KR20220166411A - Root-knot nematode egg hatching rate inhibition microorganism neobacillus drentensis jc05 strain and use thereof - Google Patents
Root-knot nematode egg hatching rate inhibition microorganism neobacillus drentensis jc05 strain and use thereof Download PDFInfo
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Abstract
Description
본 발명은 네오바실러스 드렌텐시스(Neobacillus drentensis) JC05 균주에 관한 것이다.The present invention relates to Neobacillus drentensis JC05 strain.
뿌리혹선충은 토마토, 고구마, 참외, 오이 등 주요 작물의 뿌리에 기생하면서 혹처럼 덩어리를 형성하여 물과 영양분의 흡수, 이동 등을 방해한다. 이렇게 뿌리혹선충이 침입한 식물체는 수확량이 줄어드는 피해뿐 아니라 정상적인 생장이 어려워진다.Root knot nematodes parasitize on the roots of major crops such as tomatoes, sweet potatoes, melons, and cucumbers and form lumps like lumps to hinder the absorption and movement of water and nutrients. Plants infested with root knot nematodes not only suffer reduced yields but also have difficulty in normal growth.
최근에는 노지재배뿐만 아니라 시설재배에서도 뿌리혹선충이 많아져 농가에 큰 피해를 주고 있다. 이를 제거하기 위해 윤작, 객토, 태양열 소독, 담수 처리 등과 같은 물리적 방제를 비롯하여, 저항성 식물 품종 재배, 방선균 접종 등 화학적 또는 생물학적 방제법 등이 제안되는 추세이다.In recent years, root knot nematodes have increased not only in outdoor cultivation but also in facility cultivation, causing great damage to farmhouses. In order to eliminate it, physical control such as crop rotation, soil disinfection, solar heat disinfection, desalination, etc., as well as chemical or biological control methods such as cultivation of resistant plant varieties and inoculation of actinomycetes are being proposed.
그러나, 일부 화학적 방제는 인축에 대한 위험성, 잔류 독성, 환경 오염, 생태계 교란 등과 같은 다양한 문제점의 원인이 될 수 있어서, 보다 안전한 살선충제의 개발이 요구되고 있다.However, some chemical control may cause various problems such as danger to humans, residual toxicity, environmental pollution, ecosystem disturbance, etc., and therefore, there is a need to develop safer nematicides.
본 발명이 이루고자 하는 기술적 과제는 뿌리혹선충의 알 부화율을 억제시키는 데 효과적인 균주 및 이러한 균주를 이용해 뿌리혹선충의 알 부화율을 억제시키는 조성물 및 이를 이용한 뿌리혹선충 알 부화율 억제 방법을 제공하는 것이다.The technical problem to be achieved by the present invention is to provide a strain effective for inhibiting the hatching rate of root-knot nematodes, a composition for inhibiting the hatching rate of root-knot nematodes using the strain, and a method for inhibiting the egg hatching rate of root-knot nematodes using the same.
본 발명은 뿌리혹선충의 알 부화율을 억제시키는 데 효과적인 균주에 관한 것으로, 뿌리혹선충 알 부화 억제용 네오바실러스 드렌텐시스(Neobacillus drentensis) JC05(수탁 번호: KACC 92340P) 균주를 포함한다.The present invention relates to strains effective for inhibiting the egg hatching rate of root-knot nematodes, and includes Neobacillus drentensis JC05 (accession number: KACC 92340P) strain for inhibiting root-knot nematode egg hatching.
본 발명의 일 실시예에 있어서, 상기 균주는 서열번호 1로 표시되는 염기서열의 16S rRNA를 갖는 것을 특징으로 할 수 있다.In one embodiment of the present invention, the strain may be characterized by having 16S rRNA of the nucleotide sequence represented by SEQ ID NO: 1.
본 발명은 뿌리혹선충 알 부화 억제용 조성물에 관한 것으로, 뿌리혹선충 알 부화 억제용 네오바실러스 드렌텐시스(Neobacillus drentensis) JC05(수탁 번호: KACC 92340P) 균주, 상기 균주의 배양액, 또는 이들 모두를 유효성분으로 포함할 수 있다.The present invention relates to a composition for inhibiting root-knot nematode egg hatching, wherein Neobacillus drentensis JC05 (Accession Number: KACC 92340P) strain for inhibiting root-knot nematode egg hatching, a culture solution of the strain, or both active ingredients can be included as
본 발명의 일 실시예에 있어서, 상기 균주의 배양액은 부탄올에 의해 추출될 수 있다.In one embodiment of the present invention, the culture broth of the strain may be extracted with butanol.
본 발명은 뿌리혹선충 알 부화 억제 방법에 관한 것으로, 네오바실러스 드렌텐시스(Neobacillus drentensis) JC05(수탁 번호: KACC 92340P) 균주, 상기 균주의 배양액, 상기 균주를 포함하는 조성물, 및 상기 균주의 배양액을 포함하는 조성물로 이루어진 군에서 선택된 어느 하나 이상을 뿌리혹선충 알, 뿌리혹선충 알 주변, 식물 뿌리, 식물 뿌리 주변 또는 이들 모두에 처리하는 단계를 포함한다.The present invention relates to a method for inhibiting root-knot nematode egg hatching, and includes a Neobacillus drentensis JC05 (accession number: KACC 92340P) strain, a culture solution of the strain, a composition containing the strain, and a culture solution of the strain and treating root-knot nematode eggs, around root-knot nematode eggs, plant roots, around plant roots, or both, with at least one selected from the group consisting of compositions comprising:
본 발명의 일 실시예에 있어서, 상기 균주의 배양액은 부탄올에 의해 추출될 수 있다.In one embodiment of the present invention, the culture broth of the strain may be extracted with butanol.
본 발명의 일 실시예에 있어서, 상기 식물은 토마토 또는 오이일 수 있다.In one embodiment of the present invention, the plant may be tomato or cucumber.
본 발명의 일 실시예에 있어서, 상기 처리하는 단계에 있어서, 상기 균주, 상기 배양액, 및 상기 조성물을 상기 식물 뿌리에 접종하는 방법으로 처리하는 것을 포함할 수 있다.In one embodiment of the present invention, the treatment step may include treatment by a method of inoculating the plant root with the strain, the culture medium, and the composition.
본 발명은 뿌리혹선충 유충 사멸용 조성물에 관한 것으로, 네오바실러스 드렌텐시스(Neobacillus drentensis) JC05(수탁 번호: KACC 92340P) 균주, 상기 균주의 배양액, 또는 이들 모두를 유효성분으로 포함한다.The present invention relates to a composition for killing root-knot nematode larvae, which includes Neobacillus drentensis JC05 (Accession Number: KACC 92340P) strain, a culture solution of the strain, or both of them as active ingredients.
