KR20140117221A - Plant growth substance and anti-root knot nematode compound and its composition using Lysobacter capsici YS1215 from crab shell amended soil and manufacturing method thereof - Google Patents
Plant growth substance and anti-root knot nematode compound and its composition using Lysobacter capsici YS1215 from crab shell amended soil and manufacturing method thereof Download PDFInfo
- Publication number
- KR20140117221A KR20140117221A KR1020130032401A KR20130032401A KR20140117221A KR 20140117221 A KR20140117221 A KR 20140117221A KR 1020130032401 A KR1020130032401 A KR 1020130032401A KR 20130032401 A KR20130032401 A KR 20130032401A KR 20140117221 A KR20140117221 A KR 20140117221A
- Authority
- KR
- South Korea
- Prior art keywords
- capsici
- lysobacter
- plant growth
- soil
- nematode
- Prior art date
Links
Images
Classifications
-
- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01N—PRESERVATION OF BODIES OF HUMANS OR ANIMALS OR PLANTS OR PARTS THEREOF; BIOCIDES, e.g. AS DISINFECTANTS, AS PESTICIDES OR AS HERBICIDES; PEST REPELLANTS OR ATTRACTANTS; PLANT GROWTH REGULATORS
- A01N63/00—Biocides, pest repellants or attractants, or plant growth regulators containing microorganisms, viruses, microbial fungi, animals or substances produced by, or obtained from, microorganisms, viruses, microbial fungi or animals, e.g. enzymes or fermentates
- A01N63/20—Bacteria; Substances produced thereby or obtained therefrom
-
- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01N—PRESERVATION OF BODIES OF HUMANS OR ANIMALS OR PLANTS OR PARTS THEREOF; BIOCIDES, e.g. AS DISINFECTANTS, AS PESTICIDES OR AS HERBICIDES; PEST REPELLANTS OR ATTRACTANTS; PLANT GROWTH REGULATORS
- A01N25/00—Biocides, pest repellants or attractants, or plant growth regulators, characterised by their forms, or by their non-active ingredients or by their methods of application, e.g. seed treatment or sequential application; Substances for reducing the noxious effect of the active ingredients to organisms other than pests
- A01N25/02—Biocides, pest repellants or attractants, or plant growth regulators, characterised by their forms, or by their non-active ingredients or by their methods of application, e.g. seed treatment or sequential application; Substances for reducing the noxious effect of the active ingredients to organisms other than pests containing liquids as carriers, diluents or solvents
- A01N25/04—Dispersions, emulsions, suspoemulsions, suspension concentrates or gels
-
- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01N—PRESERVATION OF BODIES OF HUMANS OR ANIMALS OR PLANTS OR PARTS THEREOF; BIOCIDES, e.g. AS DISINFECTANTS, AS PESTICIDES OR AS HERBICIDES; PEST REPELLANTS OR ATTRACTANTS; PLANT GROWTH REGULATORS
- A01N63/00—Biocides, pest repellants or attractants, or plant growth regulators containing microorganisms, viruses, microbial fungi, animals or substances produced by, or obtained from, microorganisms, viruses, microbial fungi or animals, e.g. enzymes or fermentates
- A01N63/30—Microbial fungi; Substances produced thereby or obtained therefrom
- A01N63/32—Yeast
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
- C12N1/20—Bacteria; Culture media therefor
Landscapes
- Life Sciences & Earth Sciences (AREA)
- Health & Medical Sciences (AREA)
- Engineering & Computer Science (AREA)
- General Health & Medical Sciences (AREA)
- Zoology (AREA)
- Microbiology (AREA)
- Wood Science & Technology (AREA)
- Biotechnology (AREA)
- Chemical & Material Sciences (AREA)
- Pest Control & Pesticides (AREA)
- Virology (AREA)
- Dentistry (AREA)
- Plant Pathology (AREA)
- Environmental Sciences (AREA)
- Agronomy & Crop Science (AREA)
- Bioinformatics & Cheminformatics (AREA)
- Genetics & Genomics (AREA)
- Organic Chemistry (AREA)
- Mycology (AREA)
- Tropical Medicine & Parasitology (AREA)
- Medicinal Chemistry (AREA)
- Toxicology (AREA)
- Biomedical Technology (AREA)
- Dispersion Chemistry (AREA)
- Biochemistry (AREA)
- General Engineering & Computer Science (AREA)
- Agricultural Chemicals And Associated Chemicals (AREA)
Abstract
Description
본 발명은 식물 생장 촉진제 및 선충 방제제에 관한 것으로서, 특히 게껍질이 풍부하게 포함된 토양으로부터 분리한 Lysobacter capsici YS1215를 이용한 식물 생장 촉진제 및 선충 방제제와 그 제조방법에 관한 것이다.The present invention relates to a plant growth promoter and a nematode controlling agent, and more particularly, to a Lysobacter capsici A plant growth promoting agent and a nematocide prevention agent using YS1215, and a production method thereof.
본 발명은 농림수산식품부로부터 지원받아 수행된 연구(연구사업명: 생명산업기술개발, 연구과제명: 뿌리혹선충 방제를 위한 항선충 미생물 대량배양복합체 개발)의 일환으로 개발된 것이다.
The present invention has been developed as part of the research conducted under the support of the Ministry of Food, Agriculture, Forestry and Fisheries (research project: development of life industry technology, research title: development of mass culture complex of nematode microorganism for root-knot nematode control).
뿌리혹선충(Meloidogyne)은 식물 기생선충으로서, 식물 기생선충류 중에 가장 많은 경제적 손실을 가져다 주는 해충이다. 기주식물은 잔디에서부터 나무에 이르기까지 2000여종에 이른다. Meloidogyne , a plant parasitic nematode, is the most economically disadvantaged pest among plant parasitic nematodes. Host plants range from grass to trees to about 2,000 species.
뿌리혹선충들 중 전 세계적으로 광범위하게 분포하고 있으며 피해를 주고 있는 종은 고구마뿌리혹선충(M. incognita)과 자바뿌리혹선충(M. javanica), 땅콩뿌리혹선충(M. arenaria), 당근뿌리혹선충(M. hapla) 등이 있는데, 우리나라의 경우도 시설 재배지에서의 연작으로 인해 뿌리혹선충 발생이 증가하고 있으며 피해액과 피해 대상작물이 점차 확대되고 있는 실정이다.Kind of giving has extensive worldwide distribution of root-knot nematode damage, sweet potato root-knot nematode (M. incognita) and Java root-knot nematodes (M. javanica), peanut root-knot nematode (M. arenaria), carrot root-knot nematode (M hapla ) . In Korea, the occurrence of root-knot nematodes is increasing due to the sequence in the plantation area, and the amount of damages and the crops to be damaged are gradually expanding.
뿌리혹선충은 식물체 뿌리에 2령 유충 시기에 침입하여 기생하면서 영양분을 흡즙하는 식물 내부 기생성 선충으로서, 식물 내부에서 3령, 4령을 거쳐 암수가 나누어지며 암컷은 뿌리 바깥쪽에 난낭을 만들어 500-600개의 알을 낳고, 수컷은 토양으로 나간다. 난낭에서 부화된 2령 유충은 재차 뿌리에 침입하여 세대를 반복한다. The root-knot nematode is a plant internegative nematode that enters the root of the plant and feeds the parasitic nutrients into the plant roots. During the third and fourth decades, the male and female parts of the plant divide into male and female, 600 eggs are laid, and the male goes to the soil. The second - instar larvae hatching in the ovary recede into the roots again and repeat the generation.
이러한 뿌리혹선충은 주로 시설원예작물인 오이, 수박, 참외, 토마토, 멜론 등에 많은 피해를 주며, 뿌리혹선충에 감염된 식물은 선충이 대부분의 식물 양분을 흡즙함으로써, 식물체 생육이 지연되거나 감소하여 쉽게 시들거나 고사하는 증상이 나타난다. 또한, 각종 토양 병해와의 상호작용으로 2차 피해가 수반된다.These root-knot nematodes cause many damage to the horticultural crops such as cucumber, watermelon, melon, tomato, melon and the like, and the plants infected with root-knot nematode are fed by nematode most of the plant nutrients, Symptoms of death appear. In addition, secondary damage is accompanied by interaction with various soil diseases.
지금까지, 이러한 뿌리혹선충의 방제는 주로 화학적 방제에 의존해 오고 있다. 그러나 화학 살선충제는 고독성이고 토양 잔류기간이 길기 때문에, 토양 미생물에 영향을 미침으로써 토양생태계 불균형뿐만 아니라 지하수 오염과 같은 환경문제를 야기시키며, 살선충제의 남용으로 인하여 뿌리혹선충의 약제 저항성을 증가시킬 수 있는 부작용이 있다.Up to now, the control of these root-knot nematodes has been largely dependent on chemical control. However, since the chemical nematode is highly toxic and has a long soil retention period, it affects soil microorganisms, causing environmental problems such as groundwater contamination as well as imbalance of soil ecosystem. The abuse of nematocide causes chemical resistance of root nematode There is a side effect that can be increased.
