KR101153462B1 - A New bacteria to enhance rice production - Google Patents

A New bacteria to enhance rice production Download PDF

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KR101153462B1
KR101153462B1 KR1020100001743A KR20100001743A KR101153462B1 KR 101153462 B1 KR101153462 B1 KR 101153462B1 KR 1020100001743 A KR1020100001743 A KR 1020100001743A KR 20100001743 A KR20100001743 A KR 20100001743A KR 101153462 B1 KR101153462 B1 KR 101153462B1
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South Korea
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strain
rice
present invention
effect
kacc91522p
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KR1020100001743A
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KR20110081538A (en
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두 호 김
용 기 김
정 희 김
영 주 남
경 석 박
완 해 예
데레사 이
수 형 이
영 기 이
영 훈 이
용 환 이
준 성 이
인 호 정
성 숙 한
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대한민국
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Abstract

The present invention relates to a strain showing a rice increasing effect, and relates to a strain deposited with the accession number KACC91522P to the National Institute of Agricultural Science (KACC) and a method for increasing the yield of rice using the same.
According to the present invention, when cultivating rice using the new strain of the present invention, a high yield of rice yield of about 11.8% can be expected.

Description

A new bacteria to enhance rice production

The present invention relates to a new strain exhibiting the effect of rice increase, and more particularly to a strain deposited with the accession number KACC91522P to the National Institute of Agricultural Science (KACC) and a method for increasing the yield of rice using the same.

The present invention is a result of the research project of 'Pest incidence of high quality rice production and comprehensive management research', which is a research project of national technology development project, by 'improving rice pest resistance and growth promoting effect by PGPR priming'.

Increasing rice production in Korea has been mainly made of pesticides dependent on chemical pesticides. The misuse of these chemical pesticides has increased side effects such as residual toxicity of crops and destruction of agricultural ecosystem. With the economic growth and the growing public's interest in a pleasant environment and safe food, the transition to eco-friendly agriculture has recently come to the fore. As a result, methods for controlling crop diseases are being raised, rather than chemical control. Development of eco-friendly agricultural materials such as pesticides is required.

In this regard, several studies have shown that the plant growth promoting rhizobacteria (PGPR) among the microorganisms living in the root zone of the plant are present. The plant growth-promoting myobacteria act on the roots and surrounding environment and collectively refers to the rhizosphere bacteria that have the effect of promoting plant growth and productivity through interaction with plants (Kloepper and Schroth, 1978).

Studies on the use of plant growth promoting rhizobacteria (PGPR) have been carried out mainly in terms of promoting growth and biological control using nitrogen fixation. After the report that medically found streptomycin was effective against phytopathogens (Mitchell JW et al 1952), the metabolite of Streptomyces blastmyces was isolated and applied to blastidin S, which has a selective effect on rice blasts. (Takeuchi S. et al 1958), some antifungal antibiotics were also isolated, but there were various problems such as toxicity and mass use until they were put into practical use.

Nitrogen is an important nutrient for crop production and is provided primarily by chemical fertilizers or biological methods. In this regard, it has been reported through the study that root growth flora rooted in the roots of the plant roots fixed nitrogen in the air through symbiosis or non-symbiosis, resulting in an increase in crop yield and nitrogen fixation. The term rhizosphere, first defined by Hiltner (1904), initially meant a region where bacteria were grown around legumes, but now the roots of microorganisms are grown by nutrients secreted from the roots of plants. All areas are included, and recently, the area between the epidermis and the cortex is called endogenous root zone and microorganisms distributed in these areas are also called endogenous bacteria.

Studies on rhizosphere microorganisms in soil cultivation have shown that microspheres cluster in the plant root zone to inhibit pathogen growth, protect roots of plants, and promote plant growth, while parasitic roots cause disease. It has also been reported that there are also microorganisms (deleterious rhizosphere microorganisms (DRMO)) that are harmful to plants whose metabolites inhibit the activity of roots (Baker and Scher, 1987). The action of plant growth-promoting myobacteria acts differently depending on environmental factors and cultivation methods (Zhang et al., 1997).

Plant growth and rhizospheres that directly support plant growth through increased nutrient absorption, plant hormone production and suppression of plant diseases produce auxin, cytokineins and giberellins, and ethylene ( ethylene) inhibits disease by directly controlling plant growth and inducing resistance. Despite the many positive findings on the use of plant growth-promoting myobacteria, it is difficult to apply them to farmers because their stable effects, increased crop yields, and reduced need for chemical fertilizers have not been proven. It will also be necessary to analyze and manage the effects of the natural environment on the effects of plant growth-promoting myobacteria. Sanctions on the rise and use of chemical fertilizers and pesticides are increasing, and the use of microbial formulation technology is an eco-friendly product that can reduce environmental pollution and reduce costs by reducing chemical fertilizers and pesticides. Will be effective.