본 발명은 뿌리혹선충 유충 사멸 방법에 관한 것으로, 네오바실러스 드렌텐시스(Neobacillus drentensis) JC05(수탁 번호: KACC 92340P) 균주, 상기 균주의 배양액, 상기 균주를 포함하는 조성물, 및 상기 균주의 배양액을 포함하는 조성물로 이루어진 군에서 선택된 어느 하나 이상을 뿌리혹선충 알, 뿌리혹선충 알 주변, 식물 뿌리, 식물 뿌리 주변, 또는 이들 모두에 처리하는 단계를 포함한다.The present invention relates to a method for killing root-knot nematode larvae, including Neobacillus drentensis JC05 (Accession Number: KACC 92340P) strain, a culture solution of the strain, a composition containing the strain, and a culture solution of the strain and treating root-knot nematode eggs, around root-knot nematode eggs, plant roots, around plant roots, or both, with at least one selected from the group consisting of a composition to.
본 발명은 네오바실러스 드렌텐시스(Neobacillus drentensis) JC05 균주를 이용하여 고구마뿌리혹선충 알 부화율을 낮출 수 있다.The present invention can lower the hatching rate of sweet potato root knot nematode eggs using the Neobacillus drentensis JC05 strain.
도 1은 본 발명 균주의 16S rRNA 염기서열을 다른 네오바실러스(Neobacillus) 균주와 비교하여 나타낸 계통분석도이다.
도 2a는 본 발명의 일 실시예에 따라 JC05 균주 배양액의 헥산 추출물에 의한 뿌리혹선충 알 부화율 억제 효과를 나타낸 그래프이다.
도 2b는 본 발명의 일 실시예에 따라 JC05 균주 배양액의 부탄올 추출물에 의한 뿌리혹선충 알 부화율 억제 효과를 나타낸 그래프이다.
도 2c는 본 발명의 일 실시예에 따라 JC05 균주 배양액의 디클로로메탄 추출물에 의한 뿌리혹선충 알 부화율 억제 효과를 나타낸 그래프이다.
도 2d는 본 발명의 일 실시예에 따라 JC05 균주 배양액의 에틸아세테이트 추출물에 의한 뿌리혹선충 알 부화율 억제 효과를 나타낸 그래프이다.
도 3은 본 발명의 일 실시예에 따라 JC05 균주 배양액의 부탄올 추출물을 처리한 오이 유묘에서의 뿌리혹선충 난낭의 수를 나타낸 그래프이다.Figure 1 is a phylogenetic analysis showing the 16S rRNA nucleotide sequence of the strain of the present invention compared to other Neobacillus strains.
Figure 2a is a graph showing the effect of inhibiting the hatching rate of root-knot nematode eggs by the hexane extract of the JC05 strain culture medium according to an embodiment of the present invention.
Figure 2b is a graph showing the effect of inhibiting the hatching rate of root knot nematode eggs by the butanol extract of the JC05 strain culture medium according to an embodiment of the present invention.
Figure 2c is a graph showing the root-knot nematode egg hatching rate inhibition effect by the dichloromethane extract of the JC05 strain culture medium according to an embodiment of the present invention.
Figure 2d is a graph showing the root-knot nematode egg hatching rate inhibition effect by the ethyl acetate extract of the JC05 strain culture medium according to an embodiment of the present invention.
3 is a graph showing the number of root-knot nematode egg cysts in cucumber seedlings treated with the butanol extract of the JC05 strain culture medium according to an embodiment of the present invention.
이하, 첨부된 도면을 참조하여 본 발명에 따른 네오바실러스 드렌텐시스 균주, 상기 균주를 이용한 조성물, 상기 균주 및 조성물을 이용한 뿌리혹선충 알 부화 억제 방법을 상세히 설명하기로 한다.Hereinafter, a Neobacillus drentensis strain according to the present invention, a composition using the strain, and a method for inhibiting root-knot nematode egg hatching using the strain and composition will be described in detail with reference to the accompanying drawings.
이에 앞서, 본 명세서 및 청구범위에 사용된 용어는 사전적인 의미로 한정 해석되어서는 아니되며, 발명자는 자신의 발명을 최선의 방법으로 설명하기 위해 용어의 개념을 적절히 정의할 수 있다는 원칙에 입각하여, 본 발명의 기술적 사상에 부합되는 의미와 개념으로 해석되어야 한다.Prior to this, the terms used in this specification and claims should not be construed as limited in a dictionary sense, and based on the principle that the inventor can appropriately define the concept of terms in order to best explain his/her invention. , It should be interpreted as a meaning and concept consistent with the technical idea of the present invention.
상세한 설명, 도면들, 및 청구항들에서 상술하는 예시적인 실시 예들은 한정을 위한 것이 아니며, 다른 실시 예들이 이용될 수 있으며, 여기서 개시되는 기술의 사상이나 범주를 벗어나지 않는 한 다른 변경들도 가능하다.The illustrative embodiments set forth in the detailed description, drawings, and claims are not intended to be limiting, and other embodiments may be utilized, and other changes may be made without departing from the spirit or scope of the technology disclosed herein. .
따라서, 본 명세서에 기재된 실시 예 및 도면에 도시된 구성은 본 발명의 바람직한 실시 예에 불과할 뿐이고, 본 발명의 기술적 사상을 모두 표현하는 것은 아니므로, 본 출원 시점에 있어 이들을 대체할 수 있는 다양한 균등물과 변형 예들이 존재할 수 있음을 이해하여야 한다.Therefore, the embodiments described in this specification and the configurations shown in the drawings are only preferred embodiments of the present invention, and do not represent all of the technical spirit of the present invention, so various equivalents that can replace them at the time of the present application It should be understood that water and variations may exist.
당업자는 본 개시의 구성요소들, 즉 여기서 일반적으로 기술되고, 도면에 기재되는 구성요소들을 다양하게 다른 구성으로 배열, 구성, 결합, 및 도안할 수 있으며, 이것들의 모두는 명백하게 고안되며, 본 개시의 일부를 형성하고 있음을 용이하게 이해할 수 있을 것이다.One skilled in the art can arrange, construct, combine, and design the components of the present disclosure, i.e., the components generally described herein and illustrated in the drawings, in a variety of different configurations, all of which are expressly contemplated, and the present disclosure It will be readily understood that it forms part of the
본 발명은 네오바실러스 드렌텐시스(Neobacillus drentensis) JC05(수탁 번호: KACC 92340P) 균주를 제공한다. 상기 균주는 네오바실러스 드렌텐시스(Neobacillus drentensis) LMG 21831T 균주와 유사성이 99.2%인 염기서열의 16S rRNA를 가진다.The present invention provides a Neobacillus drentensis JC05 (Accession Number: KACC 92340P) strain. The strain has a 16S rRNA sequence with 99.2% similarity to Neobacillus drentensis LMG 21831 T strain.