따라서 화학적 방제법을 대체할 방법으로 토양개량, 담수 온탕처리 및 태양열 소독 등을 이용한 물리적인 방제의 적용을 고려할 수 있으나, 비용이 많이 소요될 뿐만 아니라 작기 중에는 불가능하다. Therefore, it is possible to consider the application of physical control using soil improvement, fresh water warming treatment, and solar thermal disinfection as a method of replacing chemical control method, but it is not only expensive but also impossible in small time.
또한, 윤작이나 전답윤환 등과 같은 경종적 방제법이 사용되고 있으나, 재배시기 조절 및 인력과 시간 등의 제약으로 실질적인 사용은 제한적일 수 밖에 없다.In addition, there are various kinds of anti-segregation measures such as crop rotation and anti-seismic measures, but their practical use is limited due to restrictions on manure and time.
최근, 친환경적인 방제에 관한 지속적인 연구와 관심이 높아지면서 곰팡이, 세균, 곤충, 포식성 선충 등 천적을 이용하여 생물학적으로 뿌리혹선충의 방제를 시도하고 있다. 대표적으로 미생물을 이용한 생물학적 선충방제에 Pseudomonas 속, Bacillus 속, Clostridium 속, Desulfovibrio 속, Serratia 속, Streptomyces 속 등이 이용되고 있다.In recent years, as research and interest in environmentally friendly control have been heightened, biologically root-knot nematodes have been attempted using nematodes such as fungi, bacteria, insects and predatory nematodes. Typically, Pseudomonas spp., Bacillus spp., Clostridium spp., Desulfovibrio spp., Serratia spp., Streptomyces spp. Are used for biological nematode control using microorganisms.
이러한 미생물을 이용한 생물학적 방제법은 뿌리혹선충 방제에 사용되고 있는 대부분의 미생물들이 생산한 특정 효소나 살선충 물질 등이 뿌리혹선충의 표피 성분인 콜라겐(Collagen), 난낭 성분인 젤라틴(Gelatin), 알 껍질 성분인 키틴(Chitin)을 분해하여 유충 치사와 알 부화를 억제하는 것으로 보고되고 있다.In this biological control method using microorganisms, specific enzymes and nematode substances produced by most microorganisms used in root nodule nematode control are classified into collagen (collagen), gelatin (gelatin), egg shell It has been reported that decomposing chitin inhibits larval mortality and egg hatching.
Bacillus cereus가 생산한 단백질 분해효소인 protease는 M. javanica 유충의 표피를 분해했으며, B. ambifaria의 배양액 내의 protease와 키틴 분해효소인 chitinase에 의해 선충 유충 치사 및 알 부화가 억제되었다. 또한 Yoon et al. (2012)에 따르면 Streptomyces cacaoi GY525가 생성한 3-benzyl-1, 4-diaza-2, 5-dioxobicyclo[4.3.0]nonane가 M. incognita의 유충 치사 및 알 부화 억제에 활성을 보였다. Bacillus The protease, a protease produced by cereus , degraded the epidermis of M. javanica larvae. The protease in the culture medium of B. ambifaria and the chitinase, chitinase, inhibited lethal transmission and egg hatching. Yoon et al. (2012) According to Streptomyces 3-benzyl-1,4-diaza-2, 5-dioxobicyclo [4.3.0] nonane produced by cacaoi GY525 showed activity in the larval mortality and egg hatching inhibition of M. incognita .
리소박터(Lysobacter sp.)는 대표적인 길항미생물로서 활성을 가진 그람 음성(Gram 陰性) 박테리아이며, 다양한 용균효소(Lytic Enzymes)와 항생물질을 분비하여 식물 병원성 곰팡이 및 식물 해충을 억제하고, 식물생장 호르몬을 생성하여 작물의 생장을 촉진시키는 것으로 알려져 있다. Lysobacter sp. Is a gram-negative (Gram-negative) bacterium that is active as a typical antagonistic microorganism and secretes various lytic enzymes and antibiotics to inhibit phytopathogenic fungi and plant insects, To promote the growth of crops.
L. antibioticus HS124는 다양한 2차 대사산물을 생성하며 Phytophthora capsici, Fusarium oxysporum lycopesici , Pythium aphanidermatum 및 Rhizoctonia solani의 생장을 억제할 뿐만 아니라 배추좀나방 유충을 치사한다. L. antibioticus HS124 produces a variety of secondary metabolites , including Phytophthora capsici, Fusarium oxysporum lycopesici , Pythium It inhibits the growth of aphanidermatum and Rhizoctonia solani as well as lethal larvae of cabbage.
Chen et al. (2006)에 따르면 L. enzymogenes strain C3는 H. schachtii의 알 부화를 억제시키고 유충을 치사한다. 따라서 이러한 기능성 미생물을 이용한 친환경적인 선충방제제의 개발이 요구되고 있는 실정이다.
Chen et al. (2006), L. enzymogenes strain C3 inhibits egg hatching of H. schachtii and lethal larvae. Therefore, it is required to develop an environmentally friendly nematode controlling agent using such a functional microorganism.
상술한 문제를 해결하기 위한 본 발명의 목적은 게껍질이 풍부한 토양으로부터 분리한 Lysobacter capsici YS1215의 생육 및 효소 활성 특성을 이용하여 선충의 유충 치사와 알 부화 억제가 가능한 Lysobacter capsici YS1215를 이용한 선충 방제제를 제공하는데 있다.The object of the present invention to overcome the above-mentioned problems is to provide a method for producing Lysobacter capsici Using YS1215 growth and enzyme activity, Lysobacter capsici And to provide a nematode controlling agent using YS1215.
또한, 본 발명의 다른 목적은 게껍질이 풍부한 토양으로부터 분리한 Lysobacter capsici YS1215의 생육 및 효소 활성 특성을 이용하여 선충의 유충 치사와 알 부화 억제가 가능할 뿐만 아니라 식물생장 호르몬을 생성하여 작물의 생장을 촉진시킬 수 있는 식물 생장 촉진제 및 선충 방제제를 제공하는데 있다.
Another object of the present invention is to utilize the growth and enzymatic activity characteristics of Lysobacter capsici YS1215 isolated from crab-rich soil to inhibit larval mortality and egg hatching of nematodes, as well as to produce plant growth hormone, And to provide a plant growth promoting agent and a nematode controlling agent which can be accelerated.
상술한 목적을 달성하기 위한 본 발명에 따른 식물 생장 촉진제 및 선충 방제제는 게껍질이 포함된 토양으로부터 분리한 Lysobacter capsici YS1215를 이용한 것을 특징으로 한다.In order to achieve the above-mentioned object, the plant growth promoting agent and nematocide of the present invention can be obtained by extracting Lysobacter capsici YS1215 is used.
이때, 본 발명에 따른 식물 생장 촉진제 및 선충 방제제의 게껍질이 포함된 토양으로부터 분리한 Lysobacter capsici YS1215는, 게껍질이 풍부한 토양을 채취하고, 증류수에 현탁 및 멸균수로 희석하여 토양으로부터 분리하는 것을 특징으로 한다.In this case, the plant growth promoter and the nematocide of the present invention, Lysobacter isolated from the soil containing crab shell capsici YS1215 is characterized in that the soil rich in crab shell is collected and suspended in distilled water and diluted with sterilized water and separated from the soil.
또한, 본 발명에 따른 식물 생장 촉진제 및 선충 방제제의 게껍질이 포함된 토양으로부터 분리한 Lysobacter capsici YS1215는, 키틴아제(Chitinase)의 활성을 측정하기 위한 키틴 아가 배지(Chitin agar medium)의 조성이, 0.5% colloidal chitin, 0.2% Na2HPO4, 0.1% KH2PO4, 0.5% NaCl, 0.1% NH4Cl, 0.05% MgSO4ㆍ7H2O, 0.05% CaCl2ㆍ2H2O, 0.05% 효모추출물(Yeast Extract) 및 2% agar 인 것을 특징으로 한다.The plant growth promoter according to the present invention and Lysobacter isolated from the soil containing the crab shell of the nematocide capsici YS1215 showed that the composition of chitin agar medium for measuring the activity of chitinase was 0.5% colloidal chitin, 0.2% Na 2 HPO 4 , 0.1% KH 2 PO 4 , 0.5% NaCl, 0.1 % NH 4 Cl, 0.05% MgSO 4 .7H 2 O, 0.05% CaCl 2 .2H 2 O, 0.05% Yeast Extract and 2% agar.
