JP2006333829A - RRj228 OF GENUS PSEUDOMONAS, HELPING AGRICULTURAL CROP GROWTH AND MICROORGANISM PREPARATION CONTAINING THE SAME - Google Patents
RRj228 OF GENUS PSEUDOMONAS, HELPING AGRICULTURAL CROP GROWTH AND MICROORGANISM PREPARATION CONTAINING THE SAME Download PDFInfo
- Publication number
- JP2006333829A JP2006333829A JP2005164995A JP2005164995A JP2006333829A JP 2006333829 A JP2006333829 A JP 2006333829A JP 2005164995 A JP2005164995 A JP 2005164995A JP 2005164995 A JP2005164995 A JP 2005164995A JP 2006333829 A JP2006333829 A JP 2006333829A
- Authority
- JP
- Japan
- Prior art keywords
- strain
- rrj228
- pseudomonas
- kctc
- growth
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Granted
Links
Images
Landscapes
- Agricultural Chemicals And Associated Chemicals (AREA)
- Fertilizers (AREA)
- Micro-Organisms Or Cultivation Processes Thereof (AREA)
Abstract
Description
本発明は、作物の栽培時、化学肥料の代わりに、植物根の表面と内部に定着して植物の生長を促進させる配列番号１のシュードモナス属RRj228（KCTC 10812BP）菌株に関する。なお、このシュードモナス属RRj228（Pseudomonas sp. RRj228）については、2003年6月11日付けで「寄託番号KACC 91052」の下に韓国農村振興庁農業生命工学研究院に付設の韓国農用微生物保存センター（KACC）に寄託されたが、2005年5月27日付けで『寄託番号KCTC 10812BP』の下に韓国の国際寄託機関であるＫＲＩＢＢ（Korean Research Institute of Biosceience and Biotechnology）のＫＣＴＣ（Korean Culture for Type Cultures）に再寄託されたので、以下の記載においては、「寄託番号KACC 91052」に代えて『寄託番号KCTC 10812BP』を用いる。 The present invention relates to a Pseudomonas RRj228 (KCTC 10812BP) strain of SEQ ID NO: 1, which, instead of chemical fertilizer, is grown on the surface and inside of plant roots to promote plant growth during crop cultivation. The Pseudomonas sp. RRj228 (Pseudomonas sp. RRj228) was established on June 11, 2003 under the “Deposit No. KACC 91052” at the Korea Agricultural Microorganisms Conservation Center (Agricultural Biotechnology Research Institute) KACCC (Korean Culture for Type Cultures) of KRIBB (Korean Research Institute of Biosceience and Biotechnology), Korea's international depositary institution under “Deposition No. KCTC 10812BP” on May 27, 2005 In the following description, “deposit number KCTC 10812BP” is used instead of “deposit number KACC 91052”.
また、本発明は、前記シュードモナス属RRj228（KCTC 10812BP）菌株及びこの菌株から生産される２次代謝産物を含有する環境親和的微生物製剤に関する。 The present invention also relates to an environmentally friendly microbial preparation containing the Pseudomonas RRj228 (KCTC 10812BP) strain and a secondary metabolite produced from the strain.
農業において、肥料は、作物の収量を決定づける重要な農業資材であるが、過度に多く使用する場合、河川水の富栄養化、地下水の水質汚染、土壌養分の不均衡、作物生育の不良要因として作用して、収量減少と共に農産物の品質低下を招く。それで、既存の化学肥料を部分的に代替できる植物生長促進機能を発揮しつつ、農作物を丈夫に育てる土壌微生物の適切な活用は、持続農業の発展と農業環境の保全のために必ず必要である。 In agriculture, fertilizer is an important agricultural material that determines the yield of crops, but when used excessively, fertilizers are used as eutrophication of river water, contamination of groundwater, soil nutrient imbalance, and poor growth of crops. Acts, leading to a decrease in yield and a decrease in the quality of agricultural products. Therefore, the proper use of soil microorganisms that cultivate crops robustly while demonstrating plant growth promotion functions that can partially replace existing chemical fertilizers is absolutely necessary for the development of sustainable agriculture and the preservation of the agricultural environment. .
土壌微生物のうち、宿主植物と完全な共生関係を有することなく植物の生長を促進させる微生物は、1980年KloepperなどによりＰＧＰＲ（Plant growth-promoting rhizobacteria）と命名され、これには、空中窒素を固定するAzospirrilum属菌（Okon, 1985）、Pseudomonas属菌（Kloepper et al., 1980）、Bacillus属菌（Backman et al., 1994）などが代表的な微生物に含まれる。いわゆるＰＧＰＲ菌株と呼ばれる植物生長促進微生物は、空中窒素の固定（Boddy and Dobereiner, 1995）、植物生長促進ホルモンの生産、植物の養分吸収促進（Hallmann et al., 1997）、植物の病発生抑制（Pleban et al., 1995）などの色々な機能によって植物の生長を助けると知られている。 Among the soil microorganisms, a microorganism that promotes plant growth without having a complete symbiotic relationship with the host plant was named PGPR (Plant growth-promoting rhizobacteria) by Kloepper in 1980. Azospirrilum (Okon, 1985), Pseudomonas (Kloepper et al., 1980), Bacillus (Backman et al., 1994) and the like are included in typical microorganisms. Plant growth-promoting microorganisms called so-called PGPR strains can be used to fix aerial nitrogen (Boddy and Dobereiner, 1995), produce plant growth-promoting hormones, promote plant nutrient uptake (Hallmann et al., 1997), Pleban et al., 1995) and other functions are known to help plant growth.
Wardなど（1992）は、土壌に棲息する微生物のうち、分類及び同定されたものは、全体種の５%以下であると述べており、Torisvikなど（1990）は、土壌には人為的に培養されない細菌が多いと述べた。したがって、既に発見された有用微生物より機能性に優れた他の菌株がいくらでも存在するはずであり、これについての研究が続いて進行されてきている。 Ward et al. (1992) state that less than 5% of all species of microorganisms inhabiting soil are classified and identified, and Torisvik et al. (1990) artificially cultivate in soil. Said there are many bacteria that are not. Therefore, there must be any number of other strains that are more functional than the already discovered useful microorganisms, and research on this has continued.
