KR100523991B1 - Method for preparing biological material having antimicrobial against pathogen and growth stimulator for plant, and for cultivating it thereof - Google Patents
Method for preparing biological material having antimicrobial against pathogen and growth stimulator for plant, and for cultivating it thereof Download PDFInfo
- Publication number
- KR100523991B1 KR100523991B1 KR10-2003-0075077A KR20030075077A KR100523991B1 KR 100523991 B1 KR100523991 B1 KR 100523991B1 KR 20030075077 A KR20030075077 A KR 20030075077A KR 100523991 B1 KR100523991 B1 KR 100523991B1
- Authority
- KR
- South Korea
- Prior art keywords
- present
- plant
- growth
- perilla
- strain
- Prior art date
Links
Classifications
-
- 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
-
- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01G—HORTICULTURE; CULTIVATION OF VEGETABLES, FLOWERS, RICE, FRUIT, VINES, HOPS OR SEAWEED; FORESTRY; WATERING
- A01G2/00—Vegetative propagation
-
- 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
- A01N63/10—Animals; Substances produced thereby or obtained therefrom
-
- 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
- C12N1/205—Bacterial isolates
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12R—INDEXING SCHEME ASSOCIATED WITH SUBCLASSES C12C - C12Q, RELATING TO MICROORGANISMS
- C12R2001/00—Microorganisms ; Processes using microorganisms
- C12R2001/01—Bacteria or Actinomycetales ; using bacteria or Actinomycetales
Abstract
본 발명은 식물의 생물학적 제제의 제조방법 및 이를 이용한 식물재배방법에 관한 것이다.The present invention relates to a method for producing a biological preparation of a plant and a method for plant cultivation using the same.
본 발명은 신규한 균주 브르크호데리아 속 AK-17(Burkhoderia sp.AK-17) KACC91058 균주를 배양하고, 상기 배양물로부터 식물 병원균에 대한 항균활성 및 식물의 생육 촉진 효과를 갖는 생물학적 제제의 제조방법 및 이를 이용한 식물재배방법을 제공한다.The present invention is to cultivate a novel strain of the strain Brkhoderia genus AK-17 ( Burkhoderia sp.AK-17) KACC91058, and to prepare a biological agent having an antimicrobial activity against plant pathogens and promoting the growth of plants from the culture. It provides a method and a plant cultivation method using the same.
본 발명은 식물 병원균에 대한 항균활성 및 식물의 생육을 촉진하는 작용이 있기 때문에, 농약의 잔류독성에 대한 위험성이 없는 고품질 청정 농산물의 생산이 가능하고, 생육 촉진효과로 출하시기를 단축하고, 수확량도 증대시킬 수 있는 효과가 있다.The present invention has the action of promoting the antimicrobial activity and growth of plants against plant pathogens, it is possible to produce high-quality clean agricultural products without the risk of residual toxicity of pesticides, and to shorten the shipment by the growth promoting effect, yield There is also an effect that can be increased.
Description
본 발명은 식물의 생물학적 제제의 제조방법 및 이를 이용한 식물재배방법에 관한 것으로, 보다 상세하게는 신규한 미생물을 이용하여 식물 병원균에 대한 항균 활성 및 식물 생육 촉진의 효과를 갖는 생물학적 제제의 제조방법과 이를 사용한 식물의 재배방법과 이로부터 제조된 생물학적 제제에 관한 것이다.The present invention relates to a method for producing a biological preparation of a plant, and a method for plant cultivation using the same, and more particularly, to a method for preparing a biological preparation having an antimicrobial activity against plant pathogens and promoting plant growth by using a novel microorganism. It relates to a method of cultivating a plant using the same and a biological agent prepared therefrom.
들깨는 동부 아시아지역이 원산지로 추정되고 있으며, 현재는 인도, 소련 및 미국에 걸쳐 세계적으로 널리 분포, 재배되어지고 작물이며, 우리나라에는 통일신라시대 때부터 재배되어 왔다. Perilla is believed to be native to Eastern Asia, and is now widely distributed, grown, and cultivated throughout India, the Soviet Union, and the United States, and has been cultivated in Korea since the Unified Silla Period.
최근 식생활의 변화와 들깻잎의 유효성분들에 대한 생리활성효과가 밝혀짐으로써 들깻잎의 소비가 급증하고 있으며, 전국에 걸쳐 재배면적도 매년 증가하여 재배되고 있는 실정이다. Recent changes in dietary life and the bioactive effects of perilla leaves have been found to increase the consumption of perilla leaves, and the cultivation area is also growing every year across the country.
그러나, 수요의 급증에 따른 안정적 공급을 위해서는 시설재배 등을 이용한 연중재배가 필요하게 되었으며, 화학 비료와 농약의 과다사용이 불가피한 상황이 발생하게 되었다. However, year-round cultivation using facility cultivation is required for stable supply due to a surge in demand, and overuse of chemical fertilizers and pesticides is inevitable.
이러한 문제점은 재배토양과 수질을 오염시키는 원인뿐만 아니라 시설재배에서 농약의 과다사용은 재배환경 열악으로 생산자의 건강을 해치게 되었으며, 생산된 농산물에는 잔류농약으로 식품의 안전성에 심각한 우려를 낳고 있다. This problem not only causes pollution of cultivated soil and water quality, but too much use of pesticides in facility cultivation damages the health of producers due to poor cultivation environment, and produced agricultural products have serious concerns about food safety as residual pesticides.
현재, 농산물에 대한 안전성 문제와 토양환경의 보존을 통한 지속 가능한 농업에 관한 관심으로 유기농업이 전 세계적으로 급격하게 확산되고 있는 실정이다. Currently, organic farming is rapidly spreading around the world due to safety issues for agricultural products and interest in sustainable agriculture through the preservation of soil environment.