또한, 본 발명에 따른 식물 생장 촉진제 및 선충 방제제의 게껍질이 포함된 토양으로부터 분리한 Lysobacter capsici YS1215는, 젤라틴아제(Gelatinase) 활성을 측정하기 위한 젤라틴 아가 배지(Gelatin agar medium) 조성이, 1.6 % gelatin, 2.5% Tryptic soy broth 및 2% agar 인 것을 특징으로 한다.The plant growth promoter according to the present invention and Lysobacter isolated from the soil containing the crab shell of the nematocide capsici YS1215 is characterized in that the gelatin agar medium composition for measuring the activity of gelatinase is 1.6% gelatin, 2.5% Tryptic soy broth and 2% agar.
또한, 본 발명에 따른 식물 생장 촉진제 및 선충 방제제의 게껍질이 포함된 토양으로부터 분리한 Lysobacter capsici YS1215는, 미생물 배양용 배지의 조성이, 0.1% 게껍질 분말, 0.1% 젤라틴 분말, 0.3% 복합비료(N:P:K; 21:17:17), 0.3% 설탕, 0.003% 효모추출물(Yeast Extract) 및 0.003% FeCl3ㆍ6H2O 인 것을 특징으로 한다.The plant growth promoter according to the present invention and Lysobacter isolated from the soil containing the crab shell of the nematocide capsici YS1215 was prepared by mixing 0.1% crab shell powder, 0.1% gelatin powder, 0.3% compound fertilizer (N: P: K; 21:17:17), 0.3% sugar, 0.003% yeast extract Extract) and 0.003% FeCl 3 .6H 2 O. [
또한, 상술한 목적을 달성하기 위한 본 발명에 따른 게껍질이 포함된 토양으로부터 분리한 Lysobacter capsici YS1215를 이용한 식물 생장 촉진제 및 선충 방제제의 제조방법은, 게껍질이 포함된 토양을 채취하고, 증류수에 현탁 및 멸균수로 희석하여 Lysobacter capsici YS1215를 토양으로부터 분리하는 단계; 상기 분리된 Lysobacter capsici YS1215이 포함된 희석액을 키틴 아가 배지(Chitin agar medium)에 도말 후 배양하는 단계; 미생물 배양용 배지를 만든 후 120℃에서 15분간 멸균하는 단계; 상기 멸균한 미생물 배양용 배지를 식힌 후, 상기 배양된 Lysobacter capsici YS1215를 접종하는 단계; 상기 Lysobacter capsici YS1215 접종 배지를 30℃에서 5일간 배양하는 단계;로 이루어지는 것을 특징으로 한다. In order to achieve the above object, the present invention provides a method for producing Lysobacter capsici The plant growth promoter and the nematocide preparation method using YS1215 are obtained by collecting the soil containing the crab shell, diluting the suspension with distilled water and sterilized water to obtain Lysobacter capsici Separating YS1215 from the soil; The isolated Lysobacter capsici Culturing the diluted solution containing YS1215 in a chitin agar medium and then culturing; Sterilizing the microorganism culture medium at 120 DEG C for 15 minutes; After cooling the sterilized microorganism culture medium, the cultured Lysobacter capsici Inoculating YS1215; The Lysobacter capsici And culturing the YS1215 inoculation medium at 30 DEG C for 5 days.
이때, 본 발명에 따른 게껍질이 포함된 토양으로부터 분리한 Lysobacter capsici YS1215를 이용한 식물 생장 촉진제 및 선충 방제제의 제조방법에 있어서 상기 키틴 아가 배지는, 0.5% colloidal chitin, 0.2% Na2HPO4, 0.1% KH2PO4, 0.5% NaCl, 0.1% NH4Cl, 0.05% MgSO4ㆍ7H2O, 0.05% CaCl2ㆍ2H2O, 0.05% 효모추출물(Yeast Extract) 및 2% agar를 포함하는 것을 특징으로 한다.Herein, Lysobacter isolated from the soil containing crab shell according to the present invention capsici The chitin agar medium in the plant growth promoting agent and preparation method of nematode control agents using YS1215 is, 0.5% colloidal chitin, 0.2
또한, 본 발명에 따른 게껍질이 포함된 토양으로부터 분리한 Lysobacter capsici YS1215를 이용한 식물 생장 촉진제 및 선충 방제제의 제조방법에 있어서 상기 미생물 배양용 배지는, 0.1% 게껍질 분말, 0.1% 젤라틴 분말, 0.3% 복합비료(N:P:K; 21:17:17), 0.3% 설탕, 0.003% 효모추출물(Yeast Extract) 및 0.003% FeCl3ㆍ6H2O 인 것을 특징으로 한다.
In addition, in the method for producing a plant growth promoting agent and a nematode controlling agent using Lysobacter capsici YS1215 isolated from a crab-containing soil according to the present invention, the culture medium for microbial culture is 0.1% crab shell powder, 0.1% gelatin powder, 0.3% complex fertilizer (N: P: K; 21:17:17), 0.3% sugar, 0.003% Yeast Extract and 0.003% FeCl 3 .6H 2 O.
본 발명은 식물 생장 촉진제 및 선충 방제제에 있어서, 다양한 분해효소를 생성하는 Lysobacter capsici YS1215를 게껍질이 풍부하게 포함된 토양으로부터 분리하여 이용함으로써, 뿌리혹선충의 유충 치사와 알 부화 억제가 가능한 효과가 있다. The present invention relates to a plant growth promoter and the nematode control, Lysobacter generating various lyase capsici By separating YS1215 from the soil rich in crab shell, it is possible to suppress larval mortality and egg hatching of root-knot nematodes.
또한, 본 발명은 선충의 유충 치사와 알 부화 억제가 가능할 뿐만 아니라 식물생장 호르몬을 생성하여 작물의 생장을 촉진시킬 수 있는 효과가 있다.
In addition, the present invention is not only capable of lethal transmission of the nematodes and egg hatching, but also has the effect of promoting the growth of crops by generating plant growth hormone.
도 1은 L. capsici YS1215의 키틴아제 및 젤라틴아제 활성을 측정한 사진을 나타낸 도면,
도 2는 생육배지를 이용하여 측정한 L. capsici YS1215의 세포생장(Cell growth) 및 활성 측정 그래프,
도 3a 및 도 3b는 L. capsici YS1215 배양액 처리구별 지상부 생체중(Fresh shoot weight) 및 지상부 건조중(Dry shoot weight) 조사 그래프를 각각 나타낸 도면,
도 4는 L. capsici YS1215 배양액 처리구별 지상부 길이(Fresh shoot length) 조사 그래프,
도 5a 및 도 5b는 L. capsici YS1215 배양액 처리구별 난낭수(Egg mass number) 조사 그래프,
도 6은 L. capsici YS1215 배양액 처리구별 토양내 유충수 조사 그래프.1 is L. capsici A photograph showing the chitinase activity and gelatinase activity of YS1215,
FIG. 2 is a graph showing the cell growth and activity of L. capsici YS1215 measured using a growth medium,
FIGS. 3A and 3B are graphs showing fresh shoot weight and dry shoot weight of the L. capsici YS1215 culture medium, respectively,
FIG. 4 is a graph showing the fresh shoot length of the L. capsici YS1215 medium treated,
5A and 5B are graphs showing Egg mass number of L. capsici YS1215 culture treated,
FIG. 6 is a graph showing a survey on the water content of soils in L. capsici YS1215 treated cultures.
이하 본 발명의 바람직한 실시 예들의 상세한 설명이 첨부된 도면들을 참조하여 설명될 것이다. 도면들 중 동일한 구성들은 가능한 한 어느 곳에서든지 동일한 부호들을 나타내고 있음을 유의하여야 한다. 하기 설명에서 구체적인 특정 사항들이 나타나고 있는데, 이는 본 발명의 보다 전반적인 이해를 돕기 위해 제공된 것이다. 그리고 본 발명을 설명함에 있어, 관련된 공지 기능 혹은 구성에 대한 구체적인 설명이 본 발명의 요지를 불필요하게 흐릴 수 있다고 판단되는 경우 그 상세한 설명을 생략한다. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS Hereinafter, a detailed description of preferred embodiments of the present invention will be given with reference to the accompanying drawings. It should be noted that the same configurations of the drawings denote the same reference numerals as possible whenever possible. Specific details are set forth in the following description, which is provided to provide a more thorough understanding of the present invention. In the following description of the present invention, detailed description of known functions and configurations incorporated herein will be omitted when it may make the subject matter of the present invention rather unclear.
