CN112243824A - Peanut fruit rot inoculation method - Google Patents
Peanut fruit rot inoculation method Download PDFInfo
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- CN112243824A CN112243824A CN202011140672.4A CN202011140672A CN112243824A CN 112243824 A CN112243824 A CN 112243824A CN 202011140672 A CN202011140672 A CN 202011140672A CN 112243824 A CN112243824 A CN 112243824A
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- peanut
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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
- A01G22/00—Cultivation of specific crops or plants not otherwise provided for
- A01G22/40—Fabaceae, e.g. beans or peas
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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
- A01G24/00—Growth substrates; Culture media; Apparatus or methods therefor
- A01G24/10—Growth substrates; Culture media; Apparatus or methods therefor based on or containing inorganic material
- A01G24/12—Growth substrates; Culture media; Apparatus or methods therefor based on or containing inorganic material containing soil minerals
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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
- A01G24/00—Growth substrates; Culture media; Apparatus or methods therefor
- A01G24/20—Growth substrates; Culture media; Apparatus or methods therefor based on or containing natural organic material
- A01G24/22—Growth substrates; Culture media; Apparatus or methods therefor based on or containing natural organic material containing plant material
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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
- A01G7/00—Botany in general
- A01G7/06—Treatment of growing trees or plants, e.g. for preventing decay of wood, for tingeing flowers or wood, for prolonging the life of plants
Abstract
The invention discloses a peanut rot inoculation method, and relates to the technical field of peanut rot research. The peanut rot inoculation method comprises the following steps: s1: selecting healthy and plump peanut seeds, soaking the seeds in 75% alcohol for 0.5-1 min, then soaking the seeds in 10% sodium hypochlorite for 15min, sterilizing, washing with water for 3 times, and then sowing the seeds; s2: culturing pathogenic bacteria of the peanut rot disease by using a culture medium, and inoculating the hypha of the pathogenic bacteria of the peanut rot disease to the soaked and sterilized grains after the hypha grows out; s3: and (4) after the grains are overgrown with hypha, covering the roots of the peanut samples cultured in the step S1 with a mixture of the fungus grains, the sterile sand and the sterile water in a mass ratio of 1:1:2, and inoculating the peanut rot pathogenic bacteria. According to the method, grains are used as a carrier of pathogenic bacteria, and a mixture of bacteria grains, sterile sandy soil and sterile water in a mass ratio of 1:1:2 is matched to perform covering inoculation on the root of a peanut sample, so that the morbidity of the inoculated peanut rot is obviously improved.
Description
Technical Field
The invention relates to the technical field of peanut rot research, in particular to the technical field of a peanut rot inoculation method.
Background
Peanut rot, also known as rotten fruit disease, is a worldwide disease that mainly damages the pods of peanuts, causing the pods to rot. Peanut rot occurs in Shandong, Hebei, Jilin, Henan and the like in China, the damage is serious, and particularly, continuous cropping plots have the trend of increasing weight year by year. The yield loss of the general disease field is up to 15 percent, and even the grains are not harvested in the serious disease field.
At present, no disease-resistant variety exists for the peanut rot, so that the method is of great importance for identifying the resistance of the existing resources. The artificial high-efficiency inoculation of the peanut rot is an important basis for seed culture and disease-resistant mechanism research of the peanut rot, and is a key for realizing high-efficiency prevention and control of the peanut rot.
However, in the prior art, the inoculation success rate of the peanut rot is low, and the study of the peanut rot is seriously influenced. How to solve the technical problems is a technical problem to be solved in the technical field of peanut rot research at present.
Disclosure of Invention
The invention provides a peanut rot inoculation method aiming at the technical problems.
The technical scheme adopted by the invention is as follows: the peanut rot inoculation method comprises the following steps:
s1: selecting healthy and plump peanut seeds, soaking the seeds in 75% alcohol for 0.5-1 min, then soaking the seeds in 10% sodium hypochlorite for 15min, sterilizing, washing with water for 3 times, and then sowing the seeds;
s2: culturing pathogenic bacteria of the peanut rot disease by using a culture medium, and inoculating the hypha of the pathogenic bacteria of the peanut rot disease to the soaked and sterilized grains after the hypha grows out;
s3: and (3) after the grains are overgrown with hypha, taking a mixture of bacteria grains, sterile sand and sterile water in a mass ratio of 1:1:2, covering the roots of the peanut samples cultured in the step S1, inoculating the peanut rot pathogenic bacteria, covering a layer of sterile sand on the roots, culturing until the peanuts are in a full fruit ripening period, and thus obtaining the peanut rot-resistant peanut.
