CN102061330B - Method for identifying pathogenicity of sesame stem blight and blast pathogenic bacteria - Google Patents
Method for identifying pathogenicity of sesame stem blight and blast pathogenic bacteria Download PDFInfo
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Abstract
The invention relates to a method for identifying pathogenicity of sesame stem blight and blast pathogenic bacteria. The method comprises the following steps of: (1) culturing bacterial strains for inoculation; (2) pretreating seeds, namely selecting healthy and full sesame seeds and sterilizing for later use; (3) inoculating, namely adding a sterilization culture medium into a culture bottle, laying down sterile filter paper, adding sterile water, removing bubbles, uniformly placing sesame sterile seeds along the edges of the filter paper, inoculating bacterium cakes in the center of the filter paper, and hermetically culturing in the dark; (4) investigating pathogenicity, namely dividing the pathogenic conditions into different pathogenic grades according to pathogenic degrees; and (5) evaluating the pathogenicity, wherein the pathogenic index is equal to Sigma (pathogenic grade*corresponding pathogenic bud number)/total bud number. The method is simple and convenient and practicable, the required equipment is few, the result is reliable, and the pathogenicity of different sesame stem blight pathogenic bacteria and blast pathogenic bacteria can be reflected more accurately.
Description
Technical field
The present invention relates to a kind of stalk of sesame point rot and pathogenic method of blight pathogenic bacteria identified.
Background technology
Sesame is one of China four big important oil crops, and plantation distributes extensively, and wherein the geographic sesame cultivated area in the Yellow River and Huai He River and the middle and lower reach of Yangtze River is comparatively concentrated, accounts for 70% of national sesame area.About 90 days of the breeding time of sesame belonged to the warm crop of happiness, generally was grown in for 6～August, and at this moment, many places, sesame major production areas are in high temperature and rainy season, and hot and humidly was prone to bring out stalk of sesame and selects rot and sesame blight.
Stalk of sesame point rot is claimed stem rot, charcoal rot again, and its pathogenic bacteria is Kidney bean shell ball spore (Macrophomina phaseoli (Maubl.) Ashby.), belongs to Deuteromycotina fungi (Dinakaran et al.2001; EI-Fiki at al., 2004; Saharan, 1989; Solankiet al., 2006).This pathogenic bacteria host range is wide; Can infect 500 various plants (Dhingra and the Sinclair of 75 sections; 1978); Except that sesame, its infective plant still has crudefiber crop, beans, clover class, melon, Sunflower Receptacle, tobacco, tomato, eggplant, capsicum, sugarcane, Chinese sorghum, corn, tea, coffee, coconut and banana etc.The long-term big area of stalk of sesame point rot takes place, and is one of main disease species that causes China's sesame underproduction.This disease pathogen bacterium survives the winter on the residual body of seed, soil and diseased plant with sclerotium and pycnidium.The primary infection source is main with sclerotium, and the field conidium borrows gentle the spreading of rainwater to broadcast, and repeatedly infects again.Sesame seedling stage and full-bloom stage are the easiest to be susceptible.It is 25～30 ℃ that germ grows optimum temperuture.The then seedling root browning of catching an illness seedling stage, overground part is wilted withered, close living black point on the young stem.The phase of yielding positive results catches an illness and then begins morbidity from root, and the back is spread to stem after petiole base is invaded to the stem expansion sometimes.Root is caught an illness, main root, supporting root browning, and the strip off cortex is visible to be covered with the black microsolerotium, and it is withered to cause root.Stem catches an illness and mostly occurs in the middle and lower part, just is the tawny water soaking mode, and the back expansion is very soon around one week of stem, and there is silver gray gloss at the center, and close living black small grain point on it, epidermis reach marrow down and produce a large amount of microsolerotiums, stem stalk hollow frangible (kingdom is clear etc., 2005; Pan Zhengan etc., 2008).