본 발명에 따른 게껍질이 포함된 토양으로부터 분리한 Lysobacter capsici YS1215를 이용한 식물 생장 촉진제 및 선충 방제제의 효과에 대한 검증을 위하여, 게껍질이 풍부한 토양으로부터 분리한 Lysobacter capsici YS1215의 생육 및 효소 활성 특성과 식물체 실험을 다음과 같이 수행하였다.
The Lysobacter isolated from the soil containing the crab shell according to the present invention capsici In order to examine the effect of plant growth promoter and nematode controlling agent using YS1215, Lysobacter isolated from crab shell rich soil capsici Growth and enzyme activity of YS1215 and plant experiments were performed as follows.
1단계: 토양으로부터 Lysobacter capsici YS1215 분리Stage 1: Lysobacter from soil capsici YS1215 Isolation
먼저, 본 발명에 따른 게껍질이 포함된 토양으로부터 분리한 Lysobacter capsici YS1215를 이용한 식물 생장 촉진제 및 선충 방제제는 토양으로부터 Lysobacter capsici YS1215 분리하기 위하여, 게껍질이 풍부한 순천 해안가 토양을 채취하여, 채취한 토양 10g을 증류수 90mL에 현탁하고, 이 현탁 액을 10-6까지 멸균수로 희석하였다.
First, Lysobacter isolated from the soil containing crab shell according to the present invention capsici Plant growth promoters using YS1215 and nematode control agent is to separate Lysobacter capsici YS1215 from the soil, crabs and shells collected a wealth of Suncheon coastal soil, a soil suspending 10g collected in distilled water 90mL, and to this suspension solution 10-6 And diluted with sterilized water.
2단계: 뿌리혹선충 증식 및 분리Step 2: Propagation and separation of root-knot nematodes
2010년 전남 장성군의 뿌리혹선충 피해가 심한 농가에서 감염 토마토 뿌리를 가져와 토마토(Lycopersicon esculentum) 모종에 감염 및 증식시켜 실험에 사용하였다. 선충 알 및 유충 분리는 선충에 감염된 토마토 뿌리를 흐르는 물에 잘 씻어서 1 cm 크기로 잘라 0.5% NaOCl이 들어있는 병에 넣고 진탕배양기(140rpm)에 15분간 교반한 뒤 45μm 체와 25μm 체에 부어 흐르는 물에 세척하며 알을 수거하였다. 또한, 변형된 Bearman법을 이용하여 분리한 알을 25μm 체에 올린 후, 알을 부화시켜 2령 유충을 수거하였다. 수거한 알과 유충을 실험에 사용하였다.
In 2010 a severe root-knot nematode damage farmers jangseong county of Jeonnam get the tomato roots infected tomato (Lycopersicon esculentum ) seedlings were infected and proliferated. The nematode eggs and larvae were washed in running water and washed in running water, and then they were cut into 1 cm size, placed in a bottle containing 0.5% NaOCl, stirred in a shaking incubator (140 rpm) for 15 minutes and poured into a 45 μm sieve and a 25 μm sieve The eggs were washed with water and collected. In addition, the separated eggs were transferred to a 25 μm sieve using the modified Bearman method, and eggs were hatched to collect 2nd instar larvae. Collected eggs and larvae were used for the experiment.
3단계: YS1215의 효소 활성Step 3: Enzyme activity of YS1215
L. capsici YS1215의 키틴아제(Chitinase) 및 젤라틴아제(Gelatinase) 활성을 측정하기 위해서 키틴 아가 배지(Chitin agar medium) 및 젤라틴 아가 배지(Gelatin agar medium) 100mL를 250mL 삼각플라스크에 넣고 120℃에서 15분간 멸균한 후 각각 페트리 접시(Petri-dish)에 분주하였다. L. capsici To measure the activity of chitinase and gelatinase of YS1215, 100 mL of chitin agar medium and gelatin agar medium was placed in a 250 mL Erlenmeyer flask and sterilized at 120 ° C for 15 minutes Each was dispensed into a Petri dish.
키틴 아가 배지 조성은 0.5% colloidal chitin, 0.2% Na2HPO4, 0.1% KH2PO4, 0.5% NaCl, 0.1% NH4Cl, 0.05% MgSO4ㆍ7H2O, 0.05% CaCl2ㆍ2H2O, 0.05% 효모추출물(Yeast Extract) 및 2% agar 로 하였고, 젤라틴 아가 배지 조성은 1.6 % gelatin, 2.5% Tryptic soy broth 및 2% agar 로 하였다.The composition of the chitin agar medium was 0.5% colloidal chitin, 0.2% Na 2 HPO 4 , 0.1% KH 2 PO 4 , 0.5% NaCl, 0.1% NH 4 Cl, 0.05% MgSO 4揃 7H 2 O, 0.05% CaCl 2揃 2H 2 O, 0.05% yeast extract and 2% agar. Gelatin agar medium composition was 1.6% gelatin, 2.5% Tryptic soy broth and 2% agar.
배지를 완전히 굳힌 후 YS1215를 각각의 배지에 획선(Streaking)을 긋고 30℃에서 3일간 배양하였다. 배양 3일 뒤 키틴아제 활성은 키틴 아가 배지에 투명환이 형성되었을 때 활성이 있는 것으로 간주하고, 젤라틴아제 활성은 젤라틴 아가 배지에 30% 트리클로로아세트산(Trichloroacetic acid)을 3mL 첨가하여 투명환 형성으로 판단하였다.After the medium was completely hardened, YS1215 was streaked on each medium and cultured at 30 DEG C for 3 days. Three days after the incubation, the chitinase activity was considered to be active when a transparent ring was formed in the chitin agar medium. Gelatinase activity was determined by adding 3 mL of 30% trichloroacetic acid to the gelatin agar medium Respectively.
도 1은 L. capsici YS1215의 키틴아제 및 젤라틴아제 활성을 측정한 사진을 나타낸 도면으로서, 키틴 아가 배지 및 젤라틴 아가 배지를 각각 사용하여 L. capsici YS1215의 키틴아제 및 젤라틴아제 활성을 조사한 결과, 도 1의 (a)에 보인 바와 같이, 키틴아제 활성은 키틴 아가 배지에서 투명환이 관찰됨으로 활성이 확인되었고, 젤라틴아제 활성은 도 1의 (b)에 보인 바와 같이, 젤라틴 아가 배지에 30% 트리클로로아세트산(Trichloroacetic acid)을 첨가한 후 투명환이 관찰되어 젤라틴아제 활성을 확인하였다. Park et al.(2008)에 따르면 Lysobacter capsici YS1215가 키틴 분해 활성이 있는 것으로 조사되었다.
1 is L. capsici The chitinase activity and gelatinase activity of L. capsici YS1215 were examined using chitin agar medium and gelatin agar medium, respectively. As a result, chitinase activity and gelatinase activity of YS1215 were measured. As a result, As shown in FIG. 1 (b), the chitinase activity was confirmed by observing the transparent ring in the chitin agar medium and the activity of the gelatinase activity was confirmed by adding 30% trichloroacetic acid to the gelatin agar medium After addition, a transparent ring was observed to confirm the gelatinase activity. According to Park et al. (2008), Lysobacter capsici YS1215 was found to have chitinolytic activity.
4단계: YS1215의 세포생장(Cell growth)Step 4: Cell growth of YS1215
L. capsici YS1215의 세포생장을 측정하기 위해 250mL 삼각플라스크에 생육배지를 넣고 120℃에서 15분간 멸균한 후 YS1215를 접종하고 이를 30℃에서 8일간 배양하면서, 24시간 간격으로 8일간 취하여 UV-spectrophotometer (Shimadzu)를 이용하여 600 nm에서 흡광도를 측정하였다. In order to measure the cell growth of L. capsici YS1215, the growth medium was placed in a 250-mL Erlenmeyer flask, sterilized at 120 ° C for 15 minutes, inoculated with YS1215, cultured at 30 ° C for 8 days, (Shimadzu) was used to measure the absorbance at 600 nm.
세포생장 관찰을 위한 생육배지(미생물 배양용 배지)의 조성은 0.1% 게껍질 분말(삼성키토산, 한국), 0.1% 젤라틴 분말(젤텍, 한국), 0.3% 복합비료(N:P:K; 21:17:17; 남해화학, 한국), 0.3% 설탕(백설, 한국), 0.003% 효모추출물(Yeast Extract) 및 0.003% FeCl3ㆍ6H2O 로 하였다.The composition of the growth medium (culture medium for microbial culture) for cell growth was 0.1% gelatin powder (Samsung Chitosan, Korea), 0.1% gelatin powder (Geltech, Korea), 0.3% compound fertilizer (N: P: K; , Yeast Extract (0.003%), and 0.003% FeCl 3 .6H 2 O (0.3%).