본 발명은 네오바실러스 드렌텐시스(Neobacillus drentensis) JC05(수탁 번호: KACC 92340P), 상기 균주의 배양액, 또는 이들 모두를 유효성분으로 포함하는 뿌리혹선충 알 부화 억제용 조성물을 제공한다.The present invention provides a composition for inhibiting root-knot nematode egg hatching comprising Neobacillus drentensis JC05 (accession number: KACC 92340P), a culture solution of the strain, or both of them as active ingredients.
본 발명의 뿌리혹선충 알 부화 억제용 조성물은 상술한 본 발명의 균주, 상기 균주 배양액, 또는 이들의 혼합물에 당해 분야에서 뿌리혹선충 알 부화를 억제시킬 수 있다.The composition for inhibiting root-knot nematode egg hatching of the present invention can inhibit root-knot nematode egg hatching in the field in the above-described strain of the present invention, the strain culture medium, or a mixture thereof.
본 명세서에서 “배양액”은 균주를 배지에 접종하고 일정 시간 배양하여 얻은 결과물로서, 본 발명에 따른 균주를 적합한 액체 배지에서 배양한 배양액 자체, 상기 배양액을 여과 또는 원심분리하여 균주를 제거한 여액(여과액 또는 원심분리한 상등액), 상기 배양액을 초음파 처리하거나 상기 배양액에 용해효소(lysozyme)를 처리하여 수득한 세포 파쇄액, 상기 배양액, 여액, 및 파쇄액 등을 농축한 농축액 등을 포함하나 이에 한정되는 것은 아니다.In the present specification, "culture medium" is a result obtained by inoculating a strain into a medium and culturing for a certain period of time, the culture medium itself in which the strain according to the present invention is cultured in a suitable liquid medium, and the filtrate obtained by filtering or centrifuging the culture medium to remove the strain (filtration liquid or centrifuged supernatant), a cell lysate obtained by sonicating the culture medium or treating the culture medium with lysozyme, and a concentrate obtained by concentrating the culture medium, filtrate, and lysate, etc., but are limited thereto. it is not going to be
예를 들어, 배양액은 유기용매에 의해 추출될 수 있다. 본 발명의 일 실시예에 있어서, 배양액은 헥산(hexane), 부탄올(n-butanol), 디클로로메탄(dichloromethane), 또는 에틸아세테이트(ethyl acetate)에 의해 추출될 수 있다. 바람직하게는, 본 발명의 일 실시예에 따른 배양액은 부탄올(n-butanol)에 의해 추출될 수 있다.For example, the culture broth may be extracted with an organic solvent. In one embodiment of the present invention, the culture medium may be extracted with hexane, butanol, dichloromethane, or ethyl acetate. Preferably, the culture medium according to an embodiment of the present invention may be extracted with butanol (n-butanol).
또한, 헥산(hexane), 디클로로메탄(dichloromethane), 부탄올(n-butanol), 및 에틸아세테이트(ethyl acetate)에 의해 추출된 각각의 추출물은 1ppm 내지 1,000ppm 농도로 조절될 수 있다. 예를 들어, 헥산(hexane), 디클로로메탄(dichloromethane), 부탄올(n-butanol), 및 에틸아세테이트(ethyl acetate)에 의해 추출된 각각의 추출물은 10ppm 내지 1,000ppm 농도로 조절될 수 있다. 또한, 예를 들어, 헥산(hexane), 디클로로메탄(dichloromethane), 부탄올(n-butanol), 및 에틸아세테이트(ethyl acetate)에 의해 추출된 각각의 추출물은 100ppm 내지 1,000ppm 농도로 조절될 수 있다. 또한, 예를 들어, 헥산(hexane), 디클로로메탄(dichloromethane), 부탄올(n-butanol), 및 에틸아세테이트(ethyl acetate)에 의해 추출된 각각의 추출물은 500ppm 내지 1,000ppm 농도로 조절될 수 있다.In addition, each extract extracted by hexane, dichloromethane, butanol (n-butanol), and ethyl acetate may be adjusted to a concentration of 1 ppm to 1,000 ppm. For example, each extract extracted by hexane, dichloromethane, butanol (n-butanol), and ethyl acetate may be adjusted to a concentration of 10 ppm to 1,000 ppm. In addition, for example, each extract extracted by hexane, dichloromethane, butanol (n-butanol), and ethyl acetate may be adjusted to a concentration of 100 ppm to 1,000 ppm. In addition, for example, each extract extracted by hexane, dichloromethane, butanol (n-butanol), and ethyl acetate may be adjusted to a concentration of 500 ppm to 1,000 ppm.
또한, 상기 배양액은 고체배지 배양물, 및 상기 배양액의 건조물 또는 추출물 등의 배양물을 함유할 수 있으나, 이에 한정되는 것은 아니고, 나아가 적합한 부형제 또는 담체를 추가로 포함할 수도 있다.In addition, the culture solution may contain a solid medium culture, and a culture such as a dried product or extract of the culture solution, but is not limited thereto, and may further include a suitable excipient or carrier.
본 발명의 뿌리혹선충 알 부화 억제용 조성물은 액상 비료 형태로 제조될 수 있으며 이에 증량제를 첨가하여 가루 분말의 형태로 이용하거나 이를 제형화하여 과립화시킬 수도 있다. 그러나 그 제형에 특별히 한정되지는 않는다.The composition for inhibiting root-knot nematode egg hatching of the present invention may be prepared in the form of a liquid fertilizer, and may be used in the form of a powder by adding an extender thereto, or may be formulated and granulated. However, the formulation is not particularly limited.
뿌리혹선충은 길이 0.1mm 내지 1mm 정도의 작은 동물로 지구상 모든 곳에 서식한다. 토양 1m2 당 약 40만 마리로 토양생태계에 중요한 역할을 한다. 그 중 식물에 기생하는 선충류는 10%이고, 나머지 90%는 자유생활선충이다.Root-knot nematodes are small animals ranging in length from 0.1 mm to 1 mm and live everywhere on Earth. With about 400,000 birds per 1 m 2 of soil, they play an important role in the soil ecosystem. Among them, nematodes that parasitize plants account for 10%, and the remaining 90% are free-living nematodes.
