CN105219651A - A kind ofly improve Rice Resistance arid, interior raw Aspergillus fumigatus that salt damage coerces ability and application thereof - Google Patents
A kind ofly improve Rice Resistance arid, interior raw Aspergillus fumigatus that salt damage coerces ability and application thereof Download PDFInfo
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- CN105219651A CN105219651A CN201510504330.9A CN201510504330A CN105219651A CN 105219651 A CN105219651 A CN 105219651A CN 201510504330 A CN201510504330 A CN 201510504330A CN 105219651 A CN105219651 A CN 105219651A
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Abstract
The invention discloses a kind of Aspergillus fumigatus and application thereof.Does is this Aspergillus fumigatus preservation name called <i>Aspergillus</ i><i>? fumigatus? </i>SG17; Depositary institution: China typical culture collection center; Preservation address is China, Wuhan, Wuhan University; Preservation date: on May 11st, 2015; Does is deposit number M? 2015286.The present invention the experiment proved that, aspergillus fumigatus physical efficiency improves the resistivity of coercing Drought and salt evil of paddy rice, not only can improve the growth of paddy rice under above-mentioned Abiotic stress conditions, have laid a good foundation for screening has novel anti-microorganism of coercing active substance simultaneously, there is good economic and social benefit.
Description
Technical field
The present invention relates to raw Aspergillus fumigatus and application thereof in a strain, be mainly used in paddy rice waits abiotic stress environment tolerance to Drought and salt evil, facts have proved that this Aspergillus fumigatus may be used for improving paddy rice to the resistance of above-mentioned abiotic stress factor.
Background technology
Paddy rice (Oryzasativa) originates in Tropical Asian, after China widely plants, propagates into gradually all over the world.Nearly half population be in the world all food with rice, therefore the plantation of paddy rice has material impact with production to national economy.
The endophyte of plant is the microorganism that a class certain one-phase in plant life history moves in health tissues or organ inside, research in recent years shows, endophyte is resisted in various stressful environmental assistance host plant and is play very important role, as the oxidative stress that the biotic factors such as disease, arid, high salt or abiotic factor cause.Because the species diversity of natural endogenetic fungus is very abundant, provide rich and varied secondary metabolite and biological activity class material, to industrial and agricultural production, human health, there is important value, thus there is huge economic benefit and social benefit.
Aspergillus fumigatus belongs to imperfect fungi fungi, and this bacterium is mainly present in cereal, the food of pollution, soil and mould rotten thing, or posts and be hidden on seed, can cause pulmonary aspergillosis and the other diseases of people, animal and bird, can infect cotton boll and apple etc., cause fruit rot.This bacterium can produce the bent clavine (fumigaclavines) of cigarette, Aspergillus fumigatus dardarin (fumitremorgins), the multiple secondary metabolites such as fumidil (fumigillin), FT (fumitoxins), penicillium verruculosum former (verruculogen), but about the growth-promoting functions of Aspergillus fumigatus to plant, also rarely have report both at home and abroad at present.
The present invention is separated and obtains raw Aspergillus fumigatus in a strain from M. laxiflora (Myricarialaxiflora) root, and it can improve paddy rice well to resistance that is arid and salt damage, can serve plantation and the production of paddy rice.
Summary of the invention
An object of the present invention is to provide a kind of Aspergillus fumigatus (Aspergillusfumigatus) SG17 bacterial strain, this Aspergillus fumigatus can improve the resistance of paddy rice well, can be used for Rice Resistance Drought and salt evil and coerce aspect, for the paddy rice of developing high-quality provides new direction.
Described Aspergillus fumigatus (Aspergillusfumigatus) SG17 is the endogenetic fungus obtained from the root of M. laxiflora (MyricariaLaxiflora), be preserved in China typical culture collection center on May 11st, 2015, deposit number is: M2015286.Preservation address is China, Wuhan, Wuhan University.
Aspergillus fumigatus of the present invention (Aspergillusfumigatus) SG17 has following Microbiological Characteristics on PDA substratum: bacterium colony starts as white, blue-greenish colour is transferred to after 2-3 day, but edge is still white, becomes deep green after with the passing of time cultivating, cigarette is green.Be just velvet-like or cotton-shaped, As time goes on becoming Powdered, also there is color in edge section, and the back side is colourless or be with some tawny.Only the first half produces round shape conidium, and color is the green that the depth is different, and conidiophore is smooth, and top capsule is flask shape.