이러한 배경 아래 생물학적 방제법을 이용한 안전 농산물을 생산하기 위한 노력으로 다양한 미생물을 농업에 활용하고 있다(Raghavan Charudattan and Amos Dinoor., 2000; Chung, B. K. and Hong, K. S., 1991). Against this backdrop, various microorganisms are used in agriculture in an effort to produce safe agricultural products using biological control methods (Raghavan Charudattan and Amos Dinoor., 2000; Chung, B. K. and Hong, K. S., 1991).
미생물의 이용은 크게 미생물 비료와 미생물 농약으로 농업에 활용할 수 있는데, 미생물농약은 자연계에 존재하는 미생물 그 자체, 또는 미생물이 생산하는 2차대사산물을 이용해 식물병을 억제하는 것으로, 사람이나 가축, 어패류 등에 무해하며 환경오염과 생태계 교란의 우려성이 없다는 특징을 갖고있어 세계적으로 많은 연구가 이루어지고 있다. 따라서, 미생물제제의 사용은, 비료와 농약의 사용을 줄여 토양환경을 지속적으로 보존하고 장기적인 생산성을 확보할 수 있으며, 농작물에 대한 잔류농약의 위험성을 제거해 농산물의 안전성을 확보할 수 있는 일거양득의 효과를 가질 수 있을 것으로 예상된다.The use of microorganisms can be largely used in agriculture as microbial fertilizers and microbial pesticides. Microbial pesticides suppress plant diseases by using microorganisms in nature or secondary metabolites produced by microorganisms. It is harmless to fish and shellfish, and has no characteristic of environmental pollution and disturbance of ecosystem. Therefore, the use of microbial agents can reduce the use of fertilizers and pesticides, thereby conserving the soil environment continuously and ensuring long-term productivity, and eliminating the risk of residual pesticides on crops. It is expected to have an effect.
한편, 잎들깨의 재배에 있어 가장 문제시되는 병충해를 보면, 녹병, 잿빛곰팡이병, 균핵병, 심식충, 응애류, 콩잎마름이 나방, 진딧물을 들 수 있다. 이러한 병충해를 방제하기 위하여 현재까지는 농약의 사용과 병의 내성을 극복하기 위해서는 윤작이나 약제의 혼용을 권장하고 있는 실정이다. On the other hand, the most problematic pests in the cultivation of leaf perilla, rust, gray mold, mycosis, carnivorous, mite, soybean leaf moth, aphids. In order to control these pests, the use of pesticides and the combination of drugs are currently recommended to overcome pesticide use and disease resistance.
이에, 잎들깨의 재배에 있어 가장 문제가 되는 병원균인 잿빛곰팡이병원균인 B. cinerea와 균핵병원균인 S. sclerotiorum를 방제할 수 있는 길항세균을 분리, 동정했으며, 분리한 길항균을 각종 식물병원균에 대한 생물학적방제 효과를 검정하여 본 발명을 완성하였다.Therefore, we isolated and identified the antagonistic bacteria that can control B. cinerea , the gray fungus pathogen, and S. sclerotiorum , the fungal pathogen, which are the most problematic pathogens in the cultivation of leaf perilla. The present invention was completed by assaying the biological control effect.
따라서 본 발명의 목적은 잎들깨의 재배에 있어 가장 문제가 되는 병원균인 잿빛곰팡이병원균인 B. cinerea와 균핵병원균인 S. sclerotiorum를 방제할 수 있는 길항세균인 신규한 브르크호데리아 속(Burkhoderia sp.) AK17 균주를 제공하는 것이다.Therefore, an object of the present invention is a novel Burkhoderia sp, an antagonistic bacterium capable of controlling B. cinerea , a gray fungus, and S. sclerotiorum , a fungal pathogen, which is the most problematic pathogen in cultivating leaf perilla. .) To provide the AK17 strain.
본 발명의 다른 목적은 상기 균주를 배양하여, 이로부터 균주 및 배양추출물의 제조방법 및 이로부터 생물학적 제제를 제공하는 것이다. Another object of the present invention is to culture the strain, to provide a method for producing a strain and culture extract therefrom and a biological preparation therefrom.
또한, 본 발명의 또 다른 목적은 상기 생물학적 제제를 이용한 식물재배방법을 제공하는 것이다. In addition, another object of the present invention to provide a plant cultivation method using the biological agent.
상기의 목적을 달성하기 위해서, 본 발명은 신규한 균주 브르크호데리아 속 AK-17(Burkhoderia sp.AK-17) KACC91058를 제공한다.In order to achieve the above object, the present invention provides a novel strain of the genus Brukhoderia AK-17 ( Burkhoderia sp.AK-17) KACC91058.
본 발명의 다른 목적을 달성하기 위하여, 본 발명은 상기의 브르크호데리아 속 AK-17(Burkhoderia sp.) 균주를 배양하고, 상기 배양물로부터 식물 병원균에 대한 항균활성 및 식물 생육 촉진 효과를 갖는 생물학적 제제의 제조방법을 제공한다.In order to achieve the other object of the present invention, the present invention is to cultivate the strains of the genus Akr -17 ( Brkhoderia sp.) Of the above, and has an antimicrobial activity and plant growth promoting effect against plant pathogens from the culture Provided are methods for preparing a biological.
또한, 본 발명의 또 다른 목적을 달성하기 위하여, 본 발명은 상기의 방법으로 제조된 생물학적 제제를 제공한다.In addition, to achieve another object of the present invention, the present invention provides a biological agent prepared by the above method.
더 나아가 본 발명은 신규한 브르크호데리아 속 AK-17(Burkhoderia sp.AK-17) 균주 또는 상기 균주의 배양 추출물을 생물학적 제제로 사용하는 식물재배방법을 제공한다.Furthermore, the present invention provides a novel plant cultivation method using a new strain of Burkhoderia sp.AK-17 or a culture extract of the strain as a biological agent.
본 발명은 특히 잎들깨, 상추, 배추 및 시금치와 같은 엽채류와 딸기, 고추, 오이 등의 재배에 사용하면 더 효과적이다.The present invention is particularly effective when used in the cultivation of leafy vegetables such as leaf perilla, lettuce, cabbage and spinach and strawberries, peppers, cucumbers and the like.
