CN105779373A - Ribose culture medium applicable to ustilaginoidea virens spore production and application method of ribose culture medium - Google Patents
Ribose culture medium applicable to ustilaginoidea virens spore production and application method of ribose culture medium Download PDFInfo
- Publication number
- CN105779373A CN105779373A CN201610345507.XA CN201610345507A CN105779373A CN 105779373 A CN105779373 A CN 105779373A CN 201610345507 A CN201610345507 A CN 201610345507A CN 105779373 A CN105779373 A CN 105779373A
- Authority
- CN
- China
- Prior art keywords
- ribose
- culture medium
- spore
- culture
- ustilaginoidea virens
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Pending
Links
Classifications
-
- 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
Landscapes
- Zoology (AREA)
- Life Sciences & Earth Sciences (AREA)
- Engineering & Computer Science (AREA)
- Bioinformatics & Cheminformatics (AREA)
- Health & Medical Sciences (AREA)
- Organic Chemistry (AREA)
- Genetics & Genomics (AREA)
- Chemical & Material Sciences (AREA)
- Wood Science & Technology (AREA)
- Biomedical Technology (AREA)
- Microbiology (AREA)
- Biochemistry (AREA)
- General Engineering & Computer Science (AREA)
- General Health & Medical Sciences (AREA)
- Biotechnology (AREA)
- Micro-Organisms Or Cultivation Processes Thereof (AREA)
Abstract
The invention discloses a ribose culture medium applicable to ustilaginoidea virens spore production and an application method of the ribose culture medium. Ribose which is not sterilized at high temperature is adopted to prepare a potato ribose culture medium as a spore production culture medium for culturing and preparing ustilaginoidea virens thin-wall conidia. The preparation and application method of the culture medium comprises the following operation steps: 1) preparing a basic culture medium; 2) filtering and sterilizing ribose; 3) preparing a ribose culture medium plate; 4) planting ustilaginoidea virens; 5) performing constant-temperature culture; and 6) collecting a new generation of spores. The ribose culture medium has the characteristics and advantages that the time for obtaining a great amount of spores can be shortened, the continuous spore production time is long, operation techniques are simple, convenient and rapid, no oscillation culture equipment is needed, a relatively great number of spores of the new generation can be produced, and a spore liquid is relatively pure.
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 ribose culture medium 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 ribose culture medium and the application process thereof of spore, utilize the ribose of non high temperature sterilizing to make Rhizoma Solani tuber osi ribose culture 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 ribose culture medium and the application process thereof of spore, and operating procedure is as follows:
1. prepare basal medium
In parts by weight: Rhizoma Solani tuber osi 100g, agar 20g, water 900ml, prepare basal medium, every bottled 90ml, standby after conventional high temperature sterilizing.
2. the filtration sterilization of ribose
In parts by weight: ribose 10g, water 100ml, with water, ribose is made into ribose solution, aseptically with biofilter, this ribose solution is filtered sterilizing, standby.
3. prepare ribose culture medium flat plate
Basal medium step 1 prepared adds heat fusing, water-bath balances to 60 DEG C, the ribose solution of step 2 filtration sterilization is balanced in a water bath to 60 DEG C simultaneously, then the two is simultaneously transferred in superclean bench gnotobasis, taking 10ml ribose solution and proceed in 1 bottle of basal medium, the culture dish that falls after shake mix homogeneously becomes the culture medium flat plate containing ribose；The component proportion of this culture plate is: Rhizoma Solani tuber osi 100g, ribose 10g, agar 20g, water 1000ml.
4. implant ustilaginoidea virens
Take out with the standby ustilaginoidea virens spore liquid deposited of spore form, by this ready culture plate of spore liquid implantation step 3 under aseptic condition, with T-shaped glass rod, spore liquid is spread evenly across in platen surface.
5. constant temperature culture
The culture plate of step 4 end of operation is proceeded in incubator, 28 DEG C of constant temperature culture.
6. the collection of a new generation's spore
After step 5 operation implements cultivation 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) obtain a large amount of a new generations 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.
2) operating technology is simple and easy convenient, it is not necessary to shaken cultivation equipment；
3) a new generation's 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 is the culture medium and the application process thereof that utilize the present invention, cultivates the minute colony that ustilaginoidea virens was formed after 5 days, and bacterium colony is formed a large amount of a new generations thin-walled conidium.