Accordingly, the present inventors have made intensive research efforts to identify new strains having excellent rice yielding effect. As a result, the present inventors have isolated bacteria from samples collected from various rice cultivation sites in the country, such as organic farming and duck farming, and increased rice by laboratory and greenhouse tests. It was possible to identify new strains having excellent effects, and when the strains were applied to rice cultivation in actual packaging, it was confirmed that a high rice yield increase effect appeared and completed the present invention.

Therefore, the main object of the present invention is to provide a new strain showing a high water effect in the rice cultivation.

Another object of the present invention to provide a method for increasing rice yield using the strain.

According to one aspect of the present invention, the present invention provides a strain deposited with accession number KACC91522P to the National Institute of Agricultural Science (KACC) on January 6, 2010.

The strain of the present invention exhibits a gram negative response, and is characterized by having a 16s rDNA consisting of the nucleotide sequence of SEQ ID NO: 1.

The strain of the present invention is characterized by exhibiting the effect of increasing rice.

According to another aspect of the present invention, the present invention provides a method for growing rice using the strain.

Hereinafter, the strain of the present invention and a rice cultivation method using the same will be described in more detail.

The strain of the present invention is a strain isolated from the rice plantation, and exhibits an inhibitory effect against rice fever and sesame seed germ in a test tube, and is a strain showing the effect of increasing the rice.

The strains of the present invention are Gram negative bacteria, weak to heat, cause hypersensitivity reactions on tomato leaves, and similar availability of Chromobacterium violaceum and carbon source, Pseudomonas-fluorescens-biotype C / P. Mandelii and fatty acid components are similar strains.

16S rRNA sequence is Chromobacterium sp. Similar to (see SEQ ID NO: 1), it is a rod-like strain.

The strain of the present invention has been deposited with the accession number KACC91522P in the National Institute of Agricultural Science (KACC), and can be distributed if necessary.

The strain of the present invention is preferably cultured using TSA (Tryptic Soy Agar) or TSB (Tryptic Soy broth) medium, and may be cultured by adding a culture additive as necessary.

The culture temperature is preferably 25 to 30 ° C., more preferably 28 ° C., in order to sufficiently obtain the cells, the culture is preferably carried out for 1 to 3 days, and the culture is best performed for 2 days.

When the strain of the present invention is applied to rice cultivation, it is preferable to use the lowest rice seed, and when maximizing the seed, it is preferable to sterilize the rice seed and then minimize the seed, and immerse the seed in the strain culture suspension of the present invention. It is good to use as a sowing seed so that the seed is sufficiently absorbed and coated on the seed.

The immersion time is preferably 1 to 3 hours, most preferably 2 hours.

The seed of the present invention to which the strain of the present invention is applied may be cultivated using any rice cultivation method, but it is preferable to use a transplanting method.

As described above, according to the present invention, when cultivating rice using the new strain of the present invention, a high rice yield increase of about 11.8% can be expected.

1 is a photograph showing the antimicrobial activity of the strain of the present invention against the blast disease (A: strain of the present invention, B: control).
Figure 2 is a photograph showing the antimicrobial activity effect against rice bran pattern disease of the strain of the present invention (A: strain of the present invention, B: control).
Figure 3 is a photograph showing the morphological characteristics of the strain of the present invention by an electron microscope.

Hereinafter, the present invention will be described in more detail with reference to Examples. These examples are only for illustrating the present invention, and the scope of the present invention is not to be construed as being limited by these examples.

Example 1. Isolation and Screening of Strains

end. Isolation of strain

As a sample for the isolation of the strains, the rice roots and the harvested rice grains of the highest wheat harvester collected from organic and general farmland among rice plantations were used. Collected rice root samples were first removed from the soil attached to the roots using running tap water, thoroughly removed the remaining soil with distilled water, and then cut off the roots. After soaking for 1 minute, 1 ml of sterile water was added and crushed. The harvested rice grains were removed from the shell and used as brown rice, and then sterilized and ground in the same manner as the rice root sample. Some of the ground samples were plated in 1/5 TSA (Tryptic Soy Agar) medium and incubated at 28 ° C. for 3 days to induce different cell colonies to form and purely separated them.