On the basis of the scheme, the grains are oat grains.
On the basis of the scheme, the pathogenic bacteria of the peanut rot disease are pythium colonization.
On the basis of the scheme, the peanut seeds are sown and cultured indoors, the indoor temperature is controlled to be 28-30 ℃, and the indoor humidity is controlled to be 85%; alternating 14h/10h light and dark.
On the basis of the scheme, the inoculation time of the pathogenic bacteria of the peanut rot disease is from the growth period of the peanut to the injection period or the pod bearing period or between the injection period and the pod bearing period.
On the basis of the scheme, after the root of the peanut sample is inoculated with the pathogenic bacteria of peanut rot, the peanut sample is suitable for watering, and the water content of the mixture of the grains and the sandy soil is controlled to be more than 30 percent and less than 60 percent.
To be noted:
1. selecting the oat grains after swelling sterilization, wherein swelling is to ensure that the oat grains are easy to parasitize by a pythium strain after absorbing water and becoming soft; the sterilization is to eliminate interference of other strains during inoculation.
2. The mass ratio of the bacteria oat grains to the sterile sandy soil to the sterile water is 1:1:2, and the main reason is that the oat grains and the sandy soil are kept moist, the water is not particularly much, the water is much, the oat grains are easy to rot, and the oat grains are too dry to be beneficial to parasitizing the bacteria on the oat grains.
3. The peanut seeds are sowed and cultured indoors, the optimal temperature and humidity for peanut growth can be controlled, and reasonable illumination is provided.
The invention has the beneficial effects that:
1. according to the method, the oat grains are used as a carrier of pathogenic bacteria, and a mixture of the bacterial grains, the sterile sandy soil and the sterile water in a mass ratio of 1:1:2 is matched to perform coverage inoculation on peanut root stocks, so that the morbidity of peanut rot inoculation is obviously improved;
2. according to the method, inoculation is carried out in the peanut pod setting period, the incidence rate of peanut rot inoculation is as high as 80%, technical support can be provided for effective resistance evaluation of peanut resources, and a basis can be provided for disease-resistant breeding;
3. in addition, a large amount of zoospores generated by the pythium aggregatum are utilized for inoculation, the zoospores are greatly influenced by the environment and are difficult to be applied to field large-batch inoculation, and meanwhile, the problem of the inoculation by utilizing the pythium aggregatum mycelium blocks also exists; and the oat grains are used as a carrier for inoculation, the cost is low, only a mixture of the bacterium-carrying oat grains and sterile sandy soil needs to be scattered in the field, and watering is matched, so that the method is strong in operability, simple and convenient, and suitable for large-scale popularization.
Drawings
FIG. 1 is a graph showing the result of inoculation of Pythium clusters in different peanut growth periods.
Detailed Description
The technical solutions in the embodiments of the present invention will be clearly and completely described below with reference to the drawings in the embodiments of the present invention, and it is obvious that the described embodiments are only a part of the embodiments of the present invention, and not all of the embodiments. All other embodiments, which can be derived by a person skilled in the art from the embodiments given herein without making any creative effort, shall fall within the protection scope of the present invention.
The method is characterized in that at present, the artificial inoculation of the peanut rot disease is mainly to directly inoculate the mycelium block and the zoospore of the pathogenic bacteria of the peanut rot disease, namely, the mycelium block and the zoospore of the pathogenic bacteria of the peanut rot disease are directly inoculated to the root of the peanut. However, the inoculation mode causes low success rate of peanut rot inoculation, the incidence rate of the peanut rot is generally within 20%, and the inoculation failure is easily caused.
In order to find out the reason that the artificial inoculation rate of the peanut rot is low at present, the inventor finds out through research that the reason that the inoculation success rate of the peanut rot is low is as follows: in the prior art, the mycelium blocks and zoospores are mainly used for inoculation, because the mycelium blocks and the spores are greatly influenced by the environment, the mycelium blocks are easy to dry off, the zoospores are easy to hibernate and lose the infection activity, and if hosts cannot be quickly found in soil, the hosts cannot be colonized.
From the reasons, in order to solve the phenomenon that the inoculation success rate of the peanut rot is low, a carrier capable of continuously providing nutrient substances can be provided for pathogenic bacteria of the peanut rot; meanwhile, by using the carrier, the pathogenic bacteria can continue to absorb the nutrition provided by the carrier before peanut pods are not found, so that the survival time of hyphae in soil can be prolonged until the pathogenic bacteria find a proper host for colonization, and the success rate of inoculation is improved.