Fig. 2 is the culture medium and the application process thereof that utilize the present invention, cultivates ustilaginoidea virens and still maintains minute colony state after 20 days, and still keeps a large amount of a new generations thin-walled conidium 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, 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.
Inventor's previous work finds, adopts the Rhizoma Solani tuber osi ribose culture medium being carbon nutrition with ribose prepared by conventional method, and the growth promoter of ustilaginoidea virens is inhibited, once thinks that ribose can suppress the growth promoter of ustilaginoidea virens by mistake.But nearest work finds, ribose itself does not suppress the growth promoter of ustilaginoidea virens, is that ustilaginoidea virens is had inhibitory action by the high-temperature process product of ribose；Research process has been surprisingly found that, at ribose without (the Rhizoma Solani tuber osi ribose culture medium that the present invention will utilize made by the ribose of filtration sterilization in the Rhizoma Solani tuber osi ribose culture medium of high temperature sterilize (filtration sterilization), referred to as " ribose culture medium "), ustilaginoidea virens on the contrary can over a long time in continue to form the new spore of offspring.In real work, after this ribose culture medium is cultivated 5 days, culture plate can present the minute colony that can produce a large amount of a new generations spore, as it is shown in figure 1, cultivate backward, ustilaginoidea virens bacterium colony seldom grows non-sporogenous hyphae again, until after cultivating 20 days, remain in that the petite of big volume production spore state, as in figure 2 it is shown, and now remain to collect substantial amounts of a new generation spore.Visible, ribose culture medium is the product spore culture medium that ustilaginoidea virens is ideal, adopt ribose medium preparing spore, can so that the product spore of ustilaginoidea virens significantly extends period, thus ensureing within one period, the spore output that preparation obtains is relatively stable, and the preparation work of spore is not easy to disconnect with other work.Simultaneously as in ribose culture medium, petite does not have the phenomenon of obvious non-sporogenous hyphae growth promoter, and in the spore liquid of new generation of collection, mycelia fragment is less, and spore is more pure.
The preparation of the ribose culture medium of the present invention needs 2 kinds of sterilization technology synthesis, and basal medium (potato agar) adopts conventional high temperature autoclave techniques；Form pyrene sugar (ribose solution) and cannot pass through high-temperature process, filtration sterilization can only be taked；Product after these 2 kinds of technology sterilizings is mixed under non high temperature state, just can become effective and produce spore culture medium.
After two pieces of components (potato agar component, ribose solution component) are mixed, the valid density of all the components all declines, therefore, the concentration of these two pieces of components before mixing, necessarily be greater than the concentration of component of last culture medium (Rhizoma Solani tuber osi 100g, ribose 10g, agar 20g, water 1000ml).During actual design, the contribution calcutation that can comply with two pieces of components goes out the concentration before two pieces of components mix.
The mixed proportion of these two pieces of components can be any ratio, but needs technically to consider that Rhizoma Solani tuber osi and agar should not make higher concentration；And ribose solution its viscosity when higher concentration increases, it is unfavorable for disinfecting action.Compare after tested, with volume ratio: potato agar component: ribose solution component=9: 1, for mixed proportion conveniently.By this mixed proportion, the proportioning of component potato agar should be: Rhizoma Solani tuber osi 100g, agar 20g, water 900ml；And the proportioning of ribose solution should be: ribose 10g, water 100ml.
In real work, become solid after cooling down due to the agar sterilizing of potato agar component, it is impossible to arbitrarily measure, so that quantitatively bottle before sterilization.Every bottle of quantitative potato agar loading 90ml, adds the ribose solution of 10ml during use, the operation of such hybrid mode is relatively simple convenient.
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, desirable 10～100 μ l of every ware flat board 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 ribose culture medium 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. prepare basal medium
In parts by weight: Rhizoma Solani tuber osi 100g, agar 20g, water 900ml, prepare basal medium, every bottled 90ml, standby after conventional high temperature sterilizing.
2. the filtration sterilization of ribose
In parts by weight: xylose 10g, water 100ml, with water, xylose is made into xylose solution, aseptically with biofilter, this xylose solution is filtered sterilizing, standby.