Sesame blight cause of disease is Fusarium oxysporum sesame specialized form Fusarium oxysporum Schl.f.sp.sesami (Zap.) Cast, and different name is F.vasinfectum Atk.var.sesami Zapr., belongs to Deuteromycotina fusarium fungi.The sesame blight is a kind of vascular bundle diseases of systemic infection, all can fall ill from seedling stage to the one-tenth strain phase, and be main but fall ill with the one-tenth strain phase.The susceptible root-rot that often causes of seedling, diseased plant is dampinged off withered.Become the flavescence wilting gradually from bottom to top of strain phase susceptible then diseased plant blade, browning is withered at last comes off, and transfers to the visible root of diseased plant, basal part of stem browning to rot, and the only half of branches and leaves wilting of the diseased plant that has is withered, in diseased plant stem stalk one side generation long strip shape brown.When moist, produce the mould layer of pink clayey (germ sporodochium and conidium) in withered sick portion.Cut open the stem inspection, visible vascular bundle just is red, along with the state of an illness increases the weight of, becomes sorrel to brown.The diseased plant capsule is less, and seed is thin and hollow, is prone to burst in advance.
Different pathogenic same a kind of pathogenic bacterias are different to the harm ability of crop, often have dominant races of this pathogenic bacteria or strong pathogenic microspecies to exist in one period or certain zone.In the evaluation of crop resource material disease resistance, breeding for disease resistance and disease control process, accurately identify the pathogenic of specific pathogen bacterium, anti-stem point rot of sesame or anti-blight breeding work are shot the arrow at the target.
At present; About identifying the method and the disunity of Kidney bean shell ball spore bacteria pathogenic; Method so that field or potted plant live body plant or in vitro tissue are object of inoculation is in the majority, like hypocotyl inoculation method (Zhu Zhendong etc., 2000), cut stem method (Pabon and Hartman; 2006), soak root method (Assigbetse et al.1994), vitro method (Prat et al.1998); These methods are mainly estimated the pathogenic of pathogenic bacteria with scab size or scab propagation rate, but the plant cultivation relates to field or greenhouse management, and the cycle is than length and be subject to environmental influence.Also having certain methods in addition is object of inoculation with the germinated seeds; Severity with seed is infected is estimated the pathogenic of pathogenic bacteria; This method is comparatively easy, and mainly on the bigger species of seed, uses at present, like Sunflower Receptacle (Manici et al.; 1995), dish is with beans (Reyes-Franco et al., 2006) etc.Stalk of sesame selects rot and the pathogenic authentication method of sesame blight generally adopts the sick garden of field natural occurrence to combine artificial inoculation (to spray spore or sclerotium suspension-s flowering period; Or in soil the inoculation sick wheat); Investigate the disease index of different sesame materials; Need during inoculation the big area field to preserve moisture etc. but this land for growing field crops becomes strain phase authentication method, and the field big area is difficult to control condition, and also is vulnerable to external environment factor such as weather and disturbs; And remove the pathogenic bacteria that also contains other multiple soil-borne diseases outside these two kinds of cause of diseases in the soil; So this one-tenth strain phase land for growing field crops inoculation identification method is not only loaded down with trivial details but also safety is not high, the cycle is long, nor identifies when possibly realize a plurality of pathogenic bacteria.Therefore grope a cover simple and easy, controlled, effectively identify that stalk of sesame select rot and the pathogenic method of withered pathogenic bacteria select withered for the control stalk of sesame and blight disease, the disease resistance of evaluation sesame planting material, cultivation disease resistance new variety, the reasonable kind layout of carrying out significant.
Summary of the invention
Technical problem to be solved by this invention provides a kind of stalk of sesame point rot and pathogenic method of blight pathogenic bacteria identified.Method is simple for this, and required equipment is less, and reliable results can reflect the pathogenic of different stalk of sesame point rot pathogenic bacterias and blight pathogenic bacteria comparatively exactly.