식물기생성 선충은 전 세계 약 2,000여종이 있는데 작물의 뿌리, 줄기, 잎 등에 기생하여 양분의 이동통로를 막고 양분을 탈취해 피해를 준다. 특히 시설원예지에 연작으로 인한 선충피해가 많은데 20% 내지 30%의 수확량 감소를 가져올 수 있다.There are about 2,000 species of plant parasitic nematodes around the world, and they parasitize on the roots, stems, and leaves of crops to block the passage of nutrients and steal nutrients to cause damage. In particular, there is a lot of nematode damage due to continuous cropping in facility horticulture, which can lead to a yield reduction of 20% to 30%.
본 발명은 상기 뿌리혹선충 중 고구마뿌리혹선충이라 불리는 멜로이도기네 인코그니타(Meloidogyne incognita)를 대상으로 한다.The present invention targets Meloidogyne incognita , also called sweet potato root-knot nematode, among the root-knot nematodes.
멜로이도기네 인코그니타(Meloidogyne incognita)는 식물선충목 혹선충과의 선형동물로, 수컷의 몸길이 800㎛ 내지 1,400㎛, 암컷의 몸길이 400㎛ 내지 850㎛이다. 수컷이 암컷보다 크고 길며 꼬리가 둥글고, 암컷은 서양배 모양으로 둥글다. 암컷이 성숙하면 생식기 끝 부분에 알주머니(난낭)를 젤라틴으로 만들고 그 속에 알을 낳는다.Meloidogyne incognita ( Meloidogyne incognita ) is a linear animal belonging to the plant nematode order Knuckle Nematode, and has a male body length of 800 μm to 1,400 μm and a female body length of 400 μm to 850 μm. Males are larger and longer than females and have round tails, while females are pear-shaped and round. When a female matures, she makes an egg sac (oocyte) at the end of her genital organ into gelatin and lays eggs in it.
멜로이도기네 인코그니타(Meloidogyne incognita)의 애벌레(유충)는 몸길이 400㎛ 정도로 길고 구침이 잘 발달되어 뿌리 속으로 들어가거나 세포에서 즙을 빨아먹는 데에 유리하다. 주로 뿌리의 내부에 살면서 영양분을 빨아먹어 식물이 잘 자라지 않게 하고 다른 토양병해와 함께 2차 피해를 준다.The larvae (larvae) of Meloidogyne incognita are as long as 400㎛ in length and have well-developed stomata, which is advantageous for entering the root or sucking juice from the cells. It mainly lives inside the roots and sucks up nutrients, preventing plants from growing well and causing secondary damage along with other soil diseases.
뿌리혹선충의 피해를 받은 뿌리에는 크고 작은 혹이 생기고 이 혹들이 점차 많아지면 성장이 늦어지면서 시들기 시작한다.Large and small bumps form on the roots damaged by the root knot nematode, and when these bumps gradually increase, growth slows down and begins to wither.
종래의 방제 방법으로는 여름 땅 갈아엎기, 휴경, 담수, 윤작, 재식시기 조절, 유기질 비료 증시, 감염식물 제거, 저항성 또는 내성 품종 육종과 사용, 길항식물 심기, 유인 식물 심어 제거하기, 시비방법의 조절 등이 있다.Conventional control methods include summer land tillage, fallow, fresh water, crop rotation, planting period control, organic fertilizer stock, removal of infected plants, breeding and use of resistant or resistant varieties, planting antagonistic plants, planting and removing attracting plants, fertilization methods control, etc.
본 발명의 뿌리혹선충 알 부화 억제용 조성물은 상술한 본 발명의 균주, 상기 균주 배양액, 또는 이들의 혼합물에 당해 분야에서 뿌리혹선충 알 부화를 억제시키거나 뿌리혹선충 유충을 사멸시키는 용도로 사용되는 다른 임의의 성분을 추가로 포함할 수 있다. 일례로, 훈증제, 살선충제, 방선균(Actinomycetales)제, 항생물질로 이루어진 군에서 선택된 어느 하나 이상을 추가로 포함할 수 있다.The composition for inhibiting hatching of root-knot nematode eggs of the present invention is used in the above-described strain of the present invention, the strain culture medium, or a mixture thereof to inhibit root-knot nematode egg hatching or to kill root-knot nematode larvae in the field. It may further include the components of. For example, at least one selected from the group consisting of fumigants, nematicides, actinomycetales agents, and antibiotics may be further included.
상기 훈증제, 살선충제, 방선균제, 또는 항생물질은 화학물질 또는 박테리아의 또 다른 균주일 수 있고, 제초제, 살균제, 살충제, 비료, 또는 이들의 임의의 조합일 수 있다.The fumigant, nematicide, actinomycete, or antibiotic may be a chemical or another strain of bacteria, and may be a herbicide, fungicide, pesticide, fertilizer, or any combination thereof.
본 발명은 네오바실러스 드렌텐시스(Neobacillus drentensis) JC05(수탁 번호: KACC 92340P) 균주, 상기 균주의 배양액, 상기 균주를 포함하는 조성물, 및 상기 균주의 배양액을 포함하는 조성물로 이루어진 군에서 선택된 어느 하나 이상을 뿌리혹선충 알, 뿌리혹선충 알 주변, 식물 뿌리, 식물 뿌리 주변, 또는 이들 모두에 처리하는 단계를 포함하는 뿌리혹선충 알 부화 억제 방법을 제공한다.The present invention is any one selected from the group consisting of a Neobacillus drentensis JC05 (accession number: KACC 92340P) strain, a culture solution of the strain, a composition containing the strain, and a composition containing the culture solution of the strain Provided is a method for inhibiting root-knot nematode egg hatching comprising the step of treating the above to root-knot nematode eggs, around root-knot nematode eggs, plant roots, around plant roots, or both.
본 발명의 뿌리혹선충 알 부화 억제 방법을 적용하면 뿌리혹선충 알 부화를 억제시킬 수 있다.Root-knot nematode egg hatching can be inhibited by applying the root-knot nematode egg hatching method of the present invention.
용어 “식물”은 뿌리혹선충(Meloidogyne incognita)에 감염될 수 있는 개체이다. 뿌리혹선충이 감염될 수 있는 개체는 목, 속, 또는 종 등으로 세분화하기에는 지나치게 광범위하므로, 본 명세서에서는 상기 개체를 “식물”로 지칭하였다.The term “plant” refers to an organism that can be infected with the root knot nematode ( Meloidogyne incognita ). Since the objects that can be infected with root knot nematodes are too broad to be subdivided into orders, genera, or species, the objects are referred to as "plants" in the present specification.