The object of the invention is to provide a kind of for improving paddy rice to the microbial inoculum of Drought and salt evil stress resistance and biological modification agent, and its activeconstituents is Aspergillus fumigatus SG17.Stress tests shows, Aspergillus fumigatus SG17 can improve the resistance of wide spectrum of paddy rice.Those skilled in the art are easy to expect that the fermented liquid of bacterial strain of the present invention or fermentation broth extract are used for paddy rice opposing Drought and salt evil as the effective constituent improving paddy rice resistance coerces.Thus, the present invention also comprises the biological modification agent containing described bacterial strain or its tunning.
The present invention with the DNA of Aspergillus fumigatus (Aspergillusfumigatus) SG17 for template, upstream primer ITS1:5 '-TCCGTAGGTGAACCTGCG-3 ', downstream primer ITS4:5 '-TCCTCCGCTTATTGATATGC-3 ', carry out colony PCR amplification, obtain the 18SrDNA partial sequence (SEQIDNO:1) of 560bp.The sequence of being included by Blast and GenBank is compared, and finds that the homology of SG17 and Aspergillus fumigatus is 100%.
Accompanying drawing explanation
Fig. 1 is the colonial morphology of Aspergillus fumigatus of the present invention (Aspergillusfumigatus) SG17.
Fig. 2 is the microscopic morphology of Aspergillus fumigatus of the present invention (Aspergillusfumigatus) SG17.
Fig. 3 is that Aspergillus fumigatus of the present invention (Aspergillusfumigatus) SG17 thalline improves Rice Resistance drought stress action effect.Wherein, before A. coerces, B. arid+SG17, C. arid, D. arid+proline(Pro), E. is blank.Proline(Pro) strength of solution is 5 × 10
-5mol/l blank is tap water.
Fig. 4 is that Aspergillus fumigatus of the present invention (Aspergillusfumigatus) SG17 thalline improves Rice Resistance salt damage coercion effect.Wherein, before F. coerces, G. salt damage+SG17, H. salt damage, I. salt damage+proline(Pro), J. is blank, and proline(Pro) strength of solution is 5 × 10
-5mol/l blank is tap water.
Embodiment
Embodiment one: the separation of Aspergillus fumigatus (Aspergillusfumigatus) SG17 and preservation
Within 2012, in Yichang, basin, Three Gorges, domestic kermes dam gathers M. laxiflora root, is inoculated in and adds the PDA solid medium separation fungi that working concentration is 25 μ g/ml penicillin after surface sterilization.
Surface sterilization: get M. laxiflora root tap water 5 times, after aseptic filter paper is dried, 75% alcohol-pickled 1min, aseptic water washing 3 times, then soak 60S with 3% clorox, then use aseptic water washing 5 times, aseptic filter paper washes dry root surface-moisture.
Cultivate: by sterile scissors, the root after surface sterilization is cut into 0.5*0.5cm size, be inoculated in that to add working concentration be 25 μ g/ml penicillin P DA solid mediums (potato 200g/L, glucose 20g/L, agar 20g/L, Ph6.0-6.5), cultivate at being placed in 28 DEG C.
Bacterial strain purifying: after flat board grows bacterium colony, according to form picking list colony inoculation in new PDA solid medium, cultivates at being placed in 28 DEG C.
Bacterial strain preservation: after bacterial strain purifying, the pure inoculation of picking, in PDA slant medium, cultivates 4d for 28 DEG C, is hidden in 4 DEG C of preservations by the whiteruss envelope of sterilizing.
This bacterial strain Aspergillus fumigatus (Aspergillusfumigatus) is carried out preservation, and preservation name is called SG17; Depositary institution: China typical culture collection center; Preservation address is China, Wuhan, Wuhan University; Preservation date: on May 11st, 2015; Deposit number is, M2015286.
Embodiment two: Aspergillus fumigatus (Aspergillusfumigatus) SG17 identifies
The morphological specificity of bacterial strain and microscopic features are shown in accompanying drawing 1..
18SrDNA sequential analysis: adopt universal primer to carry out colony PCR amplification 18SrDNA sequence.Upstream primer ITS1:5 '-TCCGTAGGTGAACCTGCG-3 ', downstream primer ITS4:5 '-TCCTCCGCTTATTGATATGC-3 '.Reaction system is as follows:
Reaction conditions: 94 DEG C of denaturation 5min, 94 DEG C of sex change 1min, 56 DEG C of annealing 30sec, 72 DEG C extend 1.min, 33 circulations, and last 72 DEG C extend 10min.PCR primer 1% agarose gel electrophoresis detects, and the band of picking about 750bp carries out gel recovery, serves Hai Shenggong order-checking.