For solving the technical problem that the present invention proposes, the technical scheme that the present invention adopts is:
A kind of stalk of sesame point rot and pathogenic method of blight pathogenic bacteria identified is characterized in that may further comprise the steps: (remove and specify that following used vessel, filter paper etc. all pass through autoclaving and handles)
(1) inoculation is with the cultivation of bacterial strain: the potato dextrose agar containing through sterilization (is inoculated Kidney bean shell ball spore bacterium mycelia or Fusarium oxysporum sesame specialized form bacterium mycelia on the petridish of PDA (Potato dextroseagar); The sealing dark culturing is to forming bacterium colony; Then it is made the bacterium cake, subsequent use;
(2) seed pre-treatment: select healthy full sesame seed for use, it is subsequent use to sterilize;
(3) inoculation: in culturing bottle, add sterilising medium, the lay aseptic filter paper adds sterilized water; And drive bubble out of; Evenly put the sesame aseptic seed of process sterilising treatment in the step (2) then along the filter paper edge, cultivate the bacterium cake that makes, airtight dark culturing at filter paper central authorities' inoculation steps (1) again;
(4) pathogenic investigation: investigate the incidence of each chitting piece in the culturing bottle, and according to occurring degree incidence is divided into different morbidity grades, this morbidity grade numerical value is big more, and occurring degree is serious more;
(5) pathogenic evaluation: calculate the pathogenic index of each bacterial strain, the calculation formula of pathogenic index is following, and this pathogenic index numerical value shows the pathogenic strong more of this pathogenic bacteria more greatly:
Pathogenic index=∑ (morbidity grade * corresponding morbidity bud number)/total bud number.
Press such scheme; The concrete steps of described step (1) are: inoculation Kidney bean shell ball spore bacterium mycelia or Fusarium oxysporum sesame specialized form bacterium mycelia on the petridish of the PDA substratum that contains the process sterilization; Seal with the Parafilm film; 28～30 ℃ of dark culturing 5～7d in biochemical incubator then, the consisting of of said PDA substratum: contain 200g peeling yam filtrating, glucose 15g, agar 12g in every liter of substratum, the sterilization method of said PDA substratum is an autoclave sterilization.
Press such scheme, the mode of seed described in the step (2) sterilization is that 0.1% the mercuric chloride aqueous solution soaks 4～5min for seed being placed weight percent concentration, and during shaken several times.
Press such scheme, the substratum in the said step (3) is 2% agar (w/v), and the sterilization method of said substratum is an autoclave sterilization.
Press such scheme, when step (1) was used Kidney bean shell ball spore bacterium, the described inoculation culture of step (3) was 28-30 ℃ of sealing dark culturing 4～5d; When step (1) was used Fusarium oxysporum sesame specialized form bacterium, the described inoculation culture of step (3) was 28-30 ℃ of sealing dark culturing 8～10d.
Press such scheme, the concrete steps of said step (3) are: at diameter is in the culturing bottle of 60mm～70mm, adds sterilising medium; The lay aseptic filter paper; Add sterilized water, and drive bubble out of, evenly put the aseptic sesame seed of process sterilising treatment in 20～30 pieces of steps (2) then along the filter paper edge; Be that the step (1) of 0.5cm～1cm is cultivated the bacterium cake that obtains, airtight dark culturing at filter paper central authorities inoculation diameter again;
Press such scheme, carrying out step (3) before, step (3) is repeated 2 times at least.
Press such scheme, when repeated experiments was set, pathogenic index was on average got by the pathogenic index of repeatedly experiment.