도 2는 생육배지를 이용하여 측정한 L. capsici YS1215의 세포생장(Cell growth) 및 활성 측정 그래프로서, L. capsici YS1215의 세포생장을 측정하기 위해 생육배지를 이용하여 1일부터 8일까지 조사하였다. 조사결과, 도 2에 도시된 바와 같이 1일째부터 6일째까지 증가하다 7일째부터 서서히 감소하였다. 도 2의 (-○-)가 세포생장을 나타낸 그래프이다.
FIG. 2 is a graph showing the cell growth and activity of L. capsici YS1215 measured using a growth medium. As a graph, cell growth of L. capsici YS1215 was measured using a growth medium from
5단계: YS12154의 날짜별 효소활성Step 5: Enzyme activity of YS12154 by date
L. capsici YS1215의 배양 시간에 따른 키틴아제 및 젤라틴아제 활성을 측정하기 위하여 YS1215를 생육 배지에서 30℃에서 8일간 배양하면서 24시간 간격으로 시료를 채취하여 원심 분리(12,000rpm)한 배양 상등액을 사용하였다. To measure the activity of chitinase and gelatinase according to the incubation time of L. capsici YS1215, YS1215 was cultured in growth medium at 30 ° C for 8 days, and samples were taken at intervals of 24 hours and centrifuged (12,000 rpm) Respectively.
키틴아제 활성 측정을 위해 원심 분리한 상등액에 0.5% colloidal chitin을 넣고 37℃에서 한 시간 반응시킨다. 반응 후 1N NaOH를 가하고 반응을 정지시킨 후 12,000rpm으로 원심 분리하여 얻은 상등액을 취하여 Schales' reagent로 발색시켜 420 nm에서 측정하였다. For determination of chitinase activity, 0.5% colloidal chitin was added to the supernatant which was centrifuged, and the mixture was reacted at 37 ° C for one hour. After the reaction, 1N NaOH was added and the reaction was stopped. The supernatant was centrifuged at 12,000 rpm, and the supernatant was developed with Schales' reagent and measured at 420 nm.
젤라틴아제 활성 측정은 gelatin(sigma)을 기질로 사용하여 37℃에서 1시간 반응시킨 다음 반응 전 후의 free α-amino group을 Ninhydrin법에 따라 570nm에서 흡광도를 측정한 후 L-leucine에 대한 표준 검량선으로부터 구하였다.Gelatinase activity was determined by gelatin (Sigma) as a substrate and reacted at 37 ° C for 1 hour. The free α-amino group before and after the reaction was measured by absorbance at 570 nm according to the Ninhydrin method. Respectively.
도 2의 (-■-) 및 (-▲-)로 표시된 그래프를 참조하면, YS1215의 배양 시간에 따른 키틴아제 활성은 3일째 4.0Unit/ml까지 급격히 증가하여 최고의 활성을 나타내었고, 4일과 5일째는 약간 감소하며, 6일째부터 2.5Unit/ml까지 급격히 감소하였다. Referring to the graphs indicated by (-? -) and (-? -) in FIG. 2, the chitinase activity of YS1215 rapidly increased to 4.0 Unit / ml on
또한 젤라틴아제 활성은 2일째까지 빠르게 증가하다가, 3일부터 5일까지 조금씩 증가하여 5일째에 7.43Unit/ml로 최고활성을 보였으며, 6일째부터는 서서히 감소하였다. Gelatinase activity increased rapidly from
이로부터, YS1215 균주는 배양액 내의 균체수가 어느 정도 증가할 때 키틴아제 활성이 증가하지만 일정 시간 경과 후에는 균체의 수와 상관없이 키틴아제 활성이 감소하는 것으로 보인다. 또한, 젤라틴아제 활성은 균체가 증가함에 따라 젤라틴아제 활성도 증가하는 것으로 보인다.
From these results, it was shown that the activity of chitinase was increased when the number of cells in the culture solution was increased to some extent, but the activity of chitinase was decreased after a certain time regardless of the number of cells. In addition, the activity of gelatinase activity seems to increase with increasing cell size.
6단계: YS1215 배양액 처리가 식물생장에 미치는 영향Step 6: Effect of YS1215 medium treatment on plant growth
L. capsici YS1215 배양액의 식물생장 및 선충 감염 억제효과 조사를 위해 발아 후 4주된 토마토를 포트(10cm x 15cm)에 옮겨 심고 대략 253마리의 유충과 199여개의 알을 접종하였다. 선충 접종 후 1주 간격으로 8주차까지 시험구에 미생물 배양액(BM; bacterial medium), 미생물 배양액 반량(HBC; Half of bacterial culture, 미생물 배양액 : 물 = 1 : 1), 배지액(BC; bacterial culture) 및 살선충 농약을 각각 50ml씩 처리(CN; commercial nematicide)하고, 무처리구(TW; Tap water)는 물만 50ml 처리하였다. 이 때, 농약처리구(CN)는 테라노바(abamectin 1.68%, 신젠타 코리아(주))를 물에 녹여 선충 감염 직후 1회 처리하였다. 처리 5주 및 9주째, 각 처리구별로 식물 생장 조사항목인 지상부 길이, 지상부 생체중 및 건물중과 선충 억제 효과 조사 항목인 뿌리혹수, 난낭 수, 토양내 유충수를 측정하였다. To investigate the inhibitory effect of L. capsici YS1215 on plant growth and nematode infection, 4 weeks old tomatoes were transferred to pots (10cm x 15cm) and inoculated with approximately 253 larvae and 199 eggs. Bacterial medium (BM), Half of bacterial culture (HBC), Water (1: 1), Bacterial culture (BC) ) And pesticide pesticide were treated with CN (commercial nematicide), respectively, and TW (tap water) was treated with only 50 ml of water. At this time, terenovar (abomectin 1.68%, Syngenta Korea) was dissolved in water and treated once with nematode infectious agent (CN) immediately after nematode infection. On the 5th and 9th week of treatment, the lengths of shoots, root length, fresh weight of the ground, and plant and nematode inhibitory effects were investigated.
도 3a 및 도 3b는 L. capsici YS1215 배양액 처리구별 지상부 생체중(Fresh shoot weight) 및 지상부 건조중(Dry shoot weight) 조사 그래프를 각각 나타낸 도면으로서, L. capsici YS1215 배양액의 식물생장 및 선충 감염 억제효과를 조사하기 위해 발아 후 4주된 토마토를 이용하여 선충 접종 후 5주 및 9주째에 조사하였다. 3a and 3b L. capsici YS1215 culture process distinct shoot fresh weight (Fresh shoot weight) and the above-ground dry in a diagram illustrating the irradiation graph (Dry shoot weight), respectively, L. capsici plants YS1215 culture growth and nematode infection inhibition effect Were investigated at 5 and 9 weeks after inoculation with 4 - week - old tomatoes after germination.
먼저, 도 3a를 참조하면, 정식 후 5주 및 9주째 지상부 생체중과 지상부 건조중을 조사한 결과, 5주째 식물 지상부 생체중 및 건조중에서 미생물 배양액 반량구(HBC)에서 가장 높게 나타났다. 9주째 생체중은 미생물 배양액(BM), 미생물 배양액 반량구(HBC), 배지액(BC), 물(TW) 및 농약처리구(CN)에서 각각 8.07g, 8.62g, 8.83g, 6.85g 및 7.82g을 나타냈다. First, as shown in FIG. 3A, at the 5th week and 9th week after the formalinization, the fresh weight and the dry weight at the top of the ground were the highest in the microbial culture medium (HBC) at the 5th week. At 9 weeks, the fresh weight was 8.07 g, 8.62 g, 8.83 g, 6.85 g and 7.82 g in the microbial culture (BM), microbial culture medium (HBC), medium (BC), water (TW) Respectively.
다음으로, 도 3b를 참조하면, 9주째 건물중은 미생물 배양액(BM), 미생물 배양액 반량구(HBC), 배지액(BC), 물(TW) 및 농약처리구(CN)에서 각각 0.79g, 0.81g, 0.62g, 0.82g 및 0.72g을 나타냈다. 하지만 생체중 및 건물중의 미생물 배양액(BM), 미생물 배양액 반량구(HBC) 및 배지액 처리구(BC) 간의 유의적 차이는 나타나지 않았고, 미생물 배양액 처리구(BM)와 물 처리구(TW)간의 유의성은 나타났다. Next, referring to FIG. 3B, in the 9th week, 0.79 g and 0.81 in the microbial culture (BM), microbial culture medium (HBC), medium (BC), water (TW) g, 0.62 g, 0.82 g and 0.72 g, respectively. However, there was no significant difference between fresh and dry microbial cultures (BM), microbial culture medium (HBC) and culture broth (BC), and there was a significant difference between microbial culture medium treatment (BM) and water treatment (TW) .