The inventor selects grains as carriers through continuous screening, preferably selects oat grains containing a large amount of nutrient substances suitable for hypha growth, can allow pathogenic bacteria to grow well, is convenient for inoculation operation and has lower cost.
In order to better study the effect of oat grains as a carrier on the inoculation success rate of peanut rot, the following experiment is carried out.
The experimental procedures used in the following examples are all conventional procedures unless otherwise specified.
Materials, reagents and the like used in the following examples are commercially available unless otherwise specified.
Example one, the onset of peanut rot in different stages of peanut inoculation
The experiment was divided into 4 groups, namely: seedling stage, flowering and seeding stage, pod bearing stage and full fruit mature stage; meanwhile, each group is provided with 3 parallel experiments;
the experimental conditions are as follows: indoor temperature is controlled to be 28-30 ℃ and humidity is controlled to be 85%; performing illumination for 14h and dark culture for 10h every day;
the pathogenic bacteria of the peanut rot disease are as follows: pythium ultimum (the pathogenic bacteria of the peanut rot disease are various, wherein the pythium ultimum is one of important bacteria causing the peanut rot disease and influencing the peanut yield, and other bacteria can be inoculated);
carrier: oat grains.
The experimental procedure was as follows:
s1: selecting healthy and plump peanut seeds, soaking the seeds in 75% alcohol for 0.5-1 min, then soaking the seeds in 10% sodium hypochlorite for 15min, sterilizing and washing the seeds for 3 times, and then dividing the seeds into 4 groups, namely: seedling stage, flowering and seeding stage, pod bearing stage and full fruit mature stage, and simultaneously, 3 seeds are arranged in each group for parallel sowing;
s2: culturing the pythium aggregatum by using a V8 culture medium, and inoculating the pythium aggregatum hyphae to the soaked and sterilized oat grains after the pythium aggregatum hyphae grows out;
s3: and (3) after the oat grains are full of mycelia, taking a mixture of the bacteria oat grains, sterile sandy soil and sterile water in a mass ratio of 1:1:2, respectively performing pythium massicot inoculation on the root of the peanut sample cultured in the step S1 in a seedling stage, a flowering and injection stage, a pod forming stage and a full fruit mature stage, and continuously culturing after a layer of sterile sandy soil is coated on the root, wherein 4 groups of experiments are all cultured until the full fruit mature stage of the peanut.
The experimental results are shown in table 1 and fig. 1, wherein fig. 1 is a photograph showing typical results of peanut rot disease when peanuts are inoculated with pythium at 4 different inoculation periods.
Disease grading standard: referring to Wheeler et al, studies have classified the grading criteria for peanut rot into:
level 1, no rotten fruit, namely the rotten fruit rate is 0;
grade 3, the rotten kernel rate is less than 2.5 percent, and the rotten fruit rate is less than 10 percent when the rotten kernel rate is more than 0;
grade 5, the rotten kernel rate is less than 2.5 percent, and the rotten fruit rate is less than 25 percent more than 10 percent;
grade 7, rotten kernel rate is less than 2.5%, and rotten fruit rate is less than 50% when the percentage is more than 25%;
grade 9, rotten kernel rate is more than 2.5%, or rotten fruit rate is more than 50%;
disease index% ═ 100 × Σ (number of diseased leaves at each stage × representative value at each stage)/(total number of investigated leaves × representative value at highest stage);
the disease incidence rate is (number of infected plants/total number of investigated plants) × 100%.
Table 1: the results of the disease conditions of peanut rot in different inoculation periods
The test result shows that: the pythium aggregatum is basically inoculated to peanuts in a seedling stage, a flowering and needle sowing stage, a pod forming stage and a full fruit ripening stage, the morbidity is certain, and the morbidity of pathogenic bacteria of the peanut rot is higher than 40%; meanwhile, the inoculation is carried out from the peanut flowering period to the pod bearing period, the inoculation success rate is higher, and particularly, the morbidity is 80% in the pod bearing period.