また、植物の生長を助ける色々な植物生長促進微生物の作物接種効果は、稲で１９〜６０%（Baldani, 2000）増収されたことをはじめとして、トマト（Kloepper and Schroth, 1981）、豆（Polonenko et al., 1987; Dashti et al., 1998）、小麦（Freitas and Germida, 1990）、砂糖大根（Suslow and Schrith, 1982）などでも確認され、これらの植物生長促進微生物の実用化についての研究が進行されている。 In addition, the plant inoculation effect of various plant growth promoting microorganisms that help plant growth has been increased by 19-60% (Baldani, 2000) in rice, tomato (Kloepper and Schroth, 1981), beans (Polonenko) et al., 1987; Dashti et al., 1998), wheat (Freitas and Germida, 1990), sugar radish (Suslow and Schrith, 1982), etc., and research on the practical application of these plant growth-promoting microorganisms has been conducted. It is progressing.
これより、本発明者らは、土壌に棲息する土壌微生物について研究していたところ、稲根の表面と内部に定着する特性がある植物根圏細菌シュードモナス属RRj228（KCTC 10812BP）菌株を最初に分離・同定するようになり、この菌株が農作物の生長を促進させて、従来の化学肥料を代替できることを知見し、本発明を完成するに至った。 Based on this, the present inventors have studied soil microorganisms that inhabit the soil, and first isolated the plant rhizosphere bacterium Pseudomonas sp. RRj228 (KCTC 10812BP), which has the characteristics of being settled on the surface and inside of rice roots. -It came to identify, and it discovered that this strain promoted the growth of agricultural crops and could substitute the conventional chemical fertilizer, and came to complete this invention.
したがって、本発明の目的は、作物の生長に役立つ植物生長促進力を持ち、化学肥料の部分的な代替機能を有するシュードモナス属RRj228（KCTC 10812BP）菌株を提供することにある。 Accordingly, an object of the present invention is to provide a Pseudomonas RRj228 (KCTC 10812BP) strain having a plant growth promoting ability useful for crop growth and having a partial substitute function of chemical fertilizer.
また、本発明の目的は、前記シュードモナス属RRj228（KCTC 10812BP）菌株及びこの菌株から生産される２次代謝産物を有効成分として含有する微生物製剤を提供することにある。 Another object of the present invention is to provide a microorganism preparation containing the Pseudomonas RRj228 (KCTC 10812BP) strain and a secondary metabolite produced from this strain as active ingredients.
本発明は、植物根の表面と内部に定着して植物の生長を促進させる配列番号１のシュードモナス属RRj228（KCTC 10812BP）菌株に関する。 The present invention relates to a Pseudomonas RRj228 (KCTC 10812BP) strain of SEQ ID NO: 1, which is established on the surface and inside of a plant root to promote plant growth.
本発明によるシュードモナス属RRj228（KCTC 10812BP）菌株は、本発明者により最初に発見された土壌細菌であって、自然条件で作物根の表面と内部に定着して植物生長促進物質を分泌し、植物病原性カビや細菌に対する拮抗力を有していて、各種植物病原菌の生長抑制力を持つ。 The Pseudomonas RRj228 (KCTC 10812BP) strain according to the present invention is a soil bacterium first discovered by the present inventor, which is established on the surface and inside of a crop root under natural conditions and secretes a plant growth promoting substance. Has antagonism against pathogenic fungi and bacteria and has the ability to suppress the growth of various plant pathogens.
本発明によるシュードモナス属RRj228（KCTC 10812BP）菌株は、ロッド形状を有するグラム（gram）陰性の細菌であり、２８±２℃条件のＰＤＡ（Potato Dextrose Agar）培地でよく成長する中温菌である。 The Pseudomonas RRj228 (KCTC 10812BP) strain according to the present invention is a gram-negative bacterium having a rod shape, and is a mesophilic bacterium that grows well in a PDA (Potato Dextrose Agar) medium at 28 ± 2 ° C.
本発明のシュードモナス属RRj228（KCTC 10812BP）菌株を使用することによって得られる効果は、次の通りである。 The effects obtained by using the Pseudomonas genus RRj228 (KCTC 10812BP) strain of the present invention are as follows.
（１）本発明のシュードモナス属RRj228（KCTC 10812BP）菌株は、植物の養分供給力及び植物生長促進力に優れているので、通常の方法で微生物製剤を製作し、農作物接種剤として使用する場合、化学肥料を少なく与えても収穫量を増やすことができる。 (1) Since the Pseudomonas RRj228 (KCTC 10812BP) strain of the present invention is excellent in plant nutrient supply ability and plant growth promoting ability, when producing a microbial preparation by a usual method and using it as a crop inoculum, Yield can be increased even with less chemical fertilizer.
（２）本発明のシュードモナス属RRj228（KCTC 10812BP）菌株は、化学肥料代替資源であって、稲、唐辛子、キュウリなどの農作物に適用できるので、適用範囲が広い。 (2) The Pseudomonas sp. RRj228 (KCTC 10812BP) strain of the present invention is a chemical fertilizer alternative resource and can be applied to agricultural crops such as rice, chili, and cucumber, and thus has a wide range of applications.
（３）本発明のシュードモナス属RRj228（KCTC 10812BP）菌株は、土壌内における植物病原性微生物の生長を抑制する特性があり、作物に接種する場合、病原菌から作物の安全な生育を助ける。 (3) The Pseudomonas sp. RRj228 (KCTC 10812BP) strain of the present invention has the property of suppressing the growth of phytopathogenic microorganisms in the soil, and assists the safe growth of crops from pathogenic bacteria when inoculated into crops.
（４）本発明のシュードモナス属RRj228（KCTC 10812BP）菌株は、作物の根組織の内部に定着する特性があるので、作物接種効果発現の安定性が高い。 (4) Since the Pseudomonas sp. RRj228 (KCTC 10812BP) strain of the present invention has the property of being established in the root tissue of the crop, the stability of the crop inoculation effect expression is high.