본 발명에 따라 제조된 배양여액은 그 자체로도 식물 병원균에 항균활성 및 식물 생육 촉진 제제로서 사용할 수 있지만, 여기에 농업분야에서 사용하는 보조제, 또는 담체를 적절히 혼합하여 생물학적 제제로 사용하면 상승효과를 얻을 수 있다. The culture filtrate prepared according to the present invention can be used as an antimicrobial activity and plant growth promoting agent in plant pathogens by itself, but synergistic effect when used as a biological agent by admixing the adjuvant or carrier used in the agricultural field here Can be obtained.
이때, 혼합제로 만들어지는 제제들은 분말, 입제, 수분산입제, 습윤성 분말, 액제 등으로 유화농축제제, 용액제제, 현탁제제, 유화제제등을 사용할 수 있다. 액체 담체는 일반적인 유기용매와 같은 톨루엔, 자일렌, 아세톤, 에탄올, 메탄올 등을 사용하여 본 발명의 배양여액을 농축하고, 이를 물로 희석하여 사용할 수 있게 한다. 고체 담체로는 탈크, 점토, 실리카, 쵸크, 규조토, 라임, 벤토나이트, 콩가루, 목재가루 등을 사용할 수 있다.At this time, the formulations made of a mixed agent may be used as an emulsifier, solution, suspension, emulsifier, etc. as a powder, granules, water dispersion, wetting powder, liquid. The liquid carrier can be used by concentrating the culture filtrate of the present invention using toluene, xylene, acetone, ethanol, methanol and the like as a general organic solvent, and diluted with water. As the solid carrier, talc, clay, silica, chalk, diatomaceous earth, lime, bentonite, soy flour, wood flour and the like can be used.
또한, 본 발명은 엽면과 토양처리를 통하여, 병원균의 감염 예방과 치료효과를 얻을 수 있으며, 식물권근의 생활환경을 개선시켜 영양분 흡수를 촉진시켜 생육 촉진을 유도하는 재배방법으로 사용할 수 있다.In addition, the present invention can be obtained through the treatment of foliar and soil, to obtain the effect of preventing and treating the infection of pathogens, improve the living environment of plant roots can be used as a cultivation method to induce growth by promoting nutrient absorption.
이하, 본 발명을 보다 상세히 설명하자면 다음과 같다.Hereinafter, the present invention will be described in more detail.
1. 길항 세균의 분리 및 동정1. Isolation and Identification of Antagonist Bacteria
들깨 재배지로부터 병징을 나타내는 식물체의 뿌리, 줄기, 잎, 그리고 토양으로부터 균주들을 분리하고, 분리된 균주는 플레이트(plate) 대치배양법으로 들깨의 병원성 균주들에 대한 항균력을 비교분석하여, 그중 효과가 뛰어난 길항 세균을 선정하고, 선정된 길항세균은 버기 메뉴얼(Bergey's Manual)을 이용하여 생리, 생화학적인 특성을 분석하고, 16S rRNA 서열 검색에 의한 분자 유전학적인 방법에 의해 분석하여 동정한 결과, 본 발명은 들깨 병원균에 대한 항균력이 뛰어난 브르크호데리아 속에 속하는 신규한 균주임을 알 수 있었다. The isolates are isolated from the roots, stems, leaves, and soils of plants that exhibit the symptoms from the perilla cultivation, and the isolated strains are plate replacement cultures to compare and analyze the antibacterial activity against the pathogenic strains of perilla. As a result of selecting the antagonist bacteria, the selected antagonists were analyzed by physiological and biochemical characteristics using a buggy manual (Bergey's Manual), and analyzed by molecular genetic methods by 16S rRNA sequence search, the present invention is It was found that it is a novel strain belonging to the genus Breckhorderia excellent antibacterial activity against perilla pathogen.
이를, 본 발명자는 이 균주를 브르크호데리아 속 (Burkhoderia sp.)AK17 라 명명하였으며, 2003년 7월 11일 농업생명공학연구원에 기탁하여 KACC91058 수탁번호를 부여 받았다.This inventor named this strain Burkhoderia sp. AK17, and on July 11, 2003, was deposited with the Institute of Agricultural Biotechnology and assigned the accession number KACC91058.
2. 브르크호데리아 속 AK-17(2. AK-17 in the genus Brhoderia BurkhoderiaBurkhoderia sp. AK-17) 균주의 항균 스펙트럼 sp. Antibacterial Spectrum of AK-17 Strains
본 발명의 브르크호데리아 속 (Burkhoderia sp.)AK17 균주는 잎들깨 주요병원균인 잿빛곰팡이병원균과 균핵병원균에 대한 항균활성이 우수하며, 시들음병원균에도 양호한 항균활성을 나타낸다. Burkhoderia sp. AK17 strain of the present invention has excellent antibacterial activity against gray fungal pathogens and fungal nucleus pathogens, which are the main pathogens of leaf perilla, and show good antibacterial activity against wilted pathogens.
3. 브르크호데리아 속 AK-17(3. AK-17 in the genus Brhoderia BurkhoderiaBurkhoderia sp.AK-17) 균주의 생육 특성 sp.AK-17) Growth Characteristics of Strains
본 발명의 브르크호데리아 속 AK-17 균주는 세균 배양용 배지(pH 6.5), 30℃에서 가장 좋은 생육을 나타낸다. The strain AK-17 of the genus Breckhorderia of the present invention shows the best growth at bacterial culture medium (pH 6.5), 30 ℃.
이때, 세균 배양용 배지는 NB, LB, BHI 배지를 사용하고, BHI 배지가 더욱 바람직하다.In this case, as the medium for bacterial culture, NB, LB, BHI medium is used, and BHI medium is more preferable.
상기와 같은 조건에서, 브르크호데리아 속 AK-17 균주는 접종한 후 12∼22시간에 걸쳐 대수증식기가 형성되고, 최대생육은 접종한 지 28시간에 나타난다.Under these conditions, the AK-17 strain of the genus Breckhorderia is inoculated with a logarithmic growth over 12 to 22 hours after inoculation, the maximum growth appears 28 hours after inoculation.