The sequence recorded is committed to GenBank, and obtains accession number KR812470.
Embodiment three: the preparation of Aspergillus fumigatus (Aspergillusfumigatus) SG17 thalline
Thalline prepare: go bail for deposit 4 DEG C of refrigerators the streak inoculation of Aspergillus fumigatus kind in PDA flat board, at 28 DEG C, cultivate 4d, obtain thalline.
Prepared by fermented liquid: beat on above-mentioned bacterium colony culture medium flat plate with punch tool and get the triangular pyramidal bottle that 1 bacterium cake is placed in the 500ml containing 100mlPDA liquid nutrient medium, shake up, be then placed in 28 DEG C of shaking table shaking culture (120rpm), obtains fermented liquid.
Embodiment four: Aspergillus fumigatus (Aspergillusfumigatus) SG17 thalline improves the effect of Rice Resistance drought stress ability
Prepared by rice seedling: the fine seed of Japan taken out from 4 DEG C of refrigerators is placed in tap water and soaks 4d, impel it to root, and change a water every 12h.Be planted in after 4d and be loaded with in the plastic cup of soil, regularly watered, the fine seedling of paddy rice Japan after 7d, can be obtained.
Drought stress is tested: obtain experimental group, with 5 × 10 by joining in Japanese fine seedling after fermented liquid supersonic wave wall breaking 2h
-5mol/l proline(Pro) solution replaces fermented liquid to be positive control group, and tap water is blank group, and not supplementing any material is drought stress, often group do three parallel, cultivate under above-mentioned seedling being placed the room temperature of 25-30 DEG C.Coerce and start fully to water to each seedling the day before yesterday, only quantitatively to keep the skin wet to blank group coercing period, coerce 12 days altogether.
Embodiment five: rice drought coerces rear seedling physiology Indexs measure
Mda content: the plant lipid peroxidation brought out with accumulated active oxygen that sustains an injury under adverse circumstance is closely related, and the product of membrane lipid peroxidatio has lipid peroxide, mda, ethane etc., and wherein mda is one of most important product.Therefore the degree of membrane lipid peroxidatio is understood by measuring mda content, with the resistance of the indirect measurement membranous system extent of damage and plant.Nanjing is utilized to build up the plant Mda kit measurement mda content of Bioengineering Research Institute.
Proline content: growing plants under ambient environment, in body, the content of Proline is lower; But under adverse environmental factor, in plant materials, the content of Proline can significantly increase.Proline(Pro) wetting ability is extremely strong, can stablize protoplasma colloid and in-house metabolic process.Therefore, the content of Proline can be used as the index of stress resistance of plant.The proline(Pro) testing cassete utilizing Nanjing to build up Bioengineering Research Institute measures proline content.
The detected result of mda and proline(Pro) after drought stress
Paddy rice mda content after table one arid
Paddy rice proline content after table two arid
Positive control in table 1.2 is 5 × 10-5mol/l proline(Pro) solution, and blank is tap water.
Embodiment six: Aspergillus fumigatus (Aspergillusfumigatus) SG17 thalline improves Rice Resistance salt damage and coerces ability effect
Prepared by rice seedling: the fine seed of Japan taken out from 4 DEG C of refrigerators is placed in tap water and soaks 4d, impel it to root, and change a water every 12h.Be planted in after 4d and be loaded with in the plastic cup of soil, regularly watered, the fine seedling of paddy rice Japan after 7d, can be obtained.
Salt damage stress tests: obtain experimental group, with 5 × 10 by joining in Japanese fine seedling after fermented liquid supersonic wave wall breaking 2h
-5mol/l proline(Pro) solution replaces fermented liquid to be positive control group, and tap water is blank group, and not supplementing any material is drought stress, often group do three parallel, cultivate under above-mentioned seedling being placed the room temperature of 25-30 DEG C.Coerce and start in experimental group and positive control group, fully to add 7.5g/LNaCl solution the day before yesterday, quantitatively keep the skin wet to each seedling during coercing, coerce 12 days altogether.
Claims (7)
1. Aspergillus fumigatus (
aspergillusfumigatus) SG17, be preserved in China typical culture collection center on May 11st, 2015, deposit number is: M2015286.