상기 식물의 예로는 키위(kiwi fruit), 월하향(tuberose), 산세베리아(mother-in-law`s tongue), 알로에(aloe), 아마란스(amaranth), 캐슈넛(cashew nut), 망고(mango), 브라질 후추(Brazilian pepper), 딜(dill), 고수(coriander), 당근(carrot), 토란(taro), 코코얌(cocoyam), 야자(palms), 코코넛(coconut), 서양톱풀(yarrow), 금잔화(pot marigold), 치커리(chicory), 해바라기(sunflower), 선인장(lettuce), 양배추(cabbages, cauliflowers), 콜라비(kohlrabi), 파인애플(pineapple), 삼(hemp), 비트(beetroot), 시금치(spinach), 덩굴식물(bindweed), 고구마(sweet potato), 수박(watermelon), 멜론(melon), 오이(cucumber), 호박(pumpkin, marrow), 조롱박(cucurbit), 수세미(loofah), 참마(yam), 고무(rubber), 콩류(beans), 호두(walnuts), 라벤더(lavender), 로즈메리(rosemary), 아스파라거스(asparagus), 목화(cotton), 바나나(banana), 자스민(jasmine), 올리브(olive), 참깨(sesame), 후추(black pepper), 사탕수수(sugarcane), 쌀(rice), 강아지풀(green foxtail), 옥수수(maize), 자두(plum), 복숭아(peach), 커피(coffee), 석류(pomegranate), 피망(bell pepper), 담배(tobacco), 토마토(tomato), 가지(aubergine), 감자(potato), 포도 덩굴(grapevine), 강황(turmeric), 생강(ginger) 등이 있다.Examples of the plant include kiwi fruit, tuberose, mother-in-law's tongue, aloe, amaranth, cashew nut, mango, Brazilian pepper, dill, coriander, carrot, taro, cocoyam, palms, coconut, yarrow, marigold Pot marigold, chicory, sunflower, lettuce, cabbages, cauliflowers, kohlrabi, pineapple, hemp, beetroot, spinach spinach, bindweed, sweet potato, watermelon, melon, cucumber, pumpkin, marrow, cucurbit, loofah, yam ), rubber, beans, walnuts, lavender, rosemary, asparagus, cotton, banana, jasmine, olive ( olive, sesame, black pepper, sugar cane, rice, green foxtail, maize, plum, peach, coffee , pomegranate, bell pepper, tobacco, tomato, aubergine, potato, grapevine, turmeric, ginger, etc. .
본 발명의 일 실시예에 있어서, 식물은 토마토 또는 오이일 수 있다.In one embodiment of the present invention, the plant may be tomato or cucumber.
본 발명이 적용되는 식물의 부위는 제한되지 않는다. 따라서, 식물 전체는 물론, 식물의 부분, 예를 들어, 씨앗, 새순, 줄기, 뿌리, 잎, 과실, 및 이들의 전체는 물론 이들의 일부를 모두 포함한다. 다만, 뿌리혹선충의 주 생장 지점이 식물의 뿌리라는 점을 감안하면, 뿌리 및 뿌리 주변에 적용하는 것이 바람직하다.The part of the plant to which the present invention is applied is not limited. Thus, both whole plants as well as parts of plants such as seeds, shoots, stems, roots, leaves, fruits, and all of them as well as parts thereof are included. However, considering that the main growth point of the root-knot nematode is the root of a plant, it is preferable to apply it to the root and around the root.
본 발명의 뿌리혹선충 알 부화 억제 방법에 있어서, 상기 균주, 균주를 배양한 배양액, 및 상기 균주를 이용한 조성물을 종자나 식물에 침지, 관주, 접종, 또는 분무하여 수행할 수 있다.In the method for inhibiting hatching of root-knot nematode eggs of the present invention, the strain, a culture solution in which the strain is cultured, and a composition using the strain may be immersed, drenched, inoculated, or sprayed on seeds or plants.
접종하는 방법의 경우, 상기 균주, 균주를 배양한 배양액, 및 상기 균주를 이용한 조성물을 뿌리혹선충 알, 뿌리혹선충 알 주변, 식물 뿌리, 식물 뿌리 주변에 접종할 수 있다. 또한, 침지하는 방법의 경우, 배양액 및 제제를 식물 주변의 토양에 관주 또는 종자를 배양액 및 조성물에 담가 진탕 접종할 수 있다.In the case of the inoculation method, the strain, a culture solution in which the strain is cultured, and a composition using the strain may be inoculated around root-knot nematode eggs, root-knot nematode eggs, plant roots, and plant roots. In addition, in the case of the immersion method, the culture solution and the formulation may be drenched in the soil around the plant, or the seed may be immersed in the culture solution and the composition for shaking inoculation.
본 발명의 뿌리혹선충 알 부화 억제 방법이 적용되는 주변 영역이란 본 발명의 조성물이 효과를 발휘할 수 있는 범위를 의미하는 것으로, 지상 및 지하를 모두 포함한다. 예를 들어, 식물, 식물 부분, 또는 과실 주위의 약 2m 이내, 약 1m 이내, 약 70cm 이내, 약 50cm 이내, 약 25cm 이내, 약 10cm 이내, 또는 약 5cm 이내의 영역일 수 있다.The surrounding area to which the root-knot nematode egg hatching inhibition method of the present invention is applied means a range in which the composition of the present invention can exert an effect, and includes both aboveground and underground areas. For example, it may be an area within about 2 m, within about 1 m, within about 70 cm, within about 50 cm, within about 25 cm, within about 10 cm, or within about 5 cm around a plant, plant part, or fruit.
또한, 본 발명은 네오바실러스 드렌텐시스(Neobacillus drentensis) JC05(수탁 번호: KACC 92340P), 상기 균주의 배양액, 또는 이들 모두를 유효성분으로 포함하는 뿌리혹선충 유충 사멸용 조성물을 제공한다.In addition, the present invention provides a composition for killing root-knot nematode larvae containing Neobacillus drentensis JC05 (accession number: KACC 92340P), a culture solution of the strain, or both of them as active ingredients.
본 발명의 뿌리혹선충 유충 사멸용 조성물은 뿌리혹선충의 유충을 사멸시킬 수 있다.The composition for killing root-knot nematode larvae of the present invention can kill root-knot nematode larvae.
본 발명은 뿌리혹선충의 유충 사멸을 통하여 뿌리혹선충의 유충 수를 감소시키거나 뿌리혹선충에 의해 생기는 난낭의 수를 감소시킬 수 있다. 상기 난낭의 수는 뿌리혹선충 알주머니의 개수로서, 뿌리혹선충 유충과 함께 뿌리혹선충 감염의 척도로 사용될 수 있다.The present invention can reduce the number of root-knot nematode larvae or reduce the number of egg sacs produced by root-knot nematodes by killing root-knot nematode larvae. The number of egg sacs is the number of root-knot nematode egg sacs, and can be used as a measure of root-knot nematode infection together with root-knot nematode larvae.