4 . 들깨의 생육 촉진 효과 4 . Growth Promotion Effect of Perilla
본 발명인 브르크호데리아 속 AK-17 균주의 배양 여액을 들깨에 사용하면, 들깨의 생육을 촉진한다. When the culture filtrate of the AK-17 strain of the genus Brukderia of the present invention is used for perilla, the growth of perilla is promoted.
이때, 사용되는 브르크호데리아 속 AK-17 균주의 배양 여액은 2일 배양한 균배양액을 사용하는 것이 바람직하다.At this time, the culture filtrate of the AK-17 strain of the genus Brkhoderia is preferably used a culture medium cultured for two days.
본 발명의 브르크호데리아 속 AK-17 균주의 배양 여액을 사용한 들깨 잎의 전장(全長)은 일반 들깨잎에 비해 128%가 신장되었고, 들깨의 생체량(wet weight)과 건물량(dry weight)도 각각 164%와 163%를 증가 시켰다.The total length of the perilla leaves using the culture filtrate of the AK-17 strain of the genus Brukhoderia of the present invention was increased by 128% compared to the common perilla leaves, the wet weight and dry weight of perilla Also increased 164% and 163%, respectively.
본 발명의 브르크호데리아 속 AK-17 균주의 배양 여액은 잎들깨의 주요병원균에 대한 항균 성분과 더불어 전반적으로 들깨의 지상부 생육을 촉진하기 때문에, 들깨의 생물학적 방제제와 생육 촉진제로서 사용한다.Since the culture filtrate of the AK-17 strain of the genus Brukhoderia of the present invention, along with the antimicrobial component against the main pathogen of leaf perilla, promotes the growth of ground perilla in perilla, it is used as a biological control agent and growth promoter of perilla.
이하, 본 발명의 구체적인 방법을 실시예를 통하여 설명하고자 하나, 하기 실시예는 본 발명을 예시하기 위한 것일 뿐, 발명의 범위를 한정하는 것은 아니다.Hereinafter, one specific example of the present invention will be described by way of examples, but the following examples are only intended to illustrate the present invention and are not intended to limit the scope of the present invention.
[실시예 1] 길항 세균의 분리 및 동정Example 1 Isolation and Identification of Antagonistic Bacteria
(1) 길항 세균의 분리(1) Isolation of Antagonist Bacteria
본 발명의 균주는 경상도 밀양지역의 잎들깨 재배지로부터 병징(病徵)을 나타내는 식물체와 정상식물체, 그리고 토양을 채취하여 길항균분리의 시료로 사용하였다. 식물체의 줄기와 잎의 절편과 토양을 살균 증류수로 10-1∼10-9까지 희석하는 물질희석 아가 방법(material dilution agar methods)으로 PDA배지(potato dextrose agar media)와 NA 플레이트(nutrient agar plate)에 30℃에서 배양하면서 근접미생물의 생육을 저해시키는 수종의 곰팡이와 세균을 선별하여 1차 분리하였다. 그런다음, 선별된 곰팡이와 세균에서 가장 항균활성이 큰 균주를 단일 콜로니(colony)가 얻어질 때까지 NA배지로 계대배양을 하면서 순수분리를 하였다.The strain of the present invention was used as a sample of antagonistic bacteria by collecting plants, normal plants, and soil showing the symptom from the leaf perilla cultivation in Milyang-si, Gyeongsang-do. Potato dextrose agar media and NA plate (nutrient agar plate) using material dilution agar methods to dilute plant stem and leaf sections and soil with sterilized distilled water from 10 -1 to 10 -9 While culturing at 30 ℃ in the species of several kinds of fungi and bacteria that inhibit the growth of proximity microorganisms were selected and separated first. Then, strains with the most antimicrobial activity from the selected fungi and bacteria were purely separated by passage in NA medium until a single colony was obtained.
(2) 길항세균의 항균 스펙트럼(2) antibacterial spectrum of antagonistic bacteria
순수 분리한 단일 콜로니를 길항세균 AK-17로 임시 명명하였으며, 상기 세균은 들깨 주요 병원균인 잿빛곰팡이병원균(Botrytis cinerea), 균핵병원균(Sclerotinia sclerotiorum), 모잘록병원균(Pythium ultimum), 시들음병원균(Fusarium oxysporum), 및 줄기썩음병원균(Rhizoctonia solini)에 대해 PDA 플레이트 대치배양법으로 조사하여, 항균 스펙트럼을 확인하였다. 이때, 조건은 1/5 강도(strength)의 PDA로 각 병원균과 길항세균 AK-17을 동시에 접종하여 28℃에서 2일간 배양하고, 생육 억제 지대(inhibition zone)의 크기를 확인하였다.The purely isolated single colony was temporarily named as antagonist AK-17, and the bacteria were the major pathogens of perilla , Botrytis cinerea , Sclerotinia sclerotiorum , Pythium ultimum , and Fusarium oxyspor. ), And stem rot pathogen ( Rhizoctonia solini ) was examined by PDA plate replacement culture method, to confirm the antibacterial spectrum. At this time, the conditions were inoculated simultaneously with each pathogen and antagonistic bacteria AK-17 with 1/5 strength PDA, and cultured at 28 ° C. for 2 days, and confirmed the size of the growth inhibition zone (inhibition zone).
그 결과, 표 1과 같이, 길항세균 AK-17은 상기 들깨 병원균 모두에 항균 효과를 나타내었으며, 특히, 균핵병원균(A)과 잿빛곰팡이병원균(B)에 대한 항균효과가 월등히 우수한 것을 알 수 있었다(도 1 참조).As a result, as shown in Table 1, the antagonistic bacteria AK-17 showed an antimicrobial effect on all of the perilla pathogens, in particular, the antimicrobial effect against mycobacterial pathogens (A) and gray fungal pathogens (B) was excellent. (See Figure 1).