도 4는 L. capsici YS1215 배양액 처리구별 지상부 길이(Fresh shoot length) 조사 그래프로서 도 4를 참조하면, 9주째 지상부 길이(Fresh shoot length)는 미생물 배양액(BM), 미생물 배양액 반량구(HBC), 배지액(BC), 물(TW) 및 농약처리구(CN)에서 각각 35.00cm, 43.06cm, 36.50cm, 37.66cm, 38.66cm로 나타났다. FIG. 4 is a graph showing the fresh shoot length of the L. capsici YS1215 treated medium. As shown in FIG. 4, the fresh shoot length of the 9th week was measured by using microbial culture medium (BM), microbial culture medium (HBC) 43.06cm, 36.50cm, 37.66cm, and 38.66cm respectively in the medium (BC), water (TW) and pesticide treatment (CN)
이러한 결과는 미생물 배양액(BM), 미생물 배양액 반량구(HBC) 및 배지액(BC) 등이 비료로서의 효과는 비슷한 것으로 보인다. Broadbent et al.(1977)에 따르면 미생물(Bacillus spp.)이 뿌리혹선충의 감염을 억제하였거나 생리활성 물질을 분비함으로써 식물이 생장하는 것으로 보고하였으나 본 실험결과와는 상반되었다.These results suggest that the effects of microbial culture (BM), microbial culture medium (HBC) and culture medium (BC) as fertilizer are similar. Broadbent et al. (1977) reported that microbes ( Bacillus spp.) Inhibited root-knot nematode infection or secreted physiologically active substances, which were in contrast to the results of this experiment.
도 5a 및 도 5b는 L. capsici YS1215 배양액 처리구별 난낭수(Egg mass number) 조사 그래프로서, 미생물 배양액(BM), 미생물 배양액 반량구(HBC), 배지액(BC), 물(TW) 및 농약처리구(CN)에 대한 뿌리혹선충에 미치는 영향을 조사한 결과, 도 5a를 참조하면 각 처리구별 난낭수는 39개, 116개, 160개, 275개, 24개로 나타났고, 도 5b를 참조하면 뿌리혹수는 47개, 84개, 137개, 216개, 35로 조사되었다. 5A and 5B are graphs showing Egg mass number of L. capsici YS1215 cultured medium treated with a microorganism culture medium (BM), a microbial culture medium (HBC), a culture medium (BC), water (TW) As shown in FIG. 5A, the number of oocysts of each treatment was 39, 116, 160, 275, and 24, respectively. As shown in FIG. 5B, Were 47, 84, 137, 216, and 35, respectively.
도 6은 L. capsici YS1215 배양액 처리구별 토양내 유충수 조사 그래프로서, 도 6에 도시된 바와 같이, 토양내 유충수는 미생물 배양액(BM), 미생물 배양액 반량구(HBC), 배지액(BC), 물(TW) 및 농약처리구(CN)에서 각각 731마리, 1063마리, 4314마리, 4879마리, 413마리로 조사되었다. FIG. 6 is a graph showing the intrusion of water in the soil according to the treatment of L. capsici YS1215 culture. As shown in FIG. 6, the water in the soil contains the microbial culture medium (BM), microbial culture medium (HBC) (TW) and pesticide treatments (CN) were 731, 1063, 4314, 4879 and 413, respectively.
도 5a, 5b 및 도 6에 나타낸 바와 같이, 난낭수, 뿌리혹수 및 토양내 유충수에 대한 조사결과, 농약처리구(CN)가 가장 선충억제에 효과가 좋은 것으로 조사되었지만, 미생물 배양액 처리구(BM)와의 유의적 차이는 보이지 않았다. As shown in FIGS. 5A, 5B, and 6, the results of investigations on egg yolk, number of rootlets and soil worms indicate that the pesticide treatment (CN) is the most effective for inhibiting nematodes. However, There were no significant differences.
반면에, 미생물 배양액 처리구(BM)와 미생물 배양액 반량구 처리구(HBC)와 유의적 차이를 보였는데, 이는 배양액을 희석하여 사용하면 효과 또한 감소하는 것으로 보여진다. On the other hand, there was a significant difference between the microbial culture medium (BM) and the microbial culture medium (HBC), indicating that the effect of diluting the culture medium is also decreased.
미생물 배양액(BM) 및 반량구(HBC)가 대조구인 배지액 처리구(BC)와 물처리구(TW)보다 난낭수, 뿌리혹수 및 토양내 유충수에서 적게 나타났으며, 통계적 유의성에서도 차이를 보이는데, Devidas and Rehberger(1992)의 보고에 따르면 이러한 결과는 미생물 배양 시 배양액 속에 생성된 여러 2차 대사산물인 효소 및 항생물질이 선충 억제에 활성을 나타낸 것으로 보인다. The microbial culture medium (BM) and the half-medium (HBC) were less in the egg yolk, root and watery soil than in the control medium (BC) and water treatment (TW) and Rehberger (1992) reported that these secondary metabolites, enzymes and antibiotics, produced in the culture medium during the microbial cultivation showed activity in nematode inhibition.
비슷한 예로는 Bacillus subtilis 배양액을 토마토에 처리하였을 때 M. incognita의 난낭수와 뿌리혹수가 감소된 것과, Gautam et al.(1995) 및 Dicklow et al.(1993)의 실험에서 나타난 Streptomyces sp. 배양액이 M. incognita의 뿌리혹 생성을 감소시킨 조사결과가 있다. 또한, S. sampsonii KK1024 배양액을 처리했을 때 토마토의 뿌리혹선충 감염 피해가 감소되었다는 보고도 있다.A similar example is Bacillus subtilis When the cultures were treated with tomato, M. incognita The number of oocysts and shoots decreased, and the number of Streptomyces sp. In the experiment of Gautam et al. (1995) and Dicklow et al. (1993). Studies have shown that the culture medium reduces the incidence of root incidence of M. incognita . In addition, S. It has been reported that the damage of tomato root nodule nematode infection was reduced when the sampsonii KK1024 culture medium was treated.
결과적으로, L. capsici YS1215를 이용한 뿌리혹선충의 생물학적 방제는 가능하며, 미생물 배양액을 희석하지 않고 작물에 적용하였을 때 작물 생육저하를 일으키지 않고 높은 선충억제 효과를 보였으므로 선충 감염이 시작되는 작물 정식 초기 및 선충 번식이 활발한 작물 생육기에도 적용할 수 있다.As a result, biological control of root-knot nematodes using L. capsici YS1215 is possible, and when applied to crops without diluting the microbial culture solution, high nematode inhibitory effect was observed without causing deterioration of crop growth, And also for growing crops with active nematode propagation.
전술한 바와 같이, 본 발명에 따른 게껍질이 포함된 토양으로부터 분리한 Lysobacter capsici YS1215를 이용한 식물 생장 촉진제 및 선충 방제제의 효과를 검증하기 위하여, Lysobacter capsici YS1215의 특성 및 뿌리혹선충 방제에 미치는 영향을 조사한 결과, YS1215의 생육은 배양 6일째 최고였으며, 생육에 따른 키틴아제와 젤라틴아제의 활성은 각각 3일째와 5일째에 가장 높은 활성을 보였다. As described above, the Lysobacter capsici isolated from the soil containing crab skin according to the present invention In order to examine the effect of plant growth promoter and nematocide using YS1215, Lysobacter The effect of chitinase and gelatinase on the growth of YS1215 was highest at the 6th day of cultivation. The activity of chitinase and gelatinase was highest at 3 and 5 days, respectively.
또한, YS1215 배양액이 선충 피해 방제와 식물 생장에 미치는 영향을 조사해 본 결과, 5주째 식물 지상부 생체중 및 건조중에서 배양액 반량구(HBC)에서 가장 높게 나타났지만, 9주째에는 미생물 배양액(BM), 미생물 배양액 반량구(HBC) 및 배지액 처리구(BC)에서 차이를 보이지 않았다. 하지만 9주째 미생물 배양액(BM), 미생물 배양액 반량구(HBC) 및 배지액 처리구(BC)가 물처리구(TW) 보다 높게 나타났다. 한편, 지상부 길이에서는 미생물 배양액 반량구 처리구(HBC)가 가장 높았다. In addition, the effect of YS1215 culture medium on nematode damage control and plant growth was found to be highest in the culture supernatant (HBC) at the 5th week in the fresh weight of the plant and in the dry state. At the 9th week, the microbial culture (BM) (HBC) and medium (BC). However, in the 9th week, the microbial culture (BM), microbial culture medium (HBC) and culture medium (BC) were higher than water treatment (TW). On the other hand, microbial culture medium (HBC) was the highest in the length of the top part.