3. prepare ribose culture medium flat plate
Basal medium step 1 prepared adds heat fusing, water-bath balances to 60 DEG C, the ribose solution of step 2 filtration sterilization is balanced in a water bath to 60 DEG C simultaneously, then the two is simultaneously transferred in superclean bench gnotobasis, taking 10ml ribose solution and proceed in 1 bottle of basal medium, the culture dish (diameter is 9cm) that falls after shake mix homogeneously becomes the culture medium flat plate containing ribose；The component proportion of this culture plate is: Rhizoma Solani tuber osi 100g, ribose 10g, agar 20g, water 1000ml.
4. implant ustilaginoidea virens: take out with the spore liquid of the standby ustilaginoidea virens bacterial strain Uv-108 deposited of spore form, by this ready culture plate of spore liquid implantation step 3 under aseptic condition, spore liquid be spread evenly across in platen surface with T-shaped glass rod.
5. constant temperature culture: the culture plate of step 4 end of operation is proceeded in incubator, 28 DEG C of constant temperature culture.
6. the collection of a new generation's spore: after step 5 operation implements cultivation 5 days, bacterial strain Uv-108 can form a large amount of a new generations thin-walled conidium on culture plate；Culture plate is forwarded on superclean bench, scrub a flat-plate bacterial colony with sterilized water, 1.0 × 10 can be obtained6The new spore liquid 120ml of individual spore/ml.
Embodiment 2
Adopting a kind of suitable ustilaginoidea virens of the present invention to produce ribose culture medium 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 6 operation, and the bacterial strain that simply step 4 is implanted is Uv-110.After implementing step 6 operation, scrub a flat-plate bacterial colony with sterilized water, 1.0 × 10 can be obtained6The new spore liquid 106ml of individual spore/ml.
Embodiment 3
Adopting a kind of suitable ustilaginoidea virens of the present invention to produce ribose culture medium 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 6 operation, and the bacterial strain that simply step 4 is implanted is Uv-111.After implementing step 6 operation, scrub a flat-plate bacterial colony with sterilized water, 1.0 × 10 can be obtained6The new spore liquid 133ml of individual spore/ml.
Claims (1)
1. a suitable ustilaginoidea virens produces ribose culture medium and the application process thereof of spore, it is characterised in that utilize the ribose of non high temperature sterilizing to make Rhizoma Solani tuber osi ribose culture 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) prepare basal medium: in parts by weight: Rhizoma Solani tuber osi 100g, agar 20g, water 900ml, prepare basal medium, every bottled 90ml, standby after conventional high temperature sterilizing；
2) filtration sterilization of ribose: in parts by weight: ribose 10g, water 100ml, is made into ribose solution with water by ribose, aseptically with biofilter, this ribose solution is filtered sterilizing, standby；
3) ribose culture medium flat plate is prepared: by step 1) basal medium prepared adds heat fusing, water-bath balances to 60 DEG C, simultaneously by step 2) ribose solution of filtration sterilization balances in a water bath to 60 DEG C, then the two is simultaneously transferred in superclean bench gnotobasis, taking 10ml ribose solution and proceed in 1 bottle of basal medium, the culture dish that falls after shake mix homogeneously becomes the culture medium flat plate containing ribose；The component proportion of this culture plate is: Rhizoma Solani tuber osi 100g, ribose 10g, agar 20g, water 1000ml；
4) ustilaginoidea virens is implanted: take out with the standby ustilaginoidea virens spore liquid deposited of spore form, by this spore liquid implantation step 3 under aseptic condition) ready culture plate, with T-shaped glass rod, spore liquid is spread evenly across in platen surface；
5) constant temperature culture: by step 4) culture plate of end of operation proceeds in incubator, 28 DEG C of constant temperature culture；
6) collection of a new generation's spore: step 5) operation implements to cultivate 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 |
|---|---|---|---|
| CN201610345507.XA CN105779373A (en) | 2016-05-24 | 2016-05-24 | Ribose culture medium applicable to ustilaginoidea virens spore production and application method of ribose culture medium |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201610345507.XA CN105779373A (en) | 2016-05-24 | 2016-05-24 | Ribose culture medium applicable to ustilaginoidea virens spore production and application method of ribose culture medium |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN105779373A true CN105779373A (en) | 2016-07-20 |