（５）本発明のシュードモナス属RRj228（KCTC 10812BP）菌株は、培養物から２次代謝産物を通常の方法で分離、精製することによって、植物生長促進または植物病原菌生長抑制物質を純粋分離して、化学肥料代替効果以上の付加価値を創出することができる。 (5) The Pseudomonas genus RRj228 (KCTC 10812BP) strain of the present invention is obtained by purifying a plant growth promoting or plant pathogen growth inhibitory substance by separating and purifying a secondary metabolite from a culture by a conventional method, It can create added value that exceeds the chemical fertilizer substitution effect.
（６）本発明のシュードモナス属RRj228（KCTC 10812BP）菌株は、農業で環境汚染の可能性が全くない微生物資源であるから、農作物の栽培に使用する場合、水質及び土壌環境の保全と土壌生態系の健全性増進効果を期待することができる。 (6) The Pseudomonas sp. RRj228 (KCTC 10812BP) strain of the present invention is a microbial resource that has no possibility of environmental pollution in agriculture. Therefore, when it is used for cultivation of agricultural crops, water quality and soil environment conservation and soil ecosystem Can be expected to improve the soundness.
また、本発明は、前記シュードモナス属RRj228（KCTC 10812BP）菌株及びこの菌株から生産される２次代謝産物を含有する環境親和的微生物製剤に関する。 The present invention also relates to an environmentally friendly microbial preparation containing the Pseudomonas RRj228 (KCTC 10812BP) strain and a secondary metabolite produced from the strain.
本発明による微生物製剤は、通常の方法で植物生長促進用または病原菌生長抑制用製剤で剤型化することができ、その剤型は特に限定されるものではなく、好ましくは、化学肥料を代替するための植物生長促進有機肥料で剤型化することができる。 The microbial preparation according to the present invention can be formulated into a preparation for promoting plant growth or suppressing the growth of pathogenic bacteria by a usual method. The dosage form is not particularly limited, and preferably substitutes for chemical fertilizer. Can be formulated with plant growth promoting organic fertilizer.
また、本発明による微生物製剤を植物生長促進用製剤として乾燥粉末形態または液状肥料形態で製造する場合には、前記シュードモナス属RRj228（KCTC 10812BP）菌株を乾燥粉末ｇ当たり、又は液状肥料１mL当たり１０5〜１０7匹含有することができる。 When the microbial preparation according to the present invention is produced as a plant growth promoting preparation in dry powder form or liquid fertilizer form, the Pseudomonas RRj228 (KCTC 10812BP) strain is used per 10 g of dry powder or 10 5 ml of liquid fertilizer. -10 7 animals.
また、本発明による微生物製剤を病原菌生長抑制用製剤として乾燥粉末形態または液状形態で製造する場合には、前記シュードモナス属RRj228（KCTC 10812BP）菌株を乾燥粉末ｇ当たり、または液状１mL当たり１０7匹以上含有することができ、前記植物生長促進用製剤で剤型化する時よりも菌株の含量を多く使用することが特徴である。 In addition, when the microorganism preparation according to the present invention is produced as a preparation for controlling the growth of pathogenic bacteria in a dry powder form or a liquid form, the above Pseudomonas RRj228 (KCTC 10812BP) strain is 10 7 or more per 1 g of dry powder or 1 mL of liquid. It can be contained, and is characterized in that it uses a larger amount of the strain than when it is formulated with the above-mentioned preparation for promoting plant growth.
本発明によるシュードモナス属RRj228（KCTC 10812BP）菌株を化学肥料代用で適宜活用する場合、化学肥料の節減、作物生産性の安定化、水質及び土壌環境の保全、土壌生態系の健全性増進を期待することができる。 When the Pseudomonas RRj228 (KCTC 10812BP) strain according to the present invention is appropriately used as a substitute for chemical fertilizer, it is expected to reduce chemical fertilizer, stabilize crop productivity, preserve water quality and soil environment, and improve soil ecosystem health. be able to.
シュードモナス属RRj228（KCTC 10812BP）菌株は、一般的な農耕地土壌に適した微生物製剤形態で製造して、化学肥料の代わりに、稲、唐辛子、キュウリなどの農作物に接種することができる。 Pseudomonas RRj228 (KCTC 10812BP) strain can be produced in the form of a microbial preparation suitable for general farmland soil, and inoculated on crops such as rice, chili and cucumber instead of chemical fertilizer.
本発明によるシュードモナス属RRj228（KCTC 10812BP）菌株は、土壌から直接分離された従来の菌株とは異なって、植物体（稲）の根から直接分離したので、稲、キュウリなどの作物根の内部に定着する特性がよく再現される。これにより、作物に接種されると、根の表面または内部で自体増殖が可能であり、作物が生きている限り、雨水または乾燥による密度減少が少ないため、接種効果が長く持続されることができる。 The Pseudomonas RRj228 (KCTC 10812BP) strain according to the present invention is isolated directly from the root of the plant body (rice), unlike the conventional strain isolated directly from the soil. The fixing characteristics are well reproduced. Thus, when inoculated into a crop, it can grow itself on the surface or inside of the root, and as long as the crop is alive, the inoculation effect can be sustained for a long time because there is little decrease in density due to rainwater or drying. .
また、本発明による環境親和的微生物肥料組成物は、シュードモナス属RRj228（KCTC 10812BP）菌株を使用することによって、作物を栽培する時に、化学肥料を部分的に代替できるので、水質、土壌などのような農業環境の健全性を増大させることでき、作物の生産性の向上に寄与することができる。 In addition, the environment-friendly microbial fertilizer composition according to the present invention can partially substitute for chemical fertilizer when cultivating crops by using Pseudomonas RRj228 (KCTC 10812BP) strain. Can increase the soundness of a healthy agricultural environment and contribute to the improvement of crop productivity.
以下、実施例により韓国とロシア土壌から分離した１７２菌株のうち、ロシアのサンクト・ぺテルブルク（St. Petersburg）から分離した植物生長促進力に優れた本発明のシュードモナス属RRj228（KCTC 10812BP）菌株の分離、同定及び植物生長促進力について説明する。 Hereinafter, among the 172 strains isolated from the Korean and Russian soils according to the examples, the Pseudomonas RRj228 (KCTC 10812BP) strain of the present invention having excellent plant growth promoting ability isolated from St. Petersburg, Russia. The separation, identification, and plant growth promoting ability will be described.