I. Preparation of Target Pathogens to Investigate the Inhibitory Effect of Strains

Was used for Magnaporthe grisea (rice blast fungus), Cochliobolus miyabeanus (kkaessi pattern germs) and Thanatephorus cucumeris (sheath blight fungus) to investigate the purified a disease inhibiting effect of the present invention strains (KACC91522P) targeted pathogen, Magnaporthe grisea were incubated 7 days at 25 ℃ was inoculated on rice bran agar medium (rice polish agar, RPA), after removal of the aerial hyphae, open the lid down on the sterile fluorescent lamp by collecting the following spores for 3 days additional culture 10 5 conidia It was used after adjusting to a concentration of / ml. Cochliobolus miyabeanus was inoculated in sucrose proline agar (SPA) medium and incubated at 25 ° C. for 3 days, followed by further incubation for 2 weeks under sterile fluorescent lamps, and then adjusted to a concentration of 10 4 conidia / ml. Thanatephorus cucumeris was inoculated in PDA (potato dextrose agar) medium and incubated at 28 ° C. for 2 weeks to use mycelia as inoculum.

All. Investigation of the inhibitory effect of strains

The target pathogen was inoculated on a PDA (Potato Dextrose Agar) medium and incubated at 28 ° C. for 7 days, and the mycelia were collected with 5 mm cork borer and placed in the center of a new PDA medium. The strain of the present invention was TSA. (Tryptic Soy Agar) inoculated in the medium and incubated for 48 hours at 28 ℃, suspended in sterile water (OD 600 = 0.1) and inoculated 20μL on a paper disc (paper disc) and then put the target pathogen of the PDA medium Placed on the edge and incubated at 28 ℃ for 7 days, the size of the low support was investigated.

As a result, as shown in Table 1 and FIGS. 1 and 2, low support was formed for Magnaporthe grisea and Cochliobolus miyabeanus , respectively. However, there was no inhibitory effect on Thanatephorus cucumeris .

Target pathogen Magnaporthe grisea Thanatephorus cucumeris Cochliobolus miyabeanus KACC91522P 1.7 0.0 2.1 Control group 0.0 0.0 0.0

* Unit: mm

* KACC91522P: strain of the present invention

la. Investigation of Rice Growth Effect of Strains in Greenhouse

In order to investigate the effect of increasing the rice of the strain of the present invention in greenhouse cultivation of rice, seedlings treated with the strain of the present invention were sown in a greenhouse, and then the seedling, leaf length, leaf width, root length, root weight and dry weight of seedlings were examined.

The rice seeds to be used for greenhouse sowing were saline-dried and soaked in a 2,000-fold dilution of prochlorarcoperchloride seed treatment liquid hydrous solution for 24 hours, washed three times with sterile water and dried. The strain culture suspension was immersed for 2 hours and used for sowing. As a control, TSB medium without inoculation of the strain was used instead of the strain culture suspension.

Vermiculite and rice seedlings were mixed 2: 3 and used as culture soil, and the seed treated with the strain and the seed to be used as a control were sown in square water (15 × 15 × 10㎝) for water and then in the wet phase. (Dew chamber) was incubated for 2 days and then grown in a greenhouse. After 21 days of sowing, the height, root length, leaf length, leaf width and leaf area were examined.

Extra long (mm) Root length (mm) Leaf Length (mm) Leaf width (mm) Leaf area (㎡) KACC91522P 374.95 160.70 235.15 6.74 1196.19 Control group 359.90 148.45 219.20 6.59 1090.02

* KACC91522P: strain of the present invention

As a result, as shown in Table 2, when treated with the strain of the present invention, the height, root length, leaf length, leaf width and leaf area were all increased compared to the control group, especially the root length was increased.

hemp. Investigating the Effect of Strains on Packaging

In order to investigate the effect of the strain of the present invention in the field, the experiment was carried out in the National Agricultural Science Institute Icheon test.

Seeds were transplanted after treatment in the same manner as in the greenhouse test.

In this field of study, Benzothiadiazole (BTH) or INA (2,6-dichloroisonicotinic acid), which are well known strains of the present invention as well as inducible resistant substances and well known as pest control agents, were treated together or separately. In the experimental group treated with only the resistant expression material, the disinfected seeds were soaked in BTH (20 mg / ml) or INA (20 mg / ml) for 2 hours, respectively, and treated with the inducible resistance material and the strain of the present invention. One experimental group was sprayed on the foliar after 3 weeks of sowing of the strain of the present invention, mixed with BTH (20 mg / mL) or INA (20 mg / mL) and Tween 20 (1 / 5000-fold dilution) solution, respectively. Distilled water was used instead of the strain, BTH and INA of the present invention.

Yield survey was taken 45 days after the pan was dried, dried and then the basis weight, converted to 14% moisture content.