본 발명의 뿌리혹선충 유충 사멸용 조성물은 액상 비료 형태로 제조될 수 있으며 이에 증량제를 첨가하여 가루 분말의 형태로 이용하거나 이를 제형화하여 과립화시킬 수도 있다. 그러나 그 제형에 특별히 한정되지는 않는다.The composition for killing root-knot nematode larvae of the present invention may be prepared in the form of a liquid fertilizer, and may be used in the form of a powder by adding an extender thereto, or may be formulated and granulated. However, the formulation is not particularly limited.
본 발명의 뿌리혹선충 유충 사멸용 조성물은 상술한 본 발명의 균주, 상기 균주 배양액, 또는 이들의 혼합물에 당해 분야에서 뿌리혹선충 유충을 사멸시키는 용도로 사용되는 다른 임의의 성분을 추가로 포함할 수 있다. 일례로, 훈증제, 살선충제, 방선균(Actinomycetales)제, 항생물질로 이루어진 군에서 선택된 어느 하나 이상을 추가로 포함할 수 있다.The composition for killing root-knot nematode larvae of the present invention may further include other optional components used in the field for killing root-knot nematode larvae in the above-described strain of the present invention, the strain culture medium, or a mixture thereof. . For example, at least one selected from the group consisting of fumigants, nematicides, actinomycetales agents, and antibiotics may be further included.
본 발명은 네오바실러스 드렌텐시스(Neobacillus drentensis) JC05(수탁 번호: KACC 92340P) 균주, 상기 균주의 배양액, 상기 균주를 포함하는 조성물, 및 상기 균주의 배양액을 포함하는 조성물로 이루어진 군에서 선택된 어느 하나 이상을 뿌리혹선충 알, 뿌리혹선충 알 주변, 뿌리혹선충 유충, 뿌리혹선충 유충 주변, 식물 뿌리, 식물 뿌리 주변, 또는 이들 모두에 처리하는 단계를 포함하는 뿌리혹선충 유충 사멸 방법을 제공한다.The present invention is any one selected from the group consisting of a Neobacillus drentensis JC05 (accession number: KACC 92340P) strain, a culture solution of the strain, a composition containing the strain, and a composition containing the culture solution of the strain It provides a method for killing root-knot nematode nematodes, including the step of treating the above to root-knot nematode eggs, root-knot nematode eggs, root-knot nematode larvae, root-knot nematode larvae, plant roots, plant roots, or both.
본 발명의 뿌리혹선충 유충 사멸 방법을 적용하면 뿌리혹선충 유충을 사멸시킬 수 있다.Root-knot nematode larvae can be killed by applying the root-knot nematode larva killing method of the present invention.
본 발명의 일 실시예에 있어서, 식물은 토마토 또는 오이일 수 있다.In one embodiment of the present invention, the plant may be tomato or cucumber.
본 발명의 뿌리혹선충 유충 사멸 방법에 있어서, 상기 균주, 균주를 배양한 배양액, 및 상기 균주를 이용한 조성물을 종자나 식물에 침지, 관주, 접종, 또는 분무하여 수행할 수 있다.In the root-knot nematode larvae killing method of the present invention, the strain, a culture solution in which the strain is cultured, and a composition using the strain may be immersed, drenched, inoculated, or sprayed on seeds or plants.
접종하는 방법의 경우, 상기 균주, 균주를 배양한 배양액, 및 상기 균주를 이용한 조성물을 뿌리혹선충 알, 뿌리혹선충 알 주변, 식물 뿌리, 식물 뿌리 주변에 접종할 수 있다. 또한, 침지하는 방법의 경우, 배양액 및 제제를 식물 주변의 토양에 관주 또는 종자를 배양액 및 조성물에 담가 진탕 접종할 수 있다.In the case of the inoculation method, the strain, a culture solution in which the strain is cultured, and a composition using the strain may be inoculated around root-knot nematode eggs, root-knot nematode eggs, plant roots, and plant roots. In addition, in the case of the immersion method, the culture solution and the formulation may be drenched in the soil around the plant, or the seed may be immersed in the culture solution and the composition for shaking inoculation.
본 발명의 뿌리혹선충 유충 사멸 방법이 적용되는 주변 영역이란 본 발명의 조성물이 효과를 발휘할 수 있는 범위를 의미하는 것으로, 지상 및 지하를 모두 포함한다. 예를 들어, 식물, 식물 부분, 또는 과실 주위의 약 2m 이내, 약 1m 이내, 약 70cm 이내, 약 50cm 이내, 약 25cm 이내, 약 10cm 이내, 또는 약 5cm 이내의 영역일 수 있다.The surrounding area to which the root-knot nematode larvae killing method of the present invention is applied means a range in which the composition of the present invention can exert an effect, and includes both ground and underground areas. For example, it may be an area within about 2 m, within about 1 m, within about 70 cm, within about 50 cm, within about 25 cm, within about 10 cm, or within about 5 cm around a plant, plant part, or fruit.
이하, 실시예를 통해서 본 발명을 보다 상세히 설명하기로 한다. 하지만, 이들은 본 발명을 보다 상세하게 설명하기 위한 것일 뿐 본 발명의 권리범위가 이에 한정되는 것은 아니다.Hereinafter, the present invention will be described in more detail through examples. However, these are only for explaining the present invention in more detail and the scope of the present invention is not limited thereto.
실시예 1: 미생물의 선별 및 동정Example 1: Selection and identification of microorganisms
본 발명의 JC05 균주는 분류 동정을 위해 16S rRNA 염기서열 분석을 진행하였고, 분석된 염기서열을 EZBioCloud에서 검색하였다. 또한, MEGA 프로그램의 네이버-조이닝(neighbor-joining) 알고리즘을 이용하여 계통도를 작성하였다.The JC05 strain of the present invention was subjected to 16S rRNA sequencing for classification identification, and the analyzed nucleotide sequence was searched in EZBioCloud. In addition, a systematic diagram was created using the neighbor-joining algorithm of the MEGA program.
도 1은 본 발명 균주의 16S rRNA 염기서열을 다른 네오바실러스 균주와 비교하여 나타낸 계통분석도이다.1 is a phylogenetic diagram showing the 16S rRNA nucleotide sequence of the strain of the present invention compared to other Neobacillus strains.