2., for improving Rice Resistance arid and a salt damage modifying agent of coercing, its activeconstituents is Aspergillus fumigatus.
3. a biological modification agent, it contains the Aspergillus fumigatus of claim 1.
4. Aspergillus fumigatus according to claim 1, modifying agent according to claim 2 or biological modification agent according to claim 3 application in raising Rice Resistance arid and salt damage are coerced, its environment stress is arid and salt damage.
5. the application in ability is coerced in Aspergillus fumigatus according to claim 1 or biological modification agent according to claim 2 in raising Rice Resistance arid and salt damage.
6. application according to claim 5, wherein said biological modification agent improves the resistance that paddy rice is coerced arid and salt damage.
7. the Aspergillus fumigatus described in any one of claim 1-6, this Aspergillus fumigatus is separated and obtains in the root of M. laxiflora.
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| CN201510504330.9A CN105219651B (en) | 2015-08-17 | 2015-08-17 | It is a kind of to improve interior raw aspergillus fumigatus and its application that rice is drought-resistant, salt damage coerces ability |
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| CN201510504330.9A CN105219651B (en) | 2015-08-17 | 2015-08-17 | It is a kind of to improve interior raw aspergillus fumigatus and its application that rice is drought-resistant, salt damage coerces ability |
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Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN105754900A (en) * | 2016-04-13 | 2016-07-13 | 西北农林科技大学 | Novel species of Erwinia alhagi facilitating drought resistance of crops and application of novel species of Erwinia alhagi |
| CN107935880A (en) * | 2017-11-27 | 2018-04-20 | 三峡大学 | The separation identification and its application of a kind of anti-oxidant product for the aspergillus fumigatus for improving Rice Drought Resistence ability |
Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2009012480A2 (en) * | 2007-07-19 | 2009-01-22 | Montana State University | Fungal isolates and their use to confer salinity and drought tolerance in plants |
| CN104762216A (en) * | 2014-12-26 | 2015-07-08 | 南京晓庄学院 | Fungus strain capable of resisting salt stress, and breeding method and application thereof |
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2015
- 2015-08-17 CN CN201510504330.9A patent/CN105219651B/en active Active
Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2009012480A2 (en) * | 2007-07-19 | 2009-01-22 | Montana State University | Fungal isolates and their use to confer salinity and drought tolerance in plants |
| CN104762216A (en) * | 2014-12-26 | 2015-07-08 | 南京晓庄学院 | Fungus strain capable of resisting salt stress, and breeding method and application thereof |
Non-Patent Citations (1)
| Title |
|---|
| LIU S.等: "KR812470", 《GENBANK》 * |
Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN105754900A (en) * | 2016-04-13 | 2016-07-13 | 西北农林科技大学 | Novel species of Erwinia alhagi facilitating drought resistance of crops and application of novel species of Erwinia alhagi |
| CN105754900B (en) * | 2016-04-13 | 2020-04-17 | 西北农林科技大学 | Novel Erwinia strain for promoting crop drought resistance and application thereof |
| CN107935880A (en) * | 2017-11-27 | 2018-04-20 | 三峡大学 | The separation identification and its application of a kind of anti-oxidant product for the aspergillus fumigatus for improving Rice Drought Resistence ability |
| CN107935880B (en) * | 2017-11-27 | 2020-04-24 | 三峡大学 | Separation and identification of aspergillus fumigatus antioxidant product for improving drought resistance of rice and application thereof |
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| CN105219651B (en) | 2018-07-24 |
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Inventor after: Liu Shiping Inventor after: Qin Wang Ge pavilion Inventor after: Jiang Wei Inventor after: Kong Yushan Inventor after: Li Yuantao Inventor after: Xue Yanhong Inventor after: Gao Yuan Inventor before: Liu Shiping Inventor before: Jiang Wei Inventor before: Kong Yushan Inventor before: Qin Wang Ge pavilion Inventor before: Li Yuantao Inventor before: Xue Yanhong Inventor before: Gao Yuan |
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Effective date of registration: 20220114 Address after: No. 168, zhaoqiao Henan Road, Nanjing Chemical Industrial Park, Nanjing, Jiangsu 210000 Patentee after: TRUST CROP PROTECTION TECHNOLOGY CO.,LTD. Address before: 443002 No. 8, University Road, Yichang, Hubei Patentee before: CHINA THREE GORGES University |