但し、下記の実施例は、本発明を実施するために提示された一例に過ぎないもので、これらの実施例により本発明の範囲が限定されるものではない。 However, the following examples are merely examples provided for carrying out the present invention, and the scope of the present invention is not limited by these examples.
［実施例１］土壌微生物RRj228菌株の分離
植物根圏土壌微生物RRj228菌株の分離は、図１に示すように、無菌的に発芽させた種籾１粒を0.6%アガロース（Agarose）培地上に移し、稲根から１〜２cm距離に、菌株を分離すべき土壌試料０．５gを接種し、２８℃で４８時間恒温処理した後、根を磨いて懸濁液に作った後、ＬＣ培地｛バクトトリプトン(Bactotrypton)１０g、酵母抽出物(Yeast extract)５g、ＭｇＳＯ4・７Ｈ2Ｏ ８g、1M Tris １mL、蒸留水１L、ｐＨ６．８｝に塗抹して、菌株を分離した。
[Example 1] Isolation of soil microorganism RRj228 strain Isolation of the plant rhizosphere soil microorganism RRj228 strain was carried out by transferring one aseptically germinated seed pod onto a 0.6% Agarose medium, as shown in FIG. After inoculating 0.5 g of a soil sample to isolate the strain at a distance of 1 to 2 cm from the rice root, isothermally treating at 28 ° C. for 48 hours, polishing the root to make a suspension, and then LC medium {Bactoto The strain was isolated by smearing with 10 g of Bactotrypton, 5 g of yeast extract, 8 g of MgSO 4 .7H 2 O, 1 mL of 1M Tris, 1 L of distilled water, pH 6.8}.
［実施例２］土壌微生物RRj228 の同定
（イ）生理、生化学的特性
RRj228菌株の生理、生化学的特性は、下記表１（土壌微生物RRj228菌株の生理、生化学的特性）に示した。
[Example 2] Identification of soil microorganism RRj228
(B) Physiological and biochemical characteristics
The physiological and biochemical characteristics of the RRj228 strain are shown in Table 1 below (physiological and biochemical characteristics of the soil microorganism RRj228 strain).
前記RRj228菌株は、ロッド形状よりなり、サイズは、２．０〜３．０×０．５〜０．７μmであり、図２ａ及び図２ｂから分かるように、鞭毛を有している。ＰＤＢ培地では、２４時間培養すれば、集落色が赤黄色を呈した。グラム（Gram）陰性細菌であるRRj228菌株は、呼吸代謝をし、カタラーゼ（Catalase）とオキシダーゼ（Oxydase）活性を有する。そして、シュードモナス属RRj228菌株は、培地に添加された炭素源がアラビノース（Arabinose）、フラクトース（Fructose）、ガラクトース（Galactose）、グルコース（Glucose）、マンニトール（Mannitol）、スクロース（Sucrose）、キシロース（Xylose）である時は、生長過程で酸を生成し、炭素源がラクトース（Lactose）、デキストリン（Dextrine）、ラムノース（Ramnose）、ソルビトール（Sorbitol）、マルトース（Maltose）、ツシトール（Dulcitol）である時は、アルカリを生成した。また、RRj228菌株は、図３及び図４に示すように、植物根の表面に棲息しながら根組織の内部に入って定着する特性を有する。 The RRj228 strain has a rod shape, a size of 2.0 to 3.0 × 0.5 to 0.7 μm, and has flagella as can be seen from FIGS. 2a and 2b. In the PDB medium, the colony color was reddish yellow after 24 hours of culture. The RRj228 strain, which is a Gram-negative bacterium, undergoes respiratory metabolism and has catalase and oxidase activity. The Pseudomonas RRj228 strain has arabinose, fructose, galactose, glucose (Glucose), mannitol, sucrose, and xylose as the carbon source added to the medium. When it is, it generates acid during the growth process, and when the carbon source is lactose, dextrine, rhamnose, sorbitol, maltose, and ducitol, Alkali was produced. In addition, as shown in FIGS. 3 and 4, the RRj228 strain has the property of entering and rooting in the root tissue while inhabiting the surface of the plant root.
（ロ）塩基配列分析
RRj228菌株の塩基配列分析は、１６Ｓ ｒＤＮＡ配列分析法を用いて行ったが、まず、１６Ｓ ｒＤＮＡ領域を核酸増幅法（ＰＣＲ：Polymerase Chain Reaction）で増幅させ、ｐＧＥＭＴ−Ｔベクトル（vector）に挿入した後、自動塩基配列分析機ABI3100（Applied Biosystem, USA）を用いて塩基配列を分析した。これに使われたプライマーは、配列番号２のｆＤ１（５′−ａｇａｇｔｔｔｇａｔｇｇｃｔｃａｇ−３′）及び配列番号３のｒＰ２（５′−ａｃｇｇｃｔａｃｃｔｔｇｔｔａｃｇａｃｔｔ−３′）であり、プライマーの両末端部位の塩基配列を分析するために、ｐＵＣ／Ｍ１３の正方向プライマ （forward primer：５′−ｇｔｔｔｔｃｃｃａｇｔｃａｃｇａｃ−３′）（配列番号４）及び逆方向プライマー（reverse primer：５′−ｇｃｇｇａｔａａｃａａｔｔｔｃａｃａｃａｇｇ−３′）（配列番号５）を利用し、内部の塩基配列分析のために、４つのプライマーを利用した。
(B) Base sequence analysis
The base sequence analysis of RRj228 strain was performed using the 16S rDNA sequence analysis method. First, the 16S rDNA region was amplified by a nucleic acid amplification method (PCR: Polymerase Chain Reaction) and inserted into a pGEMT-T vector. Thereafter, the base sequence was analyzed using an automatic base sequence analyzer ABI3100 (Applied Biosystem, USA). The primers used for this were fD1 (5'-agagtttgatggctcag-3 ') of SEQ ID NO: 2 and rP2 (5'-acggctactttgttacgactt-3') of SEQ ID NO: 3, and the nucleotide sequences of both ends of the primer were analyzed. PUC / M13 forward primer (5'-gtttccccattcacgac-3 '(SEQ ID NO: 4) and reverse primer (5'-gcggataacaattttcacacagg-3') (SEQ ID NO: 5) 4 primers were used for internal base sequence analysis.