Pressing such scheme, when step (1) is used Kidney bean shell ball spore bacterium, is 6 grades according to the following grade scale grade classification of will falling ill in the said step (4), and wherein infecting stem, to select the many quality in brown position of sesame bud or root of rot moistening:
0 grade: no germ is infected symptom;
1 grade: root system browning scope≤1/3;
2 grades: root system browning scope is between 1/3～2/3, i.e. 1/3＜root system browning scope≤2/3;
3 grades: root system browning scope is 2/3 to whole root system browning, i.e. root system browning scope＞2/3 is to whole root system browning;
4 grades: the whole brownings of root system, the susceptible and bud of collar portion has 1/2 and 1/2 following browning;
5 grades: the whole brownings of root system, the susceptible and bud of collar portion has 1/2 above browning, even putrefactive phenomenon occurs.
Pressing such scheme, when step (1) is used Fusarium oxysporum sesame specialized form bacterium, is 6 grades according to the following grade scale grade classification of will fall ill in the said step (4), wherein infects the many quality dryings in brown position of the sesame bud or the root of blight:
0 grade: no germ is infected symptom;
1 grade: root system browning scope≤1/3;
2 grades: root system browning scope is between 1/3～2/3, i.e. 1/3＜root system browning scope≤2/3;
3 grades: root system browning scope is 2/3 to whole root system browning, i.e. root system browning scope＞2/3 is to whole root system browning;
4 grades: the whole brownings of root system, the susceptible and bud of collar portion has 1/2 and 1/2 following browning;
5 grades: the whole brownings of root system, the susceptible and bud of collar portion has 1/2 above browning, even putrefactive phenomenon occurs.
Beneficial effect of the present invention:
A kind of stalk of sesame point rot and pathogenic method of blight pathogenic bacteria identified provided by the invention, 1, can in the laboratory, carry out fully, simple and easy to do, required equipment is less, and cost is low, good reproducibility; 2, can reflect the pathogenic of different stalk of sesame point rots and blight pathogenic bacteria, reliable results comparatively exactly; 3, help to study quickly and accurately that thereby that stalk of sesame selects rot and sesame blight pathogenic bacteria is pathogenic for formulating the Preventing Countermeasures that is fit to, carry out rational sesame variety layout, cultivating new disease resistance sesame variety technical support is provided.
Description of drawings
Fig. 1 selects the pathogenic difference on the sesame bud of rot pathogenic bacteria for stem, wherein 0,1,2,3,4,5 represents different morbidity grades respectively;
Fig. 2 is the pathogenic difference on the sesame bud of blight pathogenic bacteria, wherein 0,1,2,3,4,5 represents different morbidity grades respectively.
Embodiment
The present invention combines following examples, so that understand content of the present invention better.Experimental technique among the following embodiment like no specified otherwise, is ordinary method.Operating process is except that specifying, all room temperature is carried out on super clean bench.The brand of used biochemical incubator is a Sanyo, and used sesame material provides by sesame resource seminar of Inst. of Oil Crops, Chinese Academy of Agriculture.
Consisting of of said PDA substratum: contain 200g peeling yam filtrating, glucose 15g, agar 12g in every liter of substratum, said PDA culture medium preparation: choose peeling yam 200g, stripping and slicing; The boiling of in zero(ppm) water, seething with excitement; After potato ball is well-done, use clean filtered through gauze, the filtrating of acquisition is peeling yam filtrating, adds 15g glucose and 12g agar; Add water to 1000mL then, and get through autoclave sterilization 15～20min.
The pathogenic evaluation of the stalk of sesame point rot pathogenic bacteria of an embodiment 1:36 different sources
(1) stalk of sesame is selected the preparation of rot pathogenic bacteria mycelia: get and infect the sick strong intersection of sesame plant that stem is selected rot; Adopt and to be inoculated into after the sterilization of 0.1% (weight percentage) mercuric chloride aqueous solution after dark culturing forms bacterium colony on the PDA substratum; Colony edge picking mycelia on the PDA substratum succeeding transfer culture until the bacterium colony homogeneous; Process lures spore to cultivate then, and obtains the Kidney bean shell ball spore bacterium mycelia of unit cell culture purified, preserves.