도 1을 참조하여 계통 분석을 진행한 결과, JC05 균주는 16S rRNA 염기서열에서 네오바실러스 드렌텐시스(Neobacillus drentensis) LMG 21831T 균주와 99.2%의 높은 상동성이 나타났으므로, JC05 균주는 네오바실러스 드렌텐시스(Neobacillus drentensis)로 동정되었다.As a result of the phylogenetic analysis with reference to FIG. 1, the JC05 strain showed a high homology of 99.2% with the Neobacillus drentensis LMG 21831 T strain in the 16S rRNA sequence, so the JC05 strain was Neobacillus It was identified as Neobacillus drentensis .
또한, 상기 네오바실러스 드렌텐시스(Neobacillus drentensis) JC05 균주를 농촌진흥청 국립농업과학원 미생물 은행에 기탁하여 수탁번호 KACC 92340P를 부여받았다.In addition, the Neobacillus drentensis JC05 strain was deposited at the Microorganism Bank of the National Institute of Agricultural Sciences of the Rural Development Administration and received accession number KACC 92340P.
실시예 2: 뿌리혹선충 알 부화 억제 효과Example 2: Root-knot nematode egg hatching inhibitory effect
뿌리혹선충(Meloidogyne incognita)의 접종원 준비를 위해 상토(부농)와 모래를 6:4(v/v)로 혼합하고 토마토('서광', 흥농종묘) 종자를 파종하였다. 파종된 토마토 종자를 25±4℃16시간 광조건, 8시간 암조건인 생장실에서 4주간 재배하였고, 뿌리혹선충(10,000eggs/plant)을 접종한 5주 내지 7주 후 혹이 형성된 뿌리를 접종원 준비에 사용하였다.To prepare the inoculum for the root-knot nematode ( Meloidogyne incognita ), soil (rich farmers) and sand were mixed at a ratio of 6:4 (v/v), and tomato ('Seokwang', Heungnong seedlings) seeds were sown. The sowed tomato seeds were grown for 4 weeks in a growth room under 25±4°C 16-hour light conditions and 8-hour dark conditions, and 5 to 7 weeks after inoculating root knot nematodes (10,000 eggs/plant), the roots with lumps were prepared as an inoculum. used in
감염된 토마토 뿌리는 흐르는 물에 씻어 흙을 제거한 뒤 1cm 크기로 잘라 0.5M 차아염소산나트륨(sodium hypochlorite) 용액을 넣고 믹서기로 2분 동안 갈아주었다. 분쇄한 뿌리를 0.63㎛, 0.45㎛, 및 0.25㎛체에 순차적으로 걸러 알을 분리하였다.Infected tomato roots were washed in running water to remove soil, cut into 1cm-sized pieces, and ground with a blender for 2 minutes with a 0.5M sodium hypochlorite solution. The crushed roots were sequentially sieved through 0.63 μm, 0.45 μm, and 0.25 μm sieves to separate eggs.
용매에 따른 미생물 네오바실러스 드렌텐시스(Neobacillus drentensis) JC05 균주의 추출물을 준비하기 위해, 네오바실러스 드렌텐시스(Neobacillus drentensis) JC05 균주를 TSB(Tryptic Soy Broth, Difco, USA) 배지에서 3일간 배양(28℃150rpm)하였다. 배양한 네오바실러스 드렌텐시스(Neobacillus drentensis) JC05 균주를 6,000rpm에서 20분간 원심분리하여 상청액을 여과(0.45㎛)하고, 유기용매(hexane, dichloromethane, n-butanol, ethyl acetate)로 각각 추출한 후 감압 농축하였다(Kwak 등, 2015).In order to prepare the extract of the microorganism Neobacillus drentensis JC05 strain according to the solvent, Neobacillus drentensis JC05 strain was cultured for 3 days in TSB (Tryptic Soy Broth, Difco, USA) medium ( 28° C. 150 rpm). The cultured Neobacillus drentensis JC05 strain was centrifuged at 6,000 rpm for 20 minutes, and the supernatant was filtered (0.45 μm), extracted with organic solvents (hexane, dichloromethane, n-butanol, ethyl acetate), and then reduced pressure. concentrated (Kwak et al., 2015).
각각의 추출물은 희석하여 다양한 농도(final concentration; 0ppm, 1ppm, 10ppm, 25ppm, 50ppm, 100ppm, 250ppm, 또는 500ppm)로 세분화하였고, 각각의 유기용매 및 다양한 농도로 제작된 추출물을 사용하여, 상기 추출물이 뿌리혹선충 알 부화율을 억제하는 효과가 있는지 검정하였다. 앞서 언급한 농도(0ppm, 1ppm, 10ppm, 25ppm, 50ppm, 100ppm, 250ppm, 또는 500ppm)는 Log[concentration(ug/mL)+1]로 변환하여 표시하였다. 상기 검정에 있어서, 뿌리혹선충의 알 부화율 억제를 평가하기 위해 희석한 네오바실러스 드렌텐시스(Neobacillus drentensis) JC05 균주의 추출물과 뿌리혹선충의 알을 혼합(1:1, v/v)하여 최종 1,500eggs/mL로 맞춘 후 3일 동안 현미경으로 알 부화율을 관찰하였다.Each extract was diluted and subdivided into various concentrations (final concentration; 0ppm, 1ppm, 10ppm, 25ppm, 50ppm, 100ppm, 250ppm, or 500ppm), and using each organic solvent and the extract prepared at various concentrations, the extract The effect of inhibiting the egg hatching rate of this root-knot nematode was tested. The aforementioned concentrations (0ppm, 1ppm, 10ppm, 25ppm, 50ppm, 100ppm, 250ppm, or 500ppm) were expressed as Log[concentration (ug/mL)+1]. In the assay, in order to evaluate the inhibition of egg hatchability of root-knot nematodes, the diluted extract of Neobacillus drentensis JC05 strain and the eggs of root-knot nematodes were mixed (1:1, v/v) to obtain a final 1,500eggs After adjusting to /mL, the egg hatchability was observed under a microscope for 3 days.
도 2a는 본 발명의 일 실시예에 따라 네오바실러스 드렌텐시스(Neobacillus drentensis) JC05 균주 배양액의 헥산 추출물에 의한 뿌리혹선충 알 부화율 억제 효과를 나타낸 그래프이다.Figure 2a is a graph showing the effect of inhibiting the hatching rate of root-knot nematode eggs by the hexane extract of Neobacillus drentensis JC05 strain culture medium according to an embodiment of the present invention.