それぞれの塩基配列は、Escherichia coli １６Ｓ ｒＤＮＡ塩基配列（GenBank No. J01695）の順序によって３１１〜３３０部位の塩基配列である１６ＳＰ−１（５′−ｇｃｃａｃａｃｔｇｇａａｃｔｇａｇａｃａｃ−３′）、６８４〜７０３部位の塩基配列である１６ＳＰ−２（５′−ｔｇｔａｇｃｇｇｔｇａａａｔｇｃｇｔｇ−３′）、９４４〜９６３部位の塩基配列である１６ＳＰ−３（５′−ｇｇａｇｃａｔｇｔｇｔｔａｔｔｃｇ−３′）及び１２２８〜１２４７部位の塩基配列である１６ＳＰ−４（５′−ｃｔａｃａｃａｃｇｔｇｃｔａｃａａｔｇｇ−３′）を利用した。 The respective base sequences are 16SP-1 (5'-gccactactgagactgagacac-3 ') and 684 to 703 base sequences which are base sequences of 311 to 330 sites according to the order of Escherichia coli 16S rDNA base sequence (GenBank No. J01695). 16SP-2 (5'-tgttagcgggtgaatgcgtg-3 '), 16SP-3 (5'-gggcatgtgtttattcg-3') having a base sequence of 944 to 963, and 16SP-4 having a base sequence of 1228 to 1247 ( 5'-ctacacacgtgctacaatgg-3 ') was utilized.
RRj228菌株は、１６Ｓ ｒＤＮＡ塩基配列は配列番号１に記載し、GenBankライブラリーの塩基配列と相同性を調べた結果、Pseudomonas chlorororaphis、P. Corrugata、P. tolaasiiなど多数のPseudomonas種と９９%以上の塩基配列相同性を示したので、暫定的にシュードモナス属（Pseudomonas Sp.）に分類した。 As for the RRj228 strain, the 16S rDNA base sequence is described in SEQ ID NO: 1, and as a result of investigating the homology with the base sequence of the GenBank library, a large number of Pseudomonas species such as Pseudomonas chlorororaphis, P. Corrugata, P. tolaasii and more than 99% Since it showed base sequence homology, it was tentatively classified as Pseudomonas sp.
したがって、本発明者らは、RRj228菌株を“シュードモナス属RRj228菌株”と命名し、2003年6月11日付で韓国農村振興庁農業生命工学研究院に付設の韓国農用微生物保存センター（KACC）に寄託し、受託番号KACC 91052を付与された。また、このシュードモナス属RRj228（Pseudomonas sp. RRj228）については、2005年5月27日付けで国際寄託機関であるＫＲＩＢＢ（Korean Research Institute of Biosceience and Biotechnology）のＫＣＴＣ（Korean Culture for Type Cultures）に再寄託され、寄託番号KCTC 10812BPを付与された。 Therefore, the present inventors named the RRj228 strain as “Pseudomonas sp. RRj228 strain” and deposited it with the Korean Agricultural Microbiology Conservation Center (KACC) attached to the Agricultural Biotechnology Research Institute of the Korean Rural Development Agency on June 11, 2003. And was given the accession number KACC 91052. The Pseudomonas sp. RRj228 is redeposited on KCTC (Korean Culture for Type Cultures) of KRIBB (Korean Research Institute of Biosceience and Biotechnology), an international depositary institution on May 27, 2005. And was given the deposit number KCTC 10812BP.
［試験例１］シュードモナス属RRj228菌株の植物生長促進効果検定
シュードモナス属RRj228菌株の植物生長促進効果検定試験は、とうもろこし幼苗を対象にして実施した。とうもろこし幼苗接種試験は、とうもろこし根の終端３ｍｍ部位を試験菌株の培養物懸濁液に１．５時間浸漬して接種処理し、対照区の幼苗は、滅菌した微生物培養液に浸漬した。次に、とうもろこし幼苗をプラスチックボックス内の湿っぽいろ過紙に載置し、根の伸張開始点（starting line）にある根終端をろ過紙で覆った後、２８℃で２４時間恒温処理して、新しく伸張した根の長さを１０回繰り返して測定し、このようなRRj228菌株のとうもろこし生長促進力を検定した結果を下記表２（シュードモナス属RRj228菌株のとうもろこし生長促進力程度）に示した。
[Test Example 1] Plant growth promoting effect test of Pseudomonas RRj228 strain The plant growth promoting effect test of Pseudomonas RRj228 strain was conducted on corn seedlings. In the corn seedling inoculation test, the 3 mm end portion of corn root was inoculated by immersing it in the culture suspension of the test strain for 1.5 hours, and the seedlings in the control group were immersed in a sterilized microorganism culture solution. Next, the corn seedlings are placed on a damp filter paper in a plastic box, and the root ends at the starting line of the root are covered with filter paper, followed by a constant temperature treatment at 28 ° C. for 24 hours. The length of the extended root was repeatedly measured 10 times, and the results of testing the corn growth promoting ability of such RRj228 strain are shown in Table 2 below (the degree of corn growth promoting ability of Pseudomonas RRj228 strain).
前記表２から明らかなように、本発明によるシュードモナス属RRj228（KCTC 10812BP）菌株の植物生長促進力は、とうもろこし根を菌株培養液に１．５時間沈積する方法で接種した時、根伸張が培養原液（4.5×109cfu/mL）の５０倍希釈接種時、２６%減少したが、１００倍希釈接種の場合、１３．０%増加したことから、接種濃度が低い条件で効果が高いことが分かる。 As apparent from Table 2, the plant growth promoting ability of Pseudomonas sp. RRj228 (KCTC 10812BP) according to the present invention is observed when root elongation is inoculated when corn root is inoculated in a strain culture solution for 1.5 hours. When inoculating the stock solution (4.5 × 10 9 cfu / mL) by 50-fold dilution, it decreased by 26%, but in the case of 100-fold dilution inoculation, it increased by 13.0%. I understand.