(2) strain culturing is used in inoculation: the central authorities at the petridish (diameter is 90mm) that contains PDA (the Potato dextrose agar) substratum of sterilizing through HTHP 15min inoculate the Kidney bean shell ball spore bacterium mycelia of having kept; The Parafilm film seals; 30 ℃ of dark culturing 5d are to forming bacterium colony;
(3) seed pre-treatment: select healthy full No. 2 seeds of Hubei Province sesame for use, with the mercuric chloride (HgCl of 0.1% (weight percentage)
2) soak sterilization 4～5min in the aqueous solution, during shaken several times, use aseptic water washing then 3 times, be tiled in the petridish that is covered with aseptic filter paper, place super clean bench to dry naturally, subsequent use;
(4) inoculation: the mass concentration of preparing 2% agar (w/v) and be agar is the substratum of 20g/L; Autoclaving 20min is poured in the wide-mouth vial about 60mm (diameter) * 95mm (height) then, to the substratum height be about 10mm; Treat that agar solidifies the back and spreads one deck aseptic filter paper (about diameter 60mm) in media surface; Add the 1mL sterilized water, pave filter paper with aseptic nipper, it is subsequent use to drive bubble out of; Evenly lay out 20 pieces of aseptic seeds along the filter paper edge; Cultivating the inoculation that obtains in step (2) then uses colony edge to buy the bacterium cake of diameter as 1cm; Be laid on the filter paper central authorities of wide-mouth vial; Again the wide-mouth vial is sealed and be placed on 30 ℃ of dark culturing 5d, each stalk of sesame point rot pathogenic strains is inoculated 3 culturing bottles respectively;
(5) pathogenic investigation and statistics: investigate the incidence of chitting piece in each inoculation wide-mouth vial, and by the strain number of following grade scale statistics different onset grade, wherein infecting stem, to select the many quality in brown position of sesame bud or root of rot moistening.
0 grade: no germ is infected symptom;
1 grade: root system browning scope≤1/3;
2 grades: root system browning scope is between 1/3～2/3, i.e. 1/3＜root system browning scope≤2/3;
3 grades: root system browning scope is 2/3 to whole root system browning, i.e. root system browning scope＞2/3 is to whole root system browning;
4 grades: the whole brownings of root system, the susceptible and bud of collar portion has 1/2 and 1/2 following browning;
5 grades: the whole brownings of root system, the susceptible and bud of collar portion has 1/2 above browning, even rots.
(6) pathogenic evaluation: by following formula calculate respectively each bacterial strain pathogenic index, see table 1.The scope of pathogenic index is between 0～5, and this numerical value shows pathogenic strong more more greatly.
Pathogenic index=∑ (the morbidity grade * bud number of falling ill accordingly)/total bud number
The pathogenic index of 36 stalk of sesame point rots of table 1 pathogenic bacteria
* the former source of bacterium: derive from the sesame disease plant in seedling stage except that 10, No. 33, other all derive from sesame growth middle and later periods disease plant.
Can find out by table 1, in 36 different stalk of sesame point rot pathogenic bacteria bacterial strains in source, pathogenic the strongest be No. 31, secondly be No. 29, a little less than is No. 5.Show through repeated experiments: the pathogenic method good reproducibility of this evaluation stalk of sesame point rot pathogenic bacteria, reliable results.
The pathogenic evaluation of the sesame blight pathogenic bacteria of an embodiment 2:28 different sources
(1) preparation of sesame blight pathogenic bacteria mycelia: get the sick strong intersection of the sesame plant that infects blight; Adopt and to be inoculated into after the sterilization of 0.1% (weight percentage) mercuric chloride aqueous solution after dark culturing forms bacterium colony on the PDA substratum; Colony edge picking mycelia on the PDA substratum succeeding transfer culture until the bacterium colony homogeneous; Through luring spore to cultivate, obtain the Fusarium oxysporum sesame specialized form bacterium mycelia of unit cell culture purified then, preserve.
