operation is cultivated 5 days, bacterial strain Uv-108 can form a large amount of a new generations thin-walled conidium on culture plate；Scrub a ware flat-plate bacterial colony with sterilized water, 1.0 × 10 can be obtained6The new spore liquid 61ml of individual spore/ml.
Embodiment 2
Adopting a kind of suitable ustilaginoidea virens of the present invention to produce potato culture and the application process thereof of spore, the thin-walled conidium of ustilaginoidea virens bacterial strain Uv-110 is prepared in cultivation, implements by the step 1 of embodiment 1 to 5 operation, and the bacterial strain that simply step 3 is implanted is Uv-110.After implementing step 5 operation, scrub a ware flat-plate bacterial colony with sterilized water, 1.0 × 10 can be obtained6The new spore liquid 54ml of individual spore/ml.
Embodiment 3
Adopting a kind of suitable ustilaginoidea virens of the present invention to produce potato culture and the application process thereof of spore, the thin-walled conidium of ustilaginoidea virens bacterial strain Uv-111 is prepared in cultivation, implements by the step 1 of embodiment 1 to 5 operation, and the bacterial strain that simply step 3 is implanted is Uv-111.After implementing step 5 operation, scrub a ware flat-plate bacterial colony with sterilized water, 1.0 × 10 can be obtained6The new spore liquid 70ml of individual spore/ml.
Claims (1)
1. a suitable ustilaginoidea virens produces potato culture and the application process thereof of spore, it is characterised in that utilize Rhizoma Solani tuber osi to make plating medium, and as producing spore culture medium, ustilaginoidea virens thin-walled conidium is prepared in cultivation；The preparation of culture medium and the operating procedure of application process thereof are as follows:
1) medium preparing: in parts by weight: Rhizoma Solani tuber osi 100g, agar 20g, water 1000ml, prepares potato culture, standby after conventional high temperature sterilizing；
2) culture plate is fallen: in the usual way by step 1) potato culture prepared melts and pours culture dish into and make culture plate；
3) ustilaginoidea virens is implanted: take out with the standby ustilaginoidea virens spore liquid deposited of spore form, by this spore liquid implantation step 2 under aseptic condition) culture plate got ready, with T-shaped glass rod, spore liquid is spread evenly across in platen surface；
4) constant temperature culture: the culture plate of step 3 end of operation is proceeded in incubator, 28 DEG C of constant temperature culture；
5) collection of a new generation's spore: step 4) operation cultivates after 5 days, and ustilaginoidea virens can form a large amount of thin-walled conidium of new generation on culture plate；Culture plate is forwarded on superclean bench, scrub the bacterium colony on flat board with sterilized water, new pure spore liquid can be obtained.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201610345478.7A CN105779372A (en) | 2016-05-24 | 2016-05-24 | Potato culture medium suitable for ustilaginoidea virens spore production and application thereof |
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| CN201610345478.7A CN105779372A (en) | 2016-05-24 | 2016-05-24 | Potato culture medium suitable for ustilaginoidea virens spore production and application thereof |
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Cited By (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN106635948A (en) * | 2017-01-10 | 2017-05-10 | 广西大学 | Pollution-free preparing and culturing method for rice-false-smut-case thin-wall conidia |
| CN106701653A (en) * | 2017-01-10 | 2017-05-24 | 广西大学 | Pollution-reduction preparation and culture method of ustilaginoidea virens thin-wall conidia |
| CN106754424A (en) * | 2017-01-10 | 2017-05-31 | 广西大学 | The standing form and its preparation and application method of a kind of ustilaginoidea virens lab material |
| CN106867957A (en) * | 2017-01-10 | 2017-06-20 | 广西大学 | A kind of ustilaginoidea virens thin-walled of Pollution protection is conidial to prepare cultural method |
| CN110511859A (en) * | 2019-08-22 | 2019-11-29 | 广西农业职业技术学院 | A kind of equipment and working method preparing potato culture |
Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103436480A (en) * | 2013-08-27 | 2013-12-11 | 广西大学 | Plate culture and preparation method of ustilaginoidea virens thin-wall conidium |
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2016
- 2016-05-24 CN CN201610345478.7A patent/CN105779372A/en active Pending
Patent Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103436480A (en) * | 2013-08-27 | 2013-12-11 | 广西大学 | Plate culture and preparation method of ustilaginoidea virens thin-wall conidium |
Non-Patent Citations (3)
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| 张君成等: "稻曲病菌分生孢子的生物学研究", 《植物病理学报》 * |
| 张君成等: "糖对稻曲病菌薄壁分生孢子萌发的影响", 《中国植物病理学会2008年学术年会论文集》 * |
| 成卓敏等: "《粮食安全与植保科科创新》", 30 September 2009 * |
Cited By (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN106635948A (en) * | 2017-01-10 | 2017-05-10 | 广西大学 | Pollution-free preparing and culturing method for rice-false-smut-case thin-wall conidia |
| CN106701653A (en) * | 2017-01-10 | 2017-05-24 | 广西大学 | Pollution-reduction preparation and culture method of ustilaginoidea virens thin-wall conidia |
| CN106754424A (en) * | 2017-01-10 | 2017-05-31 | 广西大学 | The standing form and its preparation and application method of a kind of ustilaginoidea virens lab material |
| CN106867957A (en) * | 2017-01-10 | 2017-06-20 | 广西大学 | A kind of ustilaginoidea virens thin-walled of Pollution protection is conidial to prepare cultural method |
| CN110511859A (en) * | 2019-08-22 | 2019-11-29 | 广西农业职业技术学院 | A kind of equipment and working method preparing potato culture |
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Application publication date: 20160720 |