［試験例２］シュードモナス属RRj228菌株の土壌病拮抗力検定
シュードモナス属RRj228菌株の土壌病拮抗力検定は、ＰＤＡに病原性カビ及び細菌を各々接種した後、シャーレに移して、培地を固めた後、半径４mm大きさのウェル（well）を作り、ここに液体の馬鈴薯培地で２８℃条件で５日間培養したシュードモナス属RRj228菌株を１００μLずつ接種して、２８℃条件で５日間保管した後、根圏細菌の病原菌生長阻止範囲を測定し、その結果を表３｛土壌微生物RRj228菌株の植物病原性カビ及び細菌に対する生長抑制力程度（菌株接種５日後の病原菌の生長阻止範囲）｝に示した。植物病原菌の接種は、カビの場合、１×１０6conidia/mL ５mLと細菌性病原菌１×１０8cfu/mL ５mLを各々４５℃のＰＤＡ培地２５０mLに混合し、攪拌した後、シャーレ当たり２５mLずつ３回繰り返して分株して接種した。
[Test Example 2] Soil disease antagonism test of Pseudomonas RRj228 strain The soil disease antagonism test of Pseudomonas RRj228 strain was inoculated with pathogenic fungi and bacteria on PDA, then transferred to a petri dish and solidified medium. Then, a well having a radius of 4 mm was made, inoculated with 100 μL each of Pseudomonas RRj228 strain cultured in liquid potato medium at 28 ° C. for 5 days, and stored at 28 ° C. for 5 days. The range of inhibition of the growth of pathogenic bacteria by the range bacteria was measured, and the results are shown in Table 3 {Degree of inhibition of growth of phytopathogenic fungi and bacteria of the soil microorganism RRj228 strain (range of inhibition of pathogenic bacteria 5 days after inoculation of the strain)}. Inoculation with phytopathogenic fungi, in the case of mold, mix 5 mL of 1 × 10 6 conidia / mL and 1 mL of bacterial pathogen 1 × 10 8 cfu / mL in 250 mL of 45 ° C. PDA medium, stir, then 25 mL each The inoculation was repeated 3 times.
上記結果から明らかなように、本発明によるRRj228菌株は、植物病原性カビであるフィトフトラ・カプシシ（Phytophtora capsici）とピシウム（Pythium）、植物病原性細菌であるエルウィニア・カロトボーラ（Erwinia carotovora）とシュードモナス・シリンゲ（Pseudomonas syringiae）などに対して強い生長抑制効果を示した。 As can be seen from the above results, the RRj228 strain according to the present invention comprises phytopathogenic fungi Phytophtora capsici and Pythium, phytopathogenic bacteria Erwinia carotovora and Pseudomonas. It showed a strong growth-inhibiting effect against Pseudomonas syringiae.
［試験例３］シュードモナス属RRj228菌株の根着生競合力
試験菌株の植物根着生競合力は、対照菌株であるシュードモナス・フルオレセンス（Pseudomonas fluorescens）ＷＣＳ３６５と試験菌株を稲（金烏稲）と大根（Saxa nova）の根に同時に接種した時、根に着生された全体菌株の中でRRj228菌株が占める割合で示し、その結果を表４（土壌微生物シュードモナス属RRj228 のシュードモナス・フルオレセンスWCS365菌株対比根着生競合力）に示した。
[Test Example 3] Pseudomonas RRj228 strain rooting ability Competitiveness The plant rooting ability of the test strain is Pseudomonas fluorescens WCS365 and the test strain is rice (Golden rice). When inoculated to the roots of radish (Saxa nova) at the same time, the ratio of the RRj228 strain among the total strains grown on the roots is shown. Table 4 (Results) (Competitiveness against rooting of strains).
［試験例４］シュードモナス属RRj228菌株の農作物接種効果検定
本発明の植物生長促進土壌微生物シュードモナス属RRj228菌株（KCTC 10812BP）を通常の方法で培養した培養液を、稲、唐辛子、キュウリ、トマトに接種し、その結果を下記表５（土壌微生物RRj228菌株の農作物接種効果）に示した。
[Test Example 4] Examination of crop inoculation effect of Pseudomonas sp. RRj228 strain Inoculated rice, chili pepper, cucumber, and tomato with a culture solution of the plant growth-promoting soil microorganism Pseudomonas sp. The results are shown in Table 5 below (inoculation effect of soil microorganism RRj228 strain).
上記表５から、シュードモナス属RRj228菌株（KCTC 10812BP）は、稲の収量を３３．２%、唐辛子の収量を８．５%、キュウリの生長を２１．４%、及びトマトの生長を１１７．５%向上させることが分かり、これにより、本発明によるシュードモナス属RRj228菌株が化学肥料を相当量代替できる微生物肥料資源として使われることができることを確認した。 From Table 5 above, Pseudomonas RRj228 strain (KCTC 10812BP) has a rice yield of 33.2%, a pepper yield of 8.5%, a cucumber growth of 21.4%, and a tomato growth of 117.5. As a result, it was confirmed that the Pseudomonas RRj228 strain according to the present invention can be used as a microbial fertilizer resource capable of substituting a considerable amount of chemical fertilizer.