* 저해크기: ++++ 20㎜ 이상, +++ 15∼20㎜, ++10∼15㎜, + 5∼10㎜* Inhibition size: ++++ 20 mm or more, +++ 15-20 mm, ++ 10-15 mm, + 5-10 mm
(3) 길항 세균의 동정(3) Identification of antagonistic bacteria
길항균 AK-17은 버기 메뉴얼(Bergey's Manual)을 이용하여 생리, 생화학적인 특성을 분석하고, 16S rRNA 유전자 시퀀싱 키트(제품명, 제조사)를 사용하여 서열을 결정하여, 그 결과를 ATCC 표준 균주들의 16S rRNA 분석 라이브러리와 비교하여 동정하였다.Antagonist AK-17 analyzes the physiological and biochemical properties using the buggy manual (Bergey's Manual), sequence is determined using 16S rRNA gene sequencing kit (product name, manufacturer), and the result is 16S rRNA of ATCC standard strains. Identification was made in comparison with the analysis library.
하기 표 1과 도 2에 나타난 바와 같이, 동정결과 본 발명의 길항균 AK-17은 브르크호데리아 속 (Burkhoderia sp.)에 속하는 미생물로, 이 신규한 미생물을 브르크호데리아 속 AK-17 (Burkhoderia sp. AK-17)로 명명하였으며, 2003년 7월 11일 농업생명공학연구원에 기탁하여 KACC91058 수탁번호를 부여 받았다..As shown in Table 1 and FIG. 2, the antagonist AK-17 of the present invention was identified as a microorganism belonging to the genus Burkhoderia sp., And this novel microorganism was referred to as the genus AK-17 ( Burkhoderia sp.AK -17) and was deposited with the Agricultural Biotechnology Research Institute on July 11, 2003 and received the KACC91058 accession number.
[실시예 2] 브르크호데리아 속 AK-17 (Burkhoderia sp. AK-17)의 생육 특성Example 2 Growth Characteristics of Burkhoderia sp. AK-17
본 발명의 브르크호데리아 AK-1 균주의 생육곡선을 조사하기 위해서, 먼저, BHI 배지(pH 6.5), 30℃에서 상기 균주를 24시간동안을 전배양하였다. 그런다음 NB 배지에서 본 배양을 하면서, 600nm에서 36시간동안 OD값을 측정하였다. 초기 5시간은 매시간 측정을 하고, 그 이후에는 30분마다 OD값을 측정하였다. In order to investigate the growth curve of the Brkhoderia AK-1 strain of the present invention, first, the strain was preincubated for 24 hours at 30 ° C in BHI medium (pH 6.5). Then, while the main culture in NB medium, OD value was measured for 36 hours at 600nm. The initial 5 hours were measured every hour, after which the OD value was measured every 30 minutes.
그 결과, 생육의 최적 pH는 6.5, 최적 온도는 30℃임을 알 수 있었다. 그리고, 12∼22시간에 걸쳐 대수증식기가 형성되었으며, 최대생육은 28시간이 경과되어서 나타났다.As a result, it was found that the optimum pH of growth was 6.5 and the optimum temperature was 30 ° C. The logarithmic growth stage was formed over 12 to 22 hours, and the maximum growth occurred after 28 hours.
[실시예 3] Example 3
3-1. 브르크호데리아 속 AK-17 균주 배양여액 제조3-1. Preparation of culture filtrate of AK-17 strain
브르크호데리아 속 AK-17 균주는 NA 배지(pH 6.5), 30℃에서 2일동안 진탕배양하였다. 그런다음, 배양액이 600nm에서 OD 1.0이 될 때까지 희석하고, 필터로 여과하여 본 발명의 배양여액을 완성하였다. The AK-17 strain of the genus Brukhodria was shaken for 2 days at NA medium (pH 6.5), 30 ℃. Then, the culture solution was diluted until the OD 1.0 became 600 at 600 nm and filtered through a filter to complete the culture filtrate of the present invention.
3-2. 브르크호데리아 속 AK-17의 병발생 억제와 방제효과3-2. Inhibition and Control Effects of AK-17 from the genus Bruchderia
병발생(病發生)의 억제효과는 잎들깨의 식물체와 병(pot)에 본 발명의 배양액을 처리하여, 병발생 억제와 방제효과를 아래와 같이 수행하여 확인하였다.The inhibition effect of the disease was confirmed by treating the culture solution of the present invention to the plant and the pot of the leaf perilla, by performing the disease suppression and control effect as follows.
먼저, 6엽기의 잎들깨를 심은 병(pot, 7×15.4×7.4㎝)에 상기 실시예3-1에서 제조한 배양여액 30㎖을 처리한 다음날, PDA 배지상에서 7일간 배양한 균핵 병원균과 잿빛곰팡이 병원균을 각각 다른 병에 1일 간격으로 2일동안 접종하여 병발생 억제 효과를 확인하였으며, 상기 배양한 균핵병원균과 잿빛곰팡이병원균을 들깨 잎에 접종하여, 발병시킨 다음, 상기 배양여액을 1일간격으로 3일동안 처리하여 병발생 방제효과를 확인하였다.First, after treating 30 ml of the culture filtrate prepared in Example 3-1 in a 6-leafed perilla seedling (pot, 7 × 15.4 × 7.4 cm), the next day, the bacterial pathogens and grayish-light cultured on PDA medium were cultured for 7 days. Fungal pathogens were inoculated into different bottles for 1 day at 2 days intervals to confirm the effect of inhibiting pathogenesis. The cultured fungal pathogens and gray fungal pathogens were inoculated on perilla leaves and developed, and then the culture filtrate was stored for 1 day. Treatment was performed for 3 days at intervals to determine the effect of disease control.
그 결과, 본 발명의 배양여액은 균핵 병원균에 대해서 병발생 억제와 방제효과 모두 우수한 것을 알 수가 있었다. As a result, it was found that the culture filtrate of the present invention was excellent in both pathogenesis suppression and control effects against mycobacterial pathogens.