도 2b는 본 발명의 일 실시예에 따라 네오바실러스 드렌텐시스(Neobacillus drentensis) JC05 균주 배양액의 부탄올 추출물에 의한 뿌리혹선충 알 부화율 억제 효과를 나타낸 그래프이다.Figure 2b is a graph showing the effect of inhibiting root-knot nematode egg hatching rate by butanol extract of Neobacillus drentensis JC05 strain culture medium according to an embodiment of the present invention.
도 2c는 본 발명의 일 실시예에 따라 네오바실러스 드렌텐시스(Neobacillus drentensis) JC05 균주 배양액의 디클로로메탄 추출물에 의한 뿌리혹선충 알 부화율 억제 효과를 나타낸 그래프이다.Figure 2c is a graph showing the effect of inhibiting root-knot nematode egg hatching rate by dichloromethane extract of Neobacillus drentensis JC05 strain culture medium according to an embodiment of the present invention.
도 2d는 본 발명의 일 실시예에 따라 네오바실러스 드렌텐시스(Neobacillus drentensis) JC05 균주 배양액의 에틸아세테이트 추출물에 의한 뿌리혹선충 알 부화율 억제 효과를 나타낸 그래프이다.Figure 2d is a graph showing the effect of inhibiting root-knot nematode egg hatchability by ethyl acetate extract of Neobacillus drentensis JC05 strain culture medium according to an embodiment of the present invention.
상기 도 2a 내지 도 2d를 참조하면, 다른 추출물에 비해, 부탄올(n-butanol) 추출물이 전체적으로 뿌리혹선충 알 부화율을 효과적으로 낮췄음을 확인할 수 있다.Referring to FIGS. 2A to 2D, it can be confirmed that the butanol (n-butanol) extract effectively lowered the hatching rate of root-knot nematode eggs as a whole, compared to other extracts.
더 자세하게는, 네오바실러스 드렌텐시스(Neobacillus drentensis) JC05 균주 배양액의 에틸아세테이트 추출물은 1ppm에서는 약 80%의 부화율이었고, 농도가 증가할수록 점차 감소하여 500ppm에서는 약 70%의 부화율을 기록했다. 상기 에틸아세테이트 추출물의 부화율 변화식은 하기 수학식 1과 같다.More specifically, Neobacillus drentensis ( Neobacillus drentensis ) The ethyl acetate extract of the JC05 strain culture medium had a hatching rate of about 80% at 1 ppm, and gradually decreased as the concentration increased, recording a hatching rate of about 70% at 500 ppm. The hatching rate change equation of the ethyl acetate extract is shown in
[수학식 1][Equation 1]
상기 수학식 1을 참조하면, 상기 에틸아세테이트 추출물의 부화율 변화 감소폭이 -3.91로 매우 작다는 것을 확인할 수 있다.Referring to
네오바실러스 드렌텐시스(Neobacillus drentensis) JC05 균주 배양액의 디클로로메탄 추출물은 1ppm에서는 약 78%의 부화율이었고, 농도가 증가할수록 점차 감소하여 500ppm에서는 약 70%의 부화율을 기록했다. 상기 디클로로메탄 추출물의 부화율 변화식은 하기 수학식 2와 같다.The dichloromethane extract of the Neobacillus drentensis JC05 strain culture medium had a hatching rate of about 78% at 1 ppm, and gradually decreased as the concentration increased, recording a hatching rate of about 70% at 500 ppm. The hatching rate change equation of the dichloromethane extract is shown in
[수학식 2][Equation 2]
상기 수학식 2를 참조하면, 상기 디클로로메탄 추출물의 부화율 변화 감소폭이 -4.03으로 작다는 것을 확인할 수 있다.Referring to
네오바실러스 드렌텐시스(Neobacillus drentensis) JC05 균주 배양액의 헥산 추출물은 1ppm에서는 약 80%의 부화율이었고, 농도가 증가할수록 비교적 가파르게 감소하여 500ppm에서는 약 55%의 부화율을 기록했다. 상기 헥산 추출물의 부화율 변화식은 하기 수학식 3과 같다.The hexane extract of Neobacillus drentensis JC05 strain culture medium had a hatching rate of about 80% at 1 ppm, and a relatively steep decrease as the concentration increased, recording a hatching rate of about 55% at 500 ppm. The hatching rate change equation of the hexane extract is shown in Equation 3 below.
[수학식 3][Equation 3]
상기 수학식 3을 참조하면, 상기 헥산 추출물의 부화율 변화 감소폭이 -10.47로 크다는 것을 확인할 수 있다.Referring to Equation 3, it can be seen that the reduction in hatching rate of the hexane extract is as large as -10.47.
네오바실러스 드렌텐시스(Neobacillus drentensis) JC05 균주 배양액의 부탄올 추출물은 1ppm에서는 약 75%의 부화율이었고, 농도가 증가할수록 가파르게 감소하여 500ppm에서는 42.5±0.7%의 부화율을 기록했다. 상기 부탄올 추출물의 부화율 변화식은 하기 수학식 4와 같다.Neobacillus drentensis ( Neobacillus drentensis ) The butanol extract of the JC05 strain culture medium had a hatching rate of about 75% at 1 ppm, and a steep decrease as the concentration increased, recording a hatching rate of 42.5 ± 0.7% at 500 ppm. The hatching rate change equation of the butanol extract is shown in Equation 4 below.
[수학식 4][Equation 4]
상기 수학식 4를 참조하면, 상기 부탄올 추출물의 부화율 변화 감소폭이 -11.36으로 상당히 크다는 것을 확인할 수 있다.Referring to Equation 4, it can be seen that the decrease in the hatching rate of the butanol extract is quite large at -11.36.
이에 따라, 추출 성분에 상관없이 농도가 증가할수록 알 부화율을 낮출 수 있었고, 실험한 추출 성분 중 헥산과 부탄올이 뛰어난 효과가 있음을 확인하였다. 헥산 추출물과 부탄올 추출물은 거의 유사한 감소 추세를 보이다가, 부탄올 추출물이 2.5Log(Concentration+1)를 지나는 시점부터 급격하게 감소하였다.Accordingly, it was confirmed that the egg hatching rate could be lowered as the concentration increased regardless of the extracted components, and that hexane and butanol among the extracted components tested had excellent effects. The hexane extract and the butanol extract showed almost similar decreasing trends, but the butanol extract rapidly decreased from the point of passing 2.5 Log (Concentration + 1).
따라서, 부탄올 추출물을 500ppm 농도로 사용하는 것이 뿌리혹선충 알 부화율을 현저히 낮출 수 있음을 확인하였다.Therefore, it was confirmed that using the butanol extract at a concentration of 500 ppm can significantly lower the hatching rate of root-knot nematode eggs.
실시예 3: 뿌리혹선충 억제 효과(뿌리혹선충 유