Claims (7)
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| JP2005164995A JP4246181B2 (en) | 2005-06-06 | 2005-06-06 | Pseudomonas RRj228 for supporting growth of agricultural crops and microbial preparation containing the same |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| JP2005164995A JP4246181B2 (en) | 2005-06-06 | 2005-06-06 | Pseudomonas RRj228 for supporting growth of agricultural crops and microbial preparation containing the same |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| JP2006333829A true JP2006333829A (en) | 2006-12-14 |
| JP4246181B2 JP4246181B2 (en) | 2009-04-02 |
Family
ID=37554910
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| JP2005164995A Expired - Fee Related JP4246181B2 (en) | 2005-06-06 | 2005-06-06 | Pseudomonas RRj228 for supporting growth of agricultural crops and microbial preparation containing the same |
Country Status (1)
| Country | Link |
|---|---|
| JP (1) | JP4246181B2 (en) |
Cited By (9)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| KR101086367B1 (en) | 2010-03-11 | 2011-11-23 | 강원대학교산학협력단 | Bacillus aryabhattai LS9 promoting the growth of plants and use thereof |
| JP2011250729A (en) * | 2010-06-01 | 2011-12-15 | Hokkaido Univ | Slope greening method |
| JP2013514805A (en) * | 2009-12-22 | 2013-05-02 | ラントメネン・バイオアグリ・アーベー | Novel fluorescent Pseudomonas species of the Pseudomonas azotoformans species for enhancing budding and growth of plants |
| JP2013542987A (en) * | 2010-11-16 | 2013-11-28 | ユニバーシティー オブ デラウェア | Compositions and methods for improving rice growth and limiting arsenic uptake |
| JP2015530093A (en) * | 2012-09-19 | 2015-10-15 | バイオディスカバリー・ニュージーランド・リミテッド | Methods for screening microorganisms that confer beneficial properties on plants |
| JP2017002025A (en) * | 2015-06-12 | 2017-01-05 | 株式会社前川製作所 | Novel agricultural use for pseudomonas bacteria |
| JP2019156820A (en) * | 2018-03-06 | 2019-09-19 | イビデン株式会社 | Growth promoter of tomato fruit and improver of functional component content in tomato fruit, and, production method of growth promoter of tomato fruit and improver of functional component content in tomato fruit |
| CN110300514A (en) * | 2017-01-03 | 2019-10-01 | 孟山都技术公司 | Microbial composite and method |
| CN115895954A (en) * | 2022-11-15 | 2023-04-04 | 江苏省中国科学院植物研究所 | Pseudomonas laurophilus sulfate CNBG-PGPR-8 and application thereof |
-
2005
- 2005-06-06 JP JP2005164995A patent/JP4246181B2/en not_active Expired - Fee Related
Cited By (23)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JP2013514805A (en) * | 2009-12-22 | 2013-05-02 | ラントメネン・バイオアグリ・アーベー | Novel fluorescent Pseudomonas species of the Pseudomonas azotoformans species for enhancing budding and growth of plants |
| KR101086367B1 (en) | 2010-03-11 | 2011-11-23 | 강원대학교산학협력단 | Bacillus aryabhattai LS9 promoting the growth of plants and use thereof |
| JP2011250729A (en) * | 2010-06-01 | 2011-12-15 | Hokkaido Univ | Slope greening method |
| JP2013542987A (en) * | 2010-11-16 | 2013-11-28 | ユニバーシティー オブ デラウェア | Compositions and methods for improving rice growth and limiting arsenic uptake |
| JP2018121649A (en) * | 2012-09-19 | 2018-08-09 | バイオディスカバリー・ニュージーランド・リミテッド | Method for screening microorganism imparting beneficial properties to plant |
| JP2015530093A (en) * | 2012-09-19 | 2015-10-15 | バイオディスカバリー・ニュージーランド・リミテッド | Methods for screening microorganisms that confer beneficial properties on plants |
| US11466266B2 (en) | 2012-09-19 | 2022-10-11 | Bioconsortia, Inc. | Accelerated directed evolution of microbial consortia for the development of desirable plant phenotypic traits |
| JP7423883B2 (en) | 2012-09-19 | 2024-01-30 | バイオコンソーティア・インコーポレイテッド | Screening method for microorganisms that confer beneficial properties to plants |
| US11866698B2 (en) | 2012-09-19 | 2024-01-09 | Bioconsortia, Inc. | Accelerated directed evolution of microbial consortia for the development of desirable plant phenotypic traits |
| US10526599B2 (en) | 2012-09-19 | 2020-01-07 | Biodiscovery New Zealand Limited | Accelerated directed evolution of microbial consortia for the development of desirable plant phenotypic traits |
| JP2023089056A (en) * | 2012-09-19 | 2023-06-27 | バイオディスカバリー・ニュージーランド・リミテッド | Methods of screening for microorganisms that impart beneficial properties to plants |
| JP2020156489A (en) * | 2012-09-19 | 2020-10-01 | バイオディスカバリー・ニュージーランド・リミテッド | Methods of screening for microorganisms that impart beneficial properties to plants |
| US10900029B2 (en) | 2012-09-19 | 2021-01-26 | Bioconsortia New Zealand Limited | Methods of screening for microorganisms that impart beneficial properties to plants |
| JP2022000048A (en) * | 2012-09-19 | 2022-01-04 | バイオディスカバリー・ニュージーランド・リミテッド | Screening method of microorganism in which useful properties are imparted to plant |
| JP2017002025A (en) * | 2015-06-12 | 2017-01-05 | 株式会社前川製作所 | Novel agricultural use for pseudomonas bacteria |
| CN110300514B (en) * | 2017-01-03 | 2023-04-07 | 孟山都技术公司 | Microbial compositions and methods |
| EP3565397A4 (en) * | 2017-01-03 | 2020-06-24 | Monsanto Technology LLC | Microbial compositions and methods |
| CN110300514A (en) * | 2017-01-03 | 2019-10-01 | 孟山都技术公司 | Microbial composite and method |
| JP7132785B2 (en) | 2018-03-06 | 2022-09-07 | イビデン株式会社 | Tomato fruit growth promoter, functional ingredient content improver in tomato fruit, and method for producing tomato fruit growth promoter and functional ingredient content improver in tomato fruit |
| JP7354382B2 (en) | 2018-03-06 | 2023-10-02 | イビデン株式会社 | Tomato fruit growth promoter and functional component content improver in tomato fruit, and method for producing tomato fruit growth promoter and functional component content improver in tomato fruit |
| JP2019156820A (en) * | 2018-03-06 | 2019-09-19 | イビデン株式会社 | Growth promoter of tomato fruit and improver of functional component content in tomato fruit, and, production method of growth promoter of tomato fruit and improver of functional component content in tomato fruit |
| CN115895954A (en) * | 2022-11-15 | 2023-04-04 | 江苏省中国科学院植物研究所 | Pseudomonas laurophilus sulfate CNBG-PGPR-8 and application thereof |