즉, 균핵 병원균 발생 억제 효과는 대조군의 경우, 균을 처리한지 3일이 경과하면 병징이 나타나기 시작하여 균처리 7일후에는 100% 발병되어져 9일후면 들깨가 완전히 고사하였지만, 본 발명의 배양여액으로 처리한 실험군은 2일째까지 병 발생을 거의 볼 수 없었고, 5일째부터 병징이 나타나기 시작하여 7일째에는 45%의 병발생을 나타내어, 대조군과 비교하여 55%의 병발생 억제효과를 나타내었다(도3-A 참조).In other words, the control effect of the bacterium nucleus pathogens in the case of the control group, after 3 days after the treatment of the bacteria began to appear symptom, 100 days after 7 days of the bacterium treatment, the perilla was completely killed 9 days later, but the culture filtrate of the present invention In the experimental group treated with, almost no disease occurred until day 2, symptom began to appear on day 5 and showed disease occurrence of 45% on day 7, showing 55% disease inhibition effect compared to the control group ( 3-A).
균핵 병원균 방제 효과는 대조군에서는 7일만에 모두 병징이 나타났으나, 실험군 14일까지 생육시킨 결과, 2번에 걸쳐 잎이 새로 나도 병징의 잎은 확인되지 않아, 약 90% 이상의 방제효과를 나타내었다(도 3-B참조).The control effect of mycobacterial pathogens was all symptomatic after 7 days in the control group, but as a result of growing up to 14 days in the experimental group, the leaves of the symptom were not confirmed even if the leaves were new for 2 times. (See Figure 3-B).
잿빛곰팡이 병원균 발생 억제 효과는 대조군의 경우, 2일이 경과하면서 병징이 나타나기 시작하여 9일후에는 100% 발병되어 고사하는 반면, 실험군은 2일째까지는 병징이 나타나지 않고, 3일째부터 병징이 나타나기 시작하여 9일이후에도 60%만 발병되어, 대조군과 비교하여 40%의 억제효과를 나타내었다(도 4-A 참조).The control effect of gray fungal pathogens in the control group began to develop symptom after 2 days and 100% after 9 days of death, whereas the experimental group showed no symptoms until 2 days, and began to show symptoms from 3 days. After 9 days, only 60% of the disease occurred, showing a 40% inhibitory effect compared to the control (see Fig. 4-A).
잿빛곰팡이 병원균 발생 방제 효과는 대조군에서는 9일만에 모두 발병되었으나, 실험군에서는 9일째에 새잎이 돋아나고, 더 이상의 발병은 진행되지 않고 약 70%의 방제효과를 나타내었다(도 4-B 참조).The control effect of gray mold pathogens was all developed in 9 days in the control group, but in the experimental group, young leaves sprouted on the 9th day, and no further development occurred, and the control effect was about 70% (see FIGS. 4-B).
또한, 도 5와 같이, 전반적으로 본 발명의 실험군은 대조군보다 매우 잘 생육되고 있는 것을 알 수가 있었다.In addition, as shown in Figure 5, the experimental group of the present invention as a whole it was found that the growth is much better than the control group.
[실시예 4] 생육 촉진 효과Example 4 Growth Promoting Effect
4-1. 본 발명의 생육 촉진용 배양액4-1. Growth medium of the present invention
상기 실시예 3을 통하여 진탕 배양된 배양액을 1×106(CFU)로 희석하고, 균이 살아있는 상태로 유지하면서 생육 촉진용 배양액을 제조하였다.In Example 3, the cultured shake culture was diluted to 1 × 10 6 (CFU), and cultures for promoting growth were prepared while the bacteria were kept alive.
4-2. 생육 촉진 실험4-2. Growth promotion experiment
도 4의 결과, 본 발명으로 처리한 들깨의 생육 상태가 우수한 것으로 확인됨에 따라, 들깨(Perilla ocymoides L.)를 병(Pot)에 심고, 식물 배양기에 35일동안 육종하였다. 그리고, 본 발명의 생육 촉진용 배양액이 들깨 줄기의 전장, 직경, 생체중량(fresh weight), 건물량(dry weight)과 들깨 잎의 엽수, 엽면적, 생체중량, 건물량, 엽록소의 함량에 주는 효과를 비교 분석하였다.As a result of Figure 4, the growth of the perilla treated with the present invention was confirmed to be excellent, perilla ( Perilla ocymoides L.) was planted in a bottle (Pot), and planted in a plant incubator for 35 days. In addition, the effect of the growth promoting culture of the present invention on the total length, diameter, fresh weight, dry weight and leaf number, leaf area, bio weight, dry weight, chlorophyll content of perilla stems Was analyzed comparatively.
식물 배양기의 조건은, 하루 기준으로 주간(晝間)은 16시간동안 25℃로, 야간(夜間)은 8시간동안 15℃로 유지되도록 세팅하였다.The conditions of the plant incubator were set to be maintained at 25 ° C. for 16 hours on day and 15 ° C. for 8 hours on day basis.
들깨를 병에 심고, 10일이 지난 후, 상기 실시예 4-1의 생육 촉진용 배양액을 들깨의 잎과 줄기에 촉촉하게 분무하였다. 그리고, 본 발명의 효과를 비교하기 위하여 대조군(물)과 본 발명의 균주를 제외한 배지성분만으로 처리한 배지군을 각각 준비하였다.Perilla was planted in a bottle, and after 10 days, the growth promoting culture solution of Example 4-1 was sprayed onto the leaves and stems of perilla. And, in order to compare the effects of the present invention, the control group (water) and the media group treated with only the media components except the strain of the present invention were prepared, respectively.
그 후, 25일동안 들깨를 육종한 다음, 들깨의 줄기와 잎의 변화를 측정하였다.Thereafter, perilla was bred for 25 days, and then the change of stem and leaf of the perilla was measured.