Through research on the difference of the inoculation rates of the pythium aggregatum in different peanut growth periods, the inventor finds that the peanut pythium aggregatum pathogenic bacteria, particularly the pythium aggregatum, have tissue specificity, namely the pathogenic bacteria only infect the peanut pods and do not infect the peanut roots, and even under the condition that other conditions are proper, the pathogenic bacteria can not be successfully colonized if the pathogenic bacteria do not infect the peanut pods. Therefore, when the peanut is inoculated with the pythium mass, the optimal inoculation period is the peanut pod bearing period.
The above description is only for the purpose of illustrating the preferred embodiments of the present invention and is not to be construed as limiting the invention, and any modifications, equivalents, improvements and the like that fall within the spirit and principle of the present invention are intended to be included therein.
Claims (6)
1. The peanut rot inoculation method is characterized by comprising the following steps:
s1: selecting healthy and plump peanut seeds, soaking the seeds in 75% alcohol for 0.5-1 min, then soaking the seeds in 10% sodium hypochlorite for 15min, sterilizing, washing with water for 3 times, and then sowing the seeds;
s2: culturing pathogenic bacteria of the peanut rot disease by using a culture medium, and inoculating the hypha of the pathogenic bacteria of the peanut rot disease to the soaked and sterilized grains after the hypha grows out;
s3: and (3) after the grains are overgrown with hypha, taking a mixture of bacteria grains, sterile sand and sterile water in a mass ratio of 1:1:2, covering the roots of the peanut samples cultured in the step S1, inoculating the pathogenic bacteria of the peanut rot, covering a layer of sterile sand on the roots, culturing until the peanuts are in a full fruit and mature period, and thus obtaining the peanut milk.
2. The method of claim 1, wherein the grain is oat groats.
3. The method for inoculating peanut rot disease as claimed in claim 1, wherein the pathogenic bacteria of peanut rot disease are pythium colonization.
4. The peanut rot inoculation method of claim 1, wherein the peanut seeds are sown and cultured indoors, and the indoor temperature is controlled to be 28-30 ℃ and the indoor humidity is controlled to be 85%; alternating 14h/10h light and dark.
5. The method for inoculating peanut rot disease bacteria of claim 1, wherein the peanut rot disease bacteria are inoculated for a period of time from the growth of peanuts to the stage of needle insertion or the stage of podding or between the stage of needle insertion and the stage of podding.
6. The method for inoculating peanut rot disease in accordance with claim 1, wherein the moisture content of the mixture of grains and sand is controlled to be greater than 30% and less than 60% after the roots of the peanut specimens are inoculated with the pathogenic bacteria of peanut rot disease.
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Citations (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN104762215A (en) * | 2014-12-12 | 2015-07-08 | 山东省花生研究所 | Rapid culture method for peanut pod rot pathogen |
| WO2016179047A1 (en) * | 2015-05-01 | 2016-11-10 | Indigo Agriculture, Inc. | Designed complex endophyte compositions and methods for improved plant traits |
| CN107135828A (en) * | 2017-04-30 | 2017-09-08 | 中国农业科学院油料作物研究所 | A kind of peanut sclerotium rolfsii greenhouse Seedling Inoculation method |
| CN109750080A (en) * | 2019-03-01 | 2019-05-14 | 山东省花生研究所 | A kind of method of the anti-fruit rot germ plasm resource of quick screening peanut |
| CN111296214A (en) * | 2020-03-20 | 2020-06-19 | 四川省农业科学院经济作物育种栽培研究所 | Indoor inoculation method for peanut soil-borne fungi |
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2020
- 2020-10-22 CN CN202011140672.4A patent/CN112243824A/en active Pending
Patent Citations (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN104762215A (en) * | 2014-12-12 | 2015-07-08 | 山东省花生研究所 | Rapid culture method for peanut pod rot pathogen |
| WO2016179047A1 (en) * | 2015-05-01 | 2016-11-10 | Indigo Agriculture, Inc. | Designed complex endophyte compositions and methods for improved plant traits |
| CN107135828A (en) * | 2017-04-30 | 2017-09-08 | 中国农业科学院油料作物研究所 | A kind of peanut sclerotium rolfsii greenhouse Seedling Inoculation method |
| CN109750080A (en) * | 2019-03-01 | 2019-05-14 | 山东省花生研究所 | A kind of method of the anti-fruit rot germ plasm resource of quick screening peanut |
| CN111296214A (en) * | 2020-03-20 | 2020-06-19 | 四川省农业科学院经济作物育种栽培研究所 | Indoor inoculation method for peanut soil-borne fungi |
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| 于静等: "防治花生腐霉果腐病的化学药剂筛选", 《中国油料作物学报》 * |
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