Family
ID=56380408
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201610345507.XA Pending CN105779373A (en) | 2016-05-24 | 2016-05-24 | Ribose culture medium applicable to ustilaginoidea virens spore production and application method of ribose culture medium |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN105779373A (en) |
Cited By (3)
| 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 |
| 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 |
| CN108060088A (en) * | 2018-01-11 | 2018-05-22 | 江西省农业科学院植物保护研究所 | A kind of culture medium for being used for Pyricularia oryzae culture, producing spore and preparation method thereof |
Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN105541927A (en) * | 2016-02-03 | 2016-05-04 | 广西大学 | Ribose-derived Ustilaginoidea virens inhibitor |
-
2016
- 2016-05-24 CN CN201610345507.XA patent/CN105779373A/en active Pending
Patent Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN105541927A (en) * | 2016-02-03 | 2016-05-04 | 广西大学 | Ribose-derived Ustilaginoidea virens inhibitor |
Non-Patent Citations (2)
| Title |
|---|
| 张君成 等: "糖对稻曲病菌薄壁分生孢子萌发的影响", 《植物病理学报》 * |
| 蒲相君，等: "稻曲病菌薄壁分生孢子产孢平板培养基的筛选", 《南方农业学报》 * |
Cited By (3)
| 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 |
| 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 |
| CN108060088A (en) * | 2018-01-11 | 2018-05-22 | 江西省农业科学院植物保护研究所 | A kind of culture medium for being used for Pyricularia oryzae culture, producing spore and preparation method thereof |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN101979518B (en) | Method for preparing pseudorabies virus | |
| KR101950245B1 (en) | Blue-Green Alga Spirulina Animal Cell Culture Solution, Method for Preparing the Same, and Method for Culturing Cells Using the Same | |
| Ammar | Cultivation of Microalgae Chlorella vulgaris in Airlift photobioreactor for Biomass Production using commercial NPK Nutrients | |
| CN105779372A (en) | Potato culture medium suitable for ustilaginoidea virens spore production and application thereof | |
| CN105779373A (en) | Ribose culture medium applicable to ustilaginoidea virens spore production and application method of ribose culture medium | |
| CN105462884A (en) | Mycoplasma anatis culture medium and separation and purification method for mycoplasma anatis | |
| CN102205116A (en) | Method for producing vaccine by virtue of culturing animal cells | |
| CN109913404A (en) | The preparation method of infections chicken cloacal bursa virus live vaccine | |
| CN104027798B (en) | Method for culturing and producing PVC 2 antigen through whole suspension cells | |
| CN112080435A (en) | Purple lilac spore strain and application thereof in degrading chicken feather | |
| CN109082396A (en) | Bacterium and its application in control of plant disease is quenched in a kind of DSF colony induction signaling molecule | |
| CN102286378B (en) | Composite probiotics for inhabiting aspergillus flavus growth and degrading aflatoxin and application thereof | |
| CN104004720B (en) | A kind of large scale and high density produces the method for porcine circovirus 2 type antigen | |
| CN101485300B (en) | Method for cultivating Acrobeloides nanus | |
| CN106834159B (en) | One plant of HS233 bacterial strain and its application in resistance to cadmium and/or the effective cadmium content of reduction | |
| CN108841748A (en) | Sinorhizobium nitrogen-fixing bacteria strain H6 and its application | |
| CN104604522A (en) | Method for producing cordyceps militaris | |
| CN105779374A (en) | Xylose medium suitable for ustilaginoidea virens spore production and application method thereof | |
| CN102886043A (en) | Binary inactivated vaccine against Japanese encephalitis virus and porcine parvovirus and preparation method thereof | |
| CN106635865B (en) | A kind of culture medium and its preparation method and application isolated and purified for mycoplasma bovirhinis | |
| CN105532581B (en) | A method of insect fungal component is studied in vitro through ovum vertical transmission | |
| CN107699531A (en) | A kind of hippophae plant Frankia isolated culture method | |
| CN104740627B (en) | A kind of large-scale method for producing of pseudo- mad dog attenuated live vaccines for animals | |
| CN104531548A (en) | Androstenedione substrate-tolerant mutant strain and mutation breeding method thereof | |
| CN106237324A (en) | A kind of method using full suspension technology to produce transmissible gastroenteritis of swine vaccine |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| C06 | Publication | ||
| PB01 | Publication | ||
| C10 | Entry into substantive examination | ||
| SE01 | Entry into force of request for substantive examination |