그 결과, 들깨 줄기에 있어서는 표 3과 같이, 본 발명의 전장은 대조군보다 120%, 배지군보다는 109%가 신장되었고, 생체중량과 건물량은 대조군에 비해 각각 164%, 163% 증가한 것을 알 수가 있었다.As a result, in the perilla stem, as shown in Table 3, the overall length of the present invention was increased by 120% and 109% than the medium group, and the biomass and dry weight increased by 164% and 163%, respectively, compared to the control group. there was.
또한, 들깨 잎에 있어서는 표 4와 같이, 본 발명의 엽수는 대조군보다 119%, 배지군보다 128%가 증가하였고, 엽면적은 대조군에 비해 221%, 배지군에 비해 194%가 넓어졌으며, 생체중량과 건물량은 대조군과 배지군에 비교하여 180∼210%가 증가함을 알 수가 있었다.In addition, as shown in Table 4, the number of leaves of the present invention is increased by 119% than the control group, 128% than the medium group, the leaf area is 221% compared to the control group, 194% wider than the medium group, the bio weight And dry mass was found to increase by 180 ~ 210% compared to the control and the medium group.
따라서, 본 발명의 생육 촉진용 배양액은 들깨의 생육을 촉진하여, 출하시기를 앞당길 수 있으며, 수확량도 많게 한다.Therefore, the growth promoting culture of the present invention promotes the growth of perilla, can accelerate the shipping time, and also increase the yield.
이와같이, 본 발명은 식물의 병원균에 대한 항균 활성 및 생육을 촉진하기 때문에, 농작물의 품질을 향상시키면서, 출하시기를 단축하고, 수확량도 확대되어, 농민의 소득증대에 큰 영향을 줄 것으로 기대된다.As described above, the present invention promotes the antimicrobial activity and growth of plant pathogens, thereby improving the quality of the crops, shortening the shipping time, and increasing the yield, which is expected to greatly affect the income increase of farmers.
도 1은 균핵병원균(A)과 잿빛곰팡이병원균(B)에 대한 본 발명의 길항균 AK-17의 항균효과를 고체 배지 상에서 나타낸 것이고,Figure 1 shows the antimicrobial effect of the antagonist AK-17 of the present invention against mycobacterial pathogen (A) and gray mold pathogen (B) on a solid medium,
도 2는 본 발명의 브르크호데리아 속 AK-17 균주의 16S rRNA 염기서열을 나타낸 것이고,Figure 2 shows the 16S rRNA nucleotide sequence of the genus Ak-17 strain of the genus Hokkorderia,
도 3는 균핵 병원균에 대한 본 발명의 브르크호데리아 속 AK-17 균주의 배양여액의 균핵 병원균 발생 억제 효과(A)와 균핵 병원균 방제 효과(B)를 그래프로 나타낸 것이고,Figure 3 is a graph showing the effect of inhibiting the development of mycobacterium pathogens (A) and mycobacterial pathogens control effect (B) of the culture filtrate of the genus Akr-17 strain of the present invention against mycobacterial pathogens,
도 4은 잿빛곰팡이병원균에 대한 본 발명의 브르크호데리아 속 AK-17 균주의 배양여액의 잿빛곰팡이병원균 발생 억제 효과(A)와 잿빛곰팡이병원균 방제 효과(B)를 그래프로 나타낸 것이고,Figure 4 is a graph showing the gray fungal pathogen inhibitory effect (A) and gray fungal pathogen control effect (B) of the culture filtrate of the A. K. strain AK-17 strain of the present invention against the gray fungal pathogen,
도 5는 재배 조건을 대조군(A), 잿빛곰팡이병원균으로 처리한 군(B), 본 발명의 배양여액으로 처리하고 잿빛곰팡이병원균으로 처리한 군(C), 잿빛곰팡이병원균으로 처리하고 본 발명의 배양여액으로 처리한 군(D), 및 본 발명의 배양여액으로 처리한 군(E)으로 한 후 들깨의 지상부 상태를 사진으로 나타낸 것이다. 5 is a cultivation condition of the control group (A), the group treated with gray mold pathogen (B), the group treated with the culture filtrate of the present invention and treated with gray mold pathogen (C), the gray mold pathogen of the present invention After the group (D) treated with the culture filtrate and the group (E) treated with the culture filtrate of the present invention, the state of the ground portion of the perilla is shown in the photograph.