| CN115895954B (en) * | 2022-11-15 | 2023-07-07 | 江苏省中国科学院植物研究所 | Pseudomonas lauroyl sulfate CNBG-PGPR-8 and application thereof |
Also Published As
| Publication number | Publication date |
|---|---|
| JP4246181B2 (en) | 2009-04-02 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| JP4246181B2 (en) | Pseudomonas RRj228 for supporting growth of agricultural crops and microbial preparation containing the same | |
| US9580363B2 (en) | Nitrogen-fixing bacterial inoculant for improvement of crop productivity and reduction of nitrous oxide emission | |
| Paulucci et al. | Arachis hypogaea PGPR isolated from Argentine soil modifies its lipids components in response to temperature and salinity | |
| CN110184220B (en) | Efficient phosphate and potassium solubilizing bacterium and application thereof | |
| HU230555B1 (en) | Environment-friend micro-organism produce and producing thereof | |
| US20120192605A1 (en) | Fertilizer composition and method | |
| Dahiya et al. | Resource partitioning in the rhizosphere by inoculated Bacillus spp. towards growth stimulation of wheat and suppression of wild oat (Avena fatua L.) weed | |
| Patel et al. | Growth promotion and biocontrol activity of Nocardiopsis dassonvillei strain YM12: an isolate from coastal agricultural land of Khambhat | |
| JP4102388B2 (en) | Microbial Bacillus KR083 that promotes the growth of crops and microbial preparations containing the same | |
| CN100417719C (en) | Pseudomonas RRj228 helpful for crop growth and microbe preparation containing the same | |
| Zheng et al. | Isolation and characterization of novel bacteria containing ACC Deaminase from the rhizosphere resource on Dry-Farming Lands | |
| JP6236660B2 (en) | Plant growth promoter | |
| KR100577717B1 (en) | 07691049 Bacillus megaterium KR076KACC91049 promoting the growth of crops and the microbial agent containing the same | |
| JP2019165676A (en) | Plant disease control agent and method of controlling plant disease | |
| JP2007153873A (en) | Soilborne disease controlling agent | |
| KR102085764B1 (en) | Pseudomonas plecoglossicida ANG14 having activities of promoting plant growth and controlling plant disease, and uses thereof | |
| KR102097042B1 (en) | Pseudarthrobacter equi ANG28 having activities of promoting plant growth and controlling plant disease, and uses thereof | |
| KR20220126123A (en) | Ignatzschineria strain producing auxin and promoting plant growth and uses thereof | |
| KR100646601B1 (en) | 083 91050 Bacillus sp. KR083KACC91050 promoting the growth of crops and the microbial agent containing the same | |
| JP2005151887A (en) | Agent and method for controlling soil-borne disease | |
| Siunova et al. | PGPR Pseudomonas strains promising for the development of bioformulations for plant protection and stimulation | |
| Helal et al. | Bioactive secondary metabolites extracted from the plant growth promoting bacteria Paraburkholderiatropica | |
| KR100646598B1 (en) | 181 Rhizobium sp. KR181 promoting the growth of crops and the microbial agent containing the same | |
| JP4340707B2 (en) | Method for producing filamentous fungal spores | |
| Xie et al. | Alleviating Continuous Monocropping Obstacle in Melon: Biological Elimination of Phenolic Acid |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| A131 | Notification of reasons for refusal |
Free format text: JAPANESE INTERMEDIATE CODE: A131 Effective date: 20080520 |
|
| A601 | Written request for extension of time |
Free format text: JAPANESE INTERMEDIATE CODE: A601 Effective date: 20080819 |
|
| A602 | Written permission of extension of time |
Free format text: JAPANESE INTERMEDIATE CODE: A602 Effective date: 20080822 |
|
| A601 | Written request for extension of time |
Free format text: JAPANESE INTERMEDIATE CODE: A601 Effective date: 20081014 |
|
| A602 | Written permission of extension of time |
Free format text: JAPANESE INTERMEDIATE CODE: A602 Effective date: 20081017 |
|
| TRDD | Decision of grant or rejection written | ||
| A01 | Written decision to grant a patent or to grant a registration (utility model) |
Free format text: JAPANESE INTERMEDIATE CODE: A01 Effective date: 20081209 |
|
| A01 | Written decision to grant a patent or to grant a registration (utility model) |
Free format text: JAPANESE INTERMEDIATE CODE: A01 |
|
| A61 | First payment of annual fees (during grant procedure) |
Free format text: JAPANESE INTERMEDIATE CODE: A61 Effective date: 20090107 |
|
| R150 | Certificate of patent or registration of utility model |
Ref document number: 4246181 Country of ref document: JP Free format text: JAPANESE INTERMEDIATE CODE: R150 Free format text: JAPANESE INTERMEDIATE CODE: R150 |
|
| FPAY | Renewal fee payment (event date is renewal date of database) |
Free format text: PAYMENT UNTIL: 20120116 Year of fee payment: 3 |
|
| FPAY | Renewal fee payment (event date is renewal date of database) |
Free format text: PAYMENT UNTIL: 20130116 Year of fee payment: 4 |
|
| R250 | Receipt of annual fees |
Free format text: JAPANESE INTERMEDIATE CODE: R250 |
|
| FPAY | Renewal fee payment (event date is renewal date of database) |
Free format text: PAYMENT UNTIL: 20130116 Year of fee payment: 4 |
|
| FPAY | Renewal fee payment (event date is renewal date of database) |
Free format text: PAYMENT UNTIL: 20140116 Year of fee payment: 5 |
|
| R250 | Receipt of annual fees |
Free format text: JAPANESE INTERMEDIATE CODE: R250 |
|
| R250 | Receipt of annual fees |
Free format text: JAPANESE INTERMEDIATE CODE: R250 |
|
| R250 | Receipt of annual fees |
Free format text: JAPANESE INTERMEDIATE CODE: R250 |
|
| R250 | Receipt of annual fees |
Free format text: JAPANESE INTERMEDIATE CODE: R250 |
|
| R250 | Receipt of annual fees |
Free format text: JAPANESE INTERMEDIATE CODE: R250 |
|
| R250 | Receipt of annual fees |
Free format text: JAPANESE INTERMEDIATE CODE: R250 |
|
| R250 | Receipt of annual fees |
Free format text: JAPANESE INTERMEDIATE CODE: R250 |
|
| R250 | Receipt of annual fees |
Free format text: JAPANESE INTERMEDIATE CODE: R250 |
|
| R250 | Receipt of annual fees |
Free format text: JAPANESE INTERMEDIATE CODE: R250 |
|
| R250 | Receipt of annual fees |
Free format text: JAPANESE INTERMEDIATE CODE: R250 |
|
| LAPS | Cancellation because of no payment of annual fees |