선충 피해 방제에 있어서 난낭수, 뿌리혹수 및 토양내 유충수에서 각각 농약처리구(CN)에서 가장 낮게 나타났으나, 미생물 배양액 처리구(BM)와의 유의적 차이는 보이지 않았다. 미생물 배양액 처리구(BM)는 미생물 배양액 반량구(HBC) 및 물처리구(TW)와는 유의적 차이가 있는 것으로 조사되었다. In the control of nematode damage, the lowest concentrations were found in the egg yolk, the root and the lily of the soil in the pesticide treatments (CN), but there was no significant difference with the microbial culture treatments (BM). Microbial culture medium (BM) was found to be significantly different from microbial culture medium (HBC) and water treatment (TW).
그러므로 다양한 분해효소를 생성하는 L. capsici YS1215의 뿌리혹선충 방제에 대한 충분한 가능성과 가치가 있으며, 본 발명에 따른 게껍질이 포함된 토양으로부터 분리한 Lysobacter capsici YS1215를 이용한 식물 생장 촉진제 및 선충 방제제의 가치와 효과 또한 충분히 검증된다.Therefore, L. capsici YS1215 has sufficient potential and value for root control nematode control, and Lysobacter isolated from the soil containing crab shell according to the present invention capsici The value and effectiveness of plant growth promoters and nematocides using YS1215 are also verified.
본 발명의 효과 검증을 위한 실험 데이터의 결과는 SAS프로그램 9.1 버전(2006)을 사용하여 5%의 유의수준(P≤0.05)에서 Turkey's Studentized Range Test(Tukey's HSD multiple comparison test)로 분석하였다.The results of the experimental data for the verification of the effect of the present invention were analyzed using the SAS program 9.1 version (2006) and the Turkey's Studentized Range Test (Tukey's HSD multiple comparison test) at a significance level of 5% (P? 0.05).
한편, 본 발명의 상세한 설명에서는 구체적인 실시 예에 관해 설명하였으나, 본 발명의 범위에서 벗어나지 않는 한도 내에서 여러 가지 변형이 가능함은 물론이다. 그러므로 본 발명의 범위는 설명된 실시 예에 국한되어 정해져서는 안되며 후술하는 특허청구의 범위뿐 아니라 이 특허청구의 범위와 균등한 것들에 의해서 정해져야 한다.While the invention has been shown and described with reference to certain preferred embodiments thereof, it will be understood by those skilled in the art that various changes and modifications may be made without departing from the spirit and scope of the invention. Therefore, the scope of the present invention should not be limited by the described embodiments, but should be determined by the scope of the appended claims and equivalents thereof.
Claims (8)
게껍질이 포함된 토양으로부터 분리한 Lysobacter capsici YS1215를 포함하는 것을 특징으로 하는 게껍질이 포함된 토양으로부터 분리한 Lysobacter capsici YS1215를 이용한 식물 생장 촉진제 및 선충 방제제.
In the plant growth promoting agent and the nematode controlling agent,
Lysobacter isolated from soil containing crab shells capsici A crab comprising the YS1215 separated from the shell, including soil Lysobacter capsici Plant Growth Promoter and Nematode Control Agent Using YS1215.
게껍질이 포함된 토양을 채취하고, 증류수에 현탁 및 멸균수로 희석하여 토양으로부터 분리하는 것을 특징으로 하는 게껍질이 포함된 토양으로부터 분리한 Lysobacter capsici YS1215를 이용한 식물 생장 촉진제 및 선충 방제제.
The method according to claim 1, wherein the Lysobacter capsici YS1215,
A plant growth promoting agent and a nematode controlling agent using Lysobacter capsici YS1215 isolated from a soil containing crab shell, characterized in that the soil containing crab shell is collected and suspended in distilled water and diluted with sterilized water and separated from the soil.
키틴아제(Chitinase)의 활성을 측정하기 위한 키틴 아가 배지(Chitin agar medium)의 조성이, 0.5% colloidal chitin, 0.2% Na2HPO4, 0.1% KH2PO4, 0.5% NaCl, 0.1% NH4Cl, 0.05% MgSO4ㆍ7H2O, 0.05% CaCl2ㆍ2H2O, 0.05% 효모추출물(Yeast Extract) 및 2% agar 인 것을 특징으로 하는 게껍질이 포함된 토양으로부터 분리한 Lysobacter capsici YS1215를 이용한 식물 생장 촉진제 및 선충 방제제.
The method according to claim 1, wherein the Lysobacter capsici YS1215,
The composition of the chitin agar medium for measuring the activity of chitinase was 0.5% colloidal chitin, 0.2% Na 2 HPO 4 , 0.1% KH 2 PO 4 , 0.5% NaCl, 0.1% NH 4 Cl, 0.05% MgSO 4 and 7H 2 O, 0.05% CaCl 2 and 2H 2 O, 0.05% of yeast extract (yeast extract) and two isolated from the soil to include a shell, characterized in that% agar Lysobacter capsici Plant Growth Promoter and Nematode Control Agent Using YS1215.
젤라틴아제(Gelatinase) 활성을 측정하기 위한 젤라틴 아가 배지(Gelatin agar medium) 조성이, 1.6 % gelatin, 2.5% Tryptic soy broth 및 2% agar 인 것을 특징으로 하는 게껍질이 포함된 토양으로부터 분리한 Lysobacter capsici YS1215를 이용한 식물 생장 촉진제 및 선충 방제제.
The method according to claim 1, wherein the Lysobacter capsici YS1215,
Characterized in that the gelatin agar medium composition for measuring the activity of gelatinase is 1.6% gelatin, 2.5% Tryptic soy broth and 2% agar. Lysobacter isolated from the shell- capsici Plant Growth Promoter and Nematode Control Agent Using YS1215.
미생물 배양용 배지의 조성이, 0.1% 게껍질 분말, 0.1% 젤라틴 분말, 0.3% 복합비료(N:P:K; 21:17:17), 0.3% 설탕, 0.003% 효모추출물(Yeast Extract) 및 0.003% FeCl3ㆍ6H2O 인 것을 특징으로 하는 게껍질이 포함된 토양으로부터 분리한 Lysobacter capsici YS1215를 이용한 식물 생장 촉진제 및 선충 방제제.
The method according to claim 1, wherein the Lysobacter capsici YS1215,
The composition of the culture medium for microbial culture was 0.1% crab shell powder, 0.1% gelatin powder, 0.3% compound fertilizer (N: P: K; 21:17:17), 0.3% sugar, 0.003% yeast extract, 0.003% FeCl 3 6H and separated from the soil to include a shell, characterized in that the 2 O Lysobacter capsici Plant Growth Promoter and Nematode Control Agent Using YS1215.
게껍질이 포함된 토양을 채취하고, 증류수에 현탁 및 멸균수로 희석하여 Lysobacter capsici YS1215를 토양으로부터 분리하는 단계;
상기 분리된 Lysobacter capsici YS1215이 포함된 희석액을 키틴 아가 배지(Chitin agar medium)에 도말 후 배양하는 단계;
미생물 배양용 배지를 만든 후 120℃에서 15분간 멸균하는 단계;
상기 멸균한 미생물 배양용 배지를 식힌 후, 상기 배양된 Lysobacter capsici YS1215를 접종하는 단계; 및
상기 Lysobacter capsici YS1215 접종 배지를 30℃에서 5일간 배양하는 단계;로 이루어지는 것을 특징으로 하는 게껍질이 포함된 토양으로부터 분리한 Lysobacter capsici YS1215를 이용한 식물 생장 촉진제 및 선충 방제제의 제조방법.
A method for producing a plant growth promoting agent and a nematocide,
The soil containing crab shells was collected and suspended in distilled water and diluted with sterile water to obtain Lysobacter capsici Separating YS1215 from the soil;
The isolated Lysobacter capsici Culturing the diluted solution containing YS1215 in a chitin agar medium and then culturing;
Sterilizing the microorganism culture medium at 120 DEG C for 15 minutes;
Cooling the sterilized microorganism culture medium, inoculating the cultured Lysobacter capsici YS1215; And
The Lysobacter capsici And culturing the YS1215 seeding medium at 30 DEG C for 5 days. The method for producing Lysobacter capsici Plant Growth Promoter Using YS1215 and Method for Producing Nematocide.
0.5% colloidal chitin, 0.2% Na2HPO4, 0.1% KH2PO4, 0.5% NaCl, 0.1% NH4Cl, 0.05% MgSO4ㆍ7H2O, 0.05% CaCl2ㆍ2H2O, 0.05% 효모추출물(Yeast Extract) 및 2% agar를 포함하는 것을 특징으로 하는 게껍질이 포함된 토양으로부터 분리한 Lysobacter capsici YS1215를 이용한 식물 생장 촉진제 및 선충 방제제의 제조방법.