(2) inoculating strain is cultivated: petridish (diameter the is 90mm) central authorities containing PDA (Potato dextrose agar) substratum inoculate the Fusarium oxysporum sesame specialized form bacterium mycelia of keeping; The Parafilm film seals; 30 ℃ of dark culturing 7d cause the formation bacterium colony, and are subsequent use;
(3) seed pre-treatment: select healthy full No. 2 seeds of Hubei Province sesame for use, with the HgCl of 0.1% (weight percentage)
2(mercuric chloride) aqueous solution soaking sterilization 4～5min, during shaken several times, use aseptic water washing then 4 times, be tiled in the petridish that is covered with aseptic filter paper, it is subsequent use to place super clean bench to dry naturally;
(4) inoculation: prepare 2% agar (w/v) substratum, autoclaving 20min is poured in the wide-mouth vial about 60mm (diameter) * 95mm (height) then; To the substratum height be about 10mm; Treat that agar solidifies the back and spreads one deck aseptic filter paper (about diameter 60mm) in media surface, add the 1mL sterilized water, pave filter paper with aseptic nipper; It is subsequent use to drive bubble out of; Evenly lay out 20 pieces of aseptic seeds along the filter paper edge, cultivate the inoculation obtain in step (2) then and buy the bacterium cake of diameter 1cm, be layered on wide-necked bottle filter paper central authorities with colony edge; Again the wide-mouth vial is sealed and be placed on 30 ℃ of dark culturing 10d, each sesame blight pathogenic bacteria bacterial strain is inoculated 3 culturing bottles respectively;
(5) pathogenic investigation and statistics: investigate the incidence of chitting piece in each inoculation wide-mouth vial, and add up the strain number of different onset grade, wherein infect the many quality dryings in brown position of the sesame bud or the root of blight by following grade scale;
0 grade: no germ is infected symptom;
1 grade: root system browning scope≤1/3;
2 grades: root system browning scope is between 1/3～2/3, i.e. 1/3＜root system browning scope≤2/3;
3 grades: root system browning scope is 2/3 to whole root system browning, i.e. root system browning scope＞2/3 is to whole root system browning;
4 grades: the whole brownings of root system, the susceptible and bud of collar portion has 1/2 and 1/2 following browning;
5 grades: the whole brownings of root system, the susceptible and bud of collar portion has 1/2 above browning, even rots;
(6) pathogenic evaluation: by following formula calculate respectively different each bacterial strain in source pathogenic index, see table 2.The scope of pathogenic index is between 0～5, and the big more explanation of this numerical value is pathogenic strong more:
Pathogenic index=∑ (the morbidity grade * bud number of falling ill accordingly)/total bud number
The pathogenic index of 36 sesame blights of table 2 pathogenic bacteria
* the former source of bacterium: all derive from sesame growth middle and later periods disease plant.
Can find out by table 2, in 28 different sesame blight pathogenic bacterias in source pathogenic the strongest be No. 18, secondly be No. 21, the most weak is No. 20.Show through repeated experiments: the pathogenic method good reproducibility of this evaluation sesame blight pathogenic bacteria, reliable results.