Claims (5)
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| KR10-2003-0075077A KR100523991B1 (en) | 2003-10-27 | 2003-10-27 | Method for preparing biological material having antimicrobial against pathogen and growth stimulator for plant, and for cultivating it thereof |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| KR10-2003-0075077A KR100523991B1 (en) | 2003-10-27 | 2003-10-27 | Method for preparing biological material having antimicrobial against pathogen and growth stimulator for plant, and for cultivating it thereof |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| KR20050039979A KR20050039979A (en) | 2005-05-03 |
| KR100523991B1 true KR100523991B1 (en) | 2005-10-25 |
Family
ID=37241892
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| KR10-2003-0075077A KR100523991B1 (en) | 2003-10-27 | 2003-10-27 | Method for preparing biological material having antimicrobial against pathogen and growth stimulator for plant, and for cultivating it thereof |
Country Status (1)
| Country | Link |
|---|---|
| KR (1) | KR100523991B1 (en) |
Families Citing this family (17)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| KR100685519B1 (en) * | 2006-03-06 | 2007-03-09 | 대한민국 | Multifunctional oligotrophic bacteria Burkholderia cepacia CR2 strain isolated from ginseng field producing antibiotic enzymes and industrial enzymes containing antibiotic activation |
| MX2020003136A (en) * | 2013-06-26 | 2021-03-25 | Indigo Ag Inc | Seed-origin endophyte populations, compositions, and methods of use. |
| DK3041338T3 (en) | 2013-09-04 | 2020-03-16 | Indigo Ag Inc | Agricultural endophyte plant compositions and methods of use |
| EP3653056A1 (en) | 2013-11-06 | 2020-05-20 | The Texas A&M University System | Fungal endophytes for improved crop yields and protection from pests |
| US10271554B2 (en) | 2013-12-24 | 2019-04-30 | Ait Austrian Institute Of Technology Gmbh | Plants containing beneficial endophytes |
| US9364005B2 (en) | 2014-06-26 | 2016-06-14 | Ait Austrian Institute Of Technology Gmbh | Plant-endophyte combinations and uses therefor |
| WO2015100432A2 (en) | 2013-12-24 | 2015-07-02 | Symbiota, Inc. | Method for propagating microorganisms within plant bioreactors and stably storing microorganisms within agricultural seeds |
| AU2015278238B2 (en) | 2014-06-20 | 2018-04-26 | The Flinders University Of South Australia | Inoculants and methods for use thereof |
| EP3161124B1 (en) | 2014-06-26 | 2020-06-03 | Indigo Ag, Inc. | Endophytes, associated compositions, and methods of use thereof |
| MX2017013866A (en) | 2015-05-01 | 2018-04-13 | Indigo Agriculture Inc | Designed complex endophyte compositions and methods for improved plant traits. |
| MX2017013864A (en) | 2015-05-01 | 2018-04-24 | Indigo Agriculture Inc | Isolated complex endophyte compositions and methods for improved plant traits. |
| WO2016200987A1 (en) | 2015-06-08 | 2016-12-15 | Indigo Agriculture, Inc. | Streptomyces endophyte compositions and methods for improved agronomic traits in plants |
| EP3393225A4 (en) | 2015-12-21 | 2019-11-06 | Indigo AG, Inc. | Endophyte compositions and methods for improvement of plant traits in plants of agronomic importance |
| WO2018102733A1 (en) | 2016-12-01 | 2018-06-07 | Indigo Ag, Inc. | Modulated nutritional quality traits in seeds |
| MX2019007637A (en) | 2016-12-23 | 2019-12-16 | Texas A & M Univ Sys | Fungal endophytes for improved crop yields and protection from pests. |
| AU2017401833A1 (en) | 2017-03-01 | 2019-10-03 | Indigo Ag, Inc. | Endophyte compositions and methods for improvement of plant traits |
| EP3629742A4 (en) | 2017-04-27 | 2022-01-05 | Flinders University Of South Australia | Bacterial inoculants |
-
2003
- 2003-10-27 KR KR10-2003-0075077A patent/KR100523991B1/en not_active IP Right Cessation
Also Published As
| Publication number | Publication date |
|---|---|
| KR20050039979A (en) | 2005-05-03 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| KR100523991B1 (en) | Method for preparing biological material having antimicrobial against pathogen and growth stimulator for plant, and for cultivating it thereof | |
| RU2550268C2 (en) | Novel fluorescent pseudomonade type pseudomonas azotoformans for enhancing plant germination and growth | |
| KR101279027B1 (en) | Bacillus amyloliquefaciens LM11 strain, composition for control plant disease and control method of plant disease with same | |
| KR100802494B1 (en) | The antiparasitic and antifungal agent containing violacein and the method of it | |
| KR101346698B1 (en) | Endophytic Bacillus methylotrophicus EML-CAP7 with antimicrobial activity against plant pathogens and MRSA | |
| KR101379708B1 (en) | Bacillus amyloliquefaciens EML-CAP3 strain, composition for control plant diseases and control method of plant diseases with same | |
| KR102470864B1 (en) | Promotes onion growth and drought tolerance by bacillus altitudinis h5-9 and uses thereof | |
| US6589524B1 (en) | Strains of Bacillus for biological control of pathogenic fungi | |
| KR101563645B1 (en) | Bacillus amyloliquefaciens KR-PBS6 as a novel strain with antifungal and enzyme activity | |
| KR101972079B1 (en) | A novel Pseudomonas saponiphila GHR1-1 strain that promotes plant growth and inhibits plant diseases, and microbial agent containing the same | |
| KR101224485B1 (en) | Novel Streptomyces geldanamycininus BS3283 and method for controlling plant pathogen diseases using the same | |
| KR101100685B1 (en) | Novel bacillus subtilis and microorganism agent comprising the strains for preventing sclerotinia rot and fusarium wilt of plants | |
| Sethi et al. | Screening of biocontrol potential of indigenous Bacillus spp. isolated from rice rhizosphere against R. solani, S. oryzae, S. rolfsii and response towards growth of rice. | |
| KR100773091B1 (en) | A New Strain Bacillus atrophaeus CNU05-1 and Composition for Plant Pathogens Using the Strain | |
| EP3536153B1 (en) | The composition comprising isolated strains of saprophytic soil bacteria, biopreparation containing such composition and methods and uses thereof | |
| KR101279040B1 (en) | Exiguobacterium acetylicum WCU292 strain, composition for control plant disease and control method of plant disease with same | |
| KR100997677B1 (en) | Pseudomonas geniculata mh102 strain and method for the biological control of plant diseases using same | |
| KR101389975B1 (en) | culture medium for preventing disease and promoting growth of plant, manufacturing method of microbial agent using the same | |
| KR20110120748A (en) | Bacillus amyloliquefaciens cp1 and contro method of strawberry anthracnose using the same | |
| KR101279026B1 (en) | Bacillus amyloliquefaciens KB3 strain, composition for control plant disease and control method of plant disease with same | |
| KR20080037849A (en) | Novel streptomyces sp. bacteria and biopesticide using the same | |
| KR102143334B1 (en) | Pseudomonas frederiksbergensis strain possessing antifungal activity against major pathogenic bacteria of plant and use thereof | |
| KR20190012685A (en) | Microbial agents for promoting plant growth comprising Pseudomonas nitroreducens or culture thereof as active ingredient | |
| KR101406823B1 (en) | Bacillus sp. CS-52 and Antifungal control against Anthracnose of Red Pepper using the same | |
| KR100506721B1 (en) | Bacillus licheniformis N1 and microbial agent for preventing plant-pathogenic fungi containing the same |
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: 20110401 Year of fee payment: 6 |
|
| LAPS | Lapse due to unpaid annual fee |