7. The chitin agar medium according to claim 6,
0.5% colloidal chitin, 0.2% Na 2 HPO 4 , 0.1% KH 2 PO 4 , 0.5% NaCl, 0.1% NH 4 Cl, 0.05% MgSO 4揃 7H 2 O, 0.05% CaCl 2揃 2H 2 O, 0.05% extract (Yeast extract) and two isolated from the soil that includes the crab shell, characterized in that includes Lysobacter% agar capsici Plant Growth Promoter Using YS1215 and Method for Producing Nematocide.
0.1% 게껍질 분말, 0.1% 젤라틴 분말, 0.3% 복합비료(N:P:K; 21:17:17), 0.3% 설탕, 0.003% 효모추출물(Yeast Extract) 및 0.003% FeCl3ㆍ6H2O 인 것을 특징으로 하는 게껍질이 포함된 토양으로부터 분리한 Lysobacter capsici YS1215를 이용한 식물 생장 촉진제 및 선충 방제제의 제조방법.7. The microorganism culture medium according to claim 6,
0.1% gelatin powder, 0.3% compound fertilizer (N: P: K; 21:17:17), 0.3% sugar, 0.003% Yeast Extract and 0.003% FeCl 3 .6H 2 O Characterized in that it contains Lysobacter isolated from the soil containing the crab shell capsici Plant Growth Promoter Using YS1215 and Method for Producing Nematocide.
Priority Applications (1)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
KR1020130032401A KR101503916B1 (en) | 2013-03-26 | 2013-03-26 | Plant growth substance and anti-root knot nematode compound and its composition using Lysobacter capsici YS1215 from crab shell amended soil and manufacturing method thereof |
Applications Claiming Priority (1)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
KR1020130032401A KR101503916B1 (en) | 2013-03-26 | 2013-03-26 | Plant growth substance and anti-root knot nematode compound and its composition using Lysobacter capsici YS1215 from crab shell amended soil and manufacturing method thereof |
Publications (2)
Publication Number | Publication Date |
---|---|
KR20140117221A true KR20140117221A (en) | 2014-10-07 |
KR101503916B1 KR101503916B1 (en) | 2015-03-19 |
Family
ID=51990643
Family Applications (1)
Application Number | Title | Priority Date | Filing Date |
---|---|---|---|
KR1020130032401A KR101503916B1 (en) | 2013-03-26 | 2013-03-26 | Plant growth substance and anti-root knot nematode compound and its composition using Lysobacter capsici YS1215 from crab shell amended soil and manufacturing method thereof |
Country Status (1)
Country | Link |
---|---|
KR (1) | KR101503916B1 (en) |
Cited By (2)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
CN111357734A (en) * | 2020-03-17 | 2020-07-03 | 北京市农林科学院 | Method for preventing and controlling root-knot nematodes through shallow-buried ammonia water drip irrigation disinfection |
CN116333895A (en) * | 2023-04-04 | 2023-06-27 | 重庆谷百奥生物研究院有限公司 | Special culture medium for insecticidal fungi without sterilization, production method of insecticidal fungi, and fungi preparation and application |
Families Citing this family (1)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
KR101866814B1 (en) | 2016-09-29 | 2018-06-18 | 전남대학교산학협력단 | Xylaria grammica EL 000614 strain having nematocidal activity against root knot nematode and uses thereof |
Family Cites Families (2)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
US6524998B1 (en) | 1999-03-01 | 2003-02-25 | Auburn University | Biological compositions and methods for enhancing plant growth and health and producing disease-suppressive plants |
KR100757298B1 (en) | 2006-01-09 | 2007-09-11 | 순천대학교 산학협력단 | Antagonistic microorganisms having activity against plant pathogens and plant diseases controlling agent comprising the same |
-
2013
- 2013-03-26 KR KR1020130032401A patent/KR101503916B1/en active IP Right Grant
Cited By (3)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
CN111357734A (en) * | 2020-03-17 | 2020-07-03 | 北京市农林科学院 | Method for preventing and controlling root-knot nematodes through shallow-buried ammonia water drip irrigation disinfection |
CN116333895A (en) * | 2023-04-04 | 2023-06-27 | 重庆谷百奥生物研究院有限公司 | Special culture medium for insecticidal fungi without sterilization, production method of insecticidal fungi, and fungi preparation and application |
CN116333895B (en) * | 2023-04-04 | 2024-03-15 | 重庆谷百奥生物研究院有限公司 | Special culture medium for insecticidal fungi without sterilization, production method of insecticidal fungi, and fungi preparation and application |
Also Published As
Publication number | Publication date |
---|---|
KR101503916B1 (en) | 2015-03-19 |
Similar Documents
Publication | Publication Date | Title |
---|---|---|
Ahmad et al. | Biological control: a novel strategy for the control of the plant parasitic nematodes | |
Padgham et al. | Biological control potential and modes of action of Bacillus megaterium against Meloidogyne graminicola on rice | |
CN106686983B (en) | Use of bacillus methylotrophicus as plant growth stimulator and biological control element, and isolate of said species | |
Leelavathi et al. | Antimicrobial activity of Trichoderma harzianum against bacteria and fungi | |
Kim et al. | An effective biocontrol bioformulation against Phytophthora blight of pepper using growth mixtures of combined chitinolytic bacteria under different field conditions | |
Chen et al. | 20 Biological Control of Nematodes by Fungal Antagonists | |
Duponnois et al. | Beneficial effects of Enterobacter cloacae and Pseudomonas mendocina for biocontrol of Meloidogyne incognita with the endospore-forming bacterium Pasteuria penetrans | |
Solanki et al. | Co-inoculation of different antagonists can enhance the biocontrol activity against Rhizoctonia solani in tomato | |
El-Hadad et al. | In vitro evaluation of some bacterial isolates as biofertilizers and biocontrol agents against the second stage juveniles of Meloidogyne incognita | |
Lahkar et al. | Biosurfactant of Pseudomonas aeruginosa JS29 against Alternaria solani: the causal organism of early blight of tomato | |
Nawaz et al. | Evaluation of antifungal metabolites activity from Bacillus licheniformis OE-04 against Colletotrichum gossypii | |
Yang et al. | Biocontrol activities of bacteria from cowdung against the rice sheath blight pathogen | |
Ben Khedher et al. | Biosurfactant produced by Bacillus subtilis V26: A potential biological control approach for sustainable agriculture development | |
Singh et al. | Drechslerella dactyloides and Dactylaria brochopaga mediated induction of defense related mediator molecules in tomato plants pre-challenged with Meloidogyne incognita | |
Tong-Jian et al. | Bacillus cereus X5 enhanced bio-organic fertilizers effectively control root-knot nematodes (Meloidogyne sp.) | |
Singh et al. | Combined use of Streptomyces sp. A6 and chemical fungicides against fusarium wilt of Cajanus cajan may reduce the dosage of fungicides required in the field | |
Fu et al. | Antifungal and biocontrol evaluation of four Lysobacter strains against clubroot disease | |
Zhang et al. | Pathogenicity of Serratia marcescens to hazelnut weevil (Curculio dieckmanni) | |
KR101503916B1 (en) | Plant growth substance and anti-root knot nematode compound and its composition using Lysobacter capsici YS1215 from crab shell amended soil and manufacturing method thereof | |
Neung et al. | Insecticidal potential of Paenibacillus elgii HOA73 and its combination with organic sulfur pesticide on diamondback moth, Plutella xylostella | |
Gupta et al. | Potential of wax degrading bacteria for management of the citrus mealybug, Planococcus citri | |
Al-Quwaie | The role of Streptomyces species in controlling plant diseases: a comprehensive review | |
Singh et al. | Harnessing biocontrol potential of Trichoderma harzianum for control of Meloidogyne incognita in tomato | |
Anandaraj et al. | The potential of PGPRs in disease management of spice crops | |
Chavarria-Quicaño et al. | Native Bacillus paralicheniformis isolate as a potential agent for phytopathogenic nematodes control |
Legal Events
Date | Code | Title | Description |
---|---|---|---|
A201 | Request for examination | ||
E902 | Notification of reason for refusal | ||
E701 | Decision to grant or registration of patent right | ||
GRNT | Written decision to grant | ||
FPAY | Annual fee payment |
Payment date: 20171226 Year of fee payment: 4 |
|
FPAY | Annual fee payment |
Payment date: 20181226 Year of fee payment: 5 |
|
FPAY | Annual fee payment |
Payment date: 20191231 Year of fee payment: 6 |