Treatment fungus Quantity Component Quantity Earnings per share Earnings per head Maturity rate (%) (G) KACC91522P 18.4 72.8 79.3 25.2 682.1 BTH 18.3 61.9 84.0 25.6 609.2 INA 17.9 61.5 81.5 25.4 571.8 KACC91522P + BTH 18.5 64.7 76.3 25.3 603.7 KACC91522P + INA 18.9 63.9 80.4 25.6 627.1 Control group 18.8 62.3 82.2 25.5 610.2

As a result, as shown in Table 3, when only the strain of the present invention was treated, the increase effect of 16.9% and 11.8%, respectively, in the number of grains per ear and the yield was shown. This means that the increase effect is due to the increase in pure water rather than the increase in the number of ears per week, ripening rate, and grain weight, which means that there is an increase in rice production.

bar. Identification of the strain

In order to identify the strain of the present invention, the effect of increasing rice, the Gram, hypersensitivity reaction and heat resistance were tested, the size and shape were examined through a microscope, BIOLOG analysis (consumption carbon source analysis), MIDI analysis ( Fatty acid analysis) and 16S rRNA sequencing.

The strain of the present invention was cultured in a tryptic soy agar (TSA) medium (28 ° C., 24 hours) and then subjected to a Gram test using 3% KOH. The tomato leaf was inoculated with the strain suspension of the present invention for hypersensitivity reactions. Was investigated.

For heat resistance experiment, TSB (tryptic soy broth) and MnSO 4 were added to the test tube and sterilized. The strains of the present invention were inoculated and incubated at 28 ° C. at 150 rpm for 3 days, and then at 80 ° C. in a 100 rpm water bath. After 10 minutes of treatment, the reaction was quenched to investigate the reaction.

Size and morphological characteristics were investigated using Transmission Electron Microscope and Scanning Electron Microscope.

BIOLOG analysis was inoculated strains of the present invention in GN2 microplate and incubated for 24 hours at 28 ℃ suspended in 0.85% saline solution and 150μL suspension was dispensed on microplate (BIOLOG MicroPlate TM ) and incubated for 48 hours at 28 ℃ The patterns formed were identified by cross-referencing the library patterns on the biolog microlog computer program.

MIDI analysis was performed by extracting fatty acid methyl esters from the strain of the present invention according to the standard method of MIDI-FAME, followed by Hewlett-Packard gas chromatograph (model 6890) and Microbial Identification system (MIDI; Microbial ID, Inc., Newark, Del., USA ).

16S rRNA sequencing was performed using genomic DNA prep kit (SolGent Co., Ltd.) from cells cultured at 28 ° C. for 24 hours and then identified by solgent.

Identification of the strain was performed by Berg's manual.

As a result, the strains of the present invention are Gram-negative bacteria, weak to heat and induce hypersensitivity reactions, Chromobacterium violaceum (BIOLOG test results), Pseudomonas-fluorescens-biotype C / P. mandelii (MIDI test results) and Chromobacterium sp. (Refer to 16S rRNA sequencing results, SEQ ID NO: 1), and observed by electron microscopy, it was confirmed that it is in the form of rod as shown in FIG.

Agricultural Biotechnology Research Institute KACC91522 20100107

Attach an electronic file to a sequence list

Claims (4)

  1. Chromobacterium violaceum KRYP101 strain KACC91522P.
  2. According to claim 1, wherein the strain is Gram negative reaction, characterized in that having a 16S rDNA represented by the nucleotide sequence of SEQ ID NO: 1.
  3. According to claim 1, wherein the strain is characterized in that the strain exhibits the effect of increasing the rice.
  4. A method of growing rice using the strain of any one of claims 1 to 3.
KR1020100001743A 2010-01-08 2010-01-08 A New bacteria to enhance rice production KR101153462B1 (en)

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Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
KR100646597B1 (en) 2004-06-07 2006-11-23 대한민국 228 Pseudomonas sp. RRj228 helping the growth of crops and the microbial agent containing the same
KR100646601B1 (en) 2004-06-12 2006-11-23 대한민국 083 91050 Bacillus sp. KR083KACC91050 promoting the growth of crops and the microbial agent containing the same

Patent Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
KR100646597B1 (en) 2004-06-07 2006-11-23 대한민국 228 Pseudomonas sp. RRj228 helping the growth of crops and the microbial agent containing the same
KR100646601B1 (en) 2004-06-12 2006-11-23 대한민국 083 91050 Bacillus sp. KR083KACC91050 promoting the growth of crops and the microbial agent containing the same

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