Claims (8)
1. identify stalk of sesame point rot and the pathogenic method of blight pathogenic bacteria for one kind, it is characterized in that: may further comprise the steps:
(1) inoculation is with the cultivation of bacterial strain: inoculation Kidney bean shell ball spore bacterium mycelia or Fusarium oxysporum sesame specialized form bacterium mycelia on containing through the petridish of the PDA substratum of sterilization, seal dark culturing to forming bacterium colony, and then it is made the bacterium cake, subsequent use;
(2) seed pre-treatment: select healthy full sesame seed for use, it is subsequent use to sterilize;
(3) inoculation: in culturing bottle, add sterilising medium; The lay aseptic filter paper; Add sterilized water, and drive bubble out of, evenly put the sesame aseptic seed of process sterilising treatment in the step (2) then along the filter paper edge; Cultivate the bacterium cake that makes, airtight dark culturing at filter paper central authorities' inoculation steps (1) again;
(4) pathogenic investigation: investigate the incidence of each chitting piece in the culturing bottle, and incidence is divided into different morbidity grades according to occurring degree:
Wherein, when using Kidney bean shell ball spore bacterium, it is 6 grades according to the following grade scale grade classification of will falling ill, and wherein infecting stem, to select the many quality in brown position of sesame bud or root of rot moistening:
0 grade: no germ is infected symptom;
1 grade: root system browning scope≤1/3;
2 grades: root system browning scope is between 1/3～2/3;
3 grades: root system browning scope is 2/3 to whole root system browning;
4 grades: the whole brownings of root system, the susceptible and bud of collar portion has 1/2 and 1/2 following browning;
5 grades: the whole brownings of root system, the susceptible and bud of collar portion has 1/2 above browning, even putrefactive phenomenon occurs;
When using Fusarium oxysporum sesame specialized form bacterium, it is 6 grades according to the following grade scale grade classification of will fall ill, wherein infects the many quality dryings in brown position of the sesame bud or the root of blight:
0 grade: no germ is infected symptom;
1 grade: root system browning scope≤1/3;
2 grades: root system browning scope is between 1/3～2/3;
3 grades: root system browning scope is 2/3 to whole root system browning;
4 grades: the whole brownings of root system, the susceptible and bud of collar portion has 1/2 and 1/2 following browning;
5 grades: the whole brownings of root system, the susceptible and bud of collar portion has 1/2 above browning, even putrefactive phenomenon occurs;
This morbidity grade numerical value is big more, and occurring degree is serious more;
(5) pathogenic evaluation: calculate the pathogenic index of each bacterial strain, the calculation formula of pathogenic index is following, and this pathogenic index numerical value shows the pathogenic strong more of this pathogenic bacteria more greatly:
Pathogenic index=∑ (morbidity grade * corresponding morbidity bud number)/total bud number.
2. evaluation stalk of sesame point rot according to claim 1 and the pathogenic method of blight pathogenic bacteria; It is characterized in that: the concrete steps of said step (1) are: inoculation Kidney bean shell ball spore bacterium mycelia or Fusarium oxysporum sesame specialized form bacterium mycelia on the petridish of the PDA substratum that contains the process sterilization; Seal with the Parafilm film; 28～30 ℃ of dark culturing 5～7d in biochemical incubator then; Consisting of of said PDA substratum: contain 200g peeling yam filtrating, glucose 15 g, agar 12 g in every liter of substratum, the sterilization method of said PDA substratum is an autoclave sterilization.
3. evaluation stalk of sesame point rot according to claim 1 and the pathogenic method of blight pathogenic bacteria; It is characterized in that: the mode of seed described in the step (2) sterilization is that 0.1% the mercuric chloride aqueous solution soaks 4～5min for seed being placed weight percent concentration, and during shaken several times.
4. evaluation stalk of sesame point rot according to claim 1 and the pathogenic method of blight pathogenic bacteria is characterized in that: the substratum in the said step (3) is 2% agar (w/v), and the sterilization method of said substratum is an autoclave sterilization.
5. evaluation stalk of sesame point rot according to claim 1 and the pathogenic method of blight pathogenic bacteria is characterized in that: when step (1) was used Kidney bean shell ball spore bacterium, the described inoculation culture of step (3) was 28～30 ℃ of sealing dark culturing 4～5d; When step (1) was used Fusarium oxysporum sesame specialized form bacterium, the described inoculation culture of step (3) was 28～30 ℃ of sealing dark culturing 8～10d.
6. evaluation stalk of sesame point rot according to claim 1 and the pathogenic method of blight pathogenic bacteria is characterized in that: the concrete steps of said step (3) are: at diameter is in the culturing bottle of 60mm～70mm, adds sterilising medium, lay
Aseptic filter paper; Add sterilized water, and drive bubble out of, evenly put the aseptic sesame seed of process sterilising treatment in 20～30 pieces of steps (2) then along the filter paper edge; Be that the step (1) of 0.5cm～1cm is cultivated the bacterium cake that obtains, airtight dark culturing at filter paper central authorities inoculation diameter again.
7. evaluation stalk of sesame point rot according to claim 1 and the pathogenic method of blight pathogenic bacteria is characterized in that: carrying out step (4) before, step (3) is repeated 2 times at least.
8. evaluation stalk of sesame point rot according to claim 7 and the pathogenic method of blight pathogenic bacteria is characterized in that:
Said pathogenic index is on average got by the pathogenic index of each experiment.
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| CN104160846B (en) * | 2014-07-15 | 2016-02-17 | 武汉市蔬菜科学研究所 | A kind of sprout period testifying method of cowpea anti-blight |
| CN109061057A (en) * | 2018-07-12 | 2018-12-21 | 安徽省农业科学院作物研究所 | A kind of non-destructive monitoring method of crops seedling stage root pathogens colonization status |
| CN112119895A (en) * | 2020-09-03 | 2020-12-25 | 河南省农业科学院烟草研究所 | Method for rapidly identifying pathogenicity of fusarium graminearum |
| CN114958898B (en) * | 2022-05-20 | 2023-11-14 | 中国农业科学院郑州果树研究所 | Establishment method and application of PEG-mediated genetic transformation system of aschersonia phaseoli |
| CN114921349B (en) * | 2022-06-21 | 2023-11-24 | 河南省农业科学院植物保护研究所 | Eichhornia crassipes strain and application thereof in biological control |
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| Publication number | Priority date | Publication date | Assignee | Title |
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| WO2002096204A1 (en) * | 2001-05-30 | 2002-12-05 | Her Majesty The Queen In Right Of Canada, As Represented By The Minister Of Agriculture And Agri-Food | Fungal isolates and biological control compositions for the control of weeds |
| CN101323875A (en) * | 2008-07-24 | 2008-12-17 | 江苏省农业科学院 | Wheat fusarium stalk rot seedlings inoculation identification method |
| WO2009027539A2 (en) * | 2007-08-31 | 2009-03-05 | Basf Plant Science Gmbh | Method for producing a transgenic plant cell, a plant or a part thereof with increased resistance to plant disease |
| CN101508959A (en) * | 2008-12-23 | 2009-08-19 | 西南大学 | Water hyacinth fungal biocontrol agent-eichhornia crassipes alternaria alternata |
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- 2010-11-30 CN CN201010565405A patent/CN102061330B/en not_active Expired - Fee Related
Patent Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2002096204A1 (en) * | 2001-05-30 | 2002-12-05 | Her Majesty The Queen In Right Of Canada, As Represented By The Minister Of Agriculture And Agri-Food | Fungal isolates and biological control compositions for the control of weeds |
| WO2009027539A2 (en) * | 2007-08-31 | 2009-03-05 | Basf Plant Science Gmbh | Method for producing a transgenic plant cell, a plant or a part thereof with increased resistance to plant disease |
| CN101323875A (en) * | 2008-07-24 | 2008-12-17 | 江苏省农业科学院 | Wheat fusarium stalk rot seedlings inoculation identification method |
| CN101508959A (en) * | 2008-12-23 | 2009-08-19 | 西南大学 | Water hyacinth fungal biocontrol agent-eichhornia crassipes alternaria alternata |
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| 胡小文等.播期对春箭筈豌豆种子质量的影响.《生态学报》.2004,第24卷(第03期),409-413. * |
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| CN102061330A (en) | 2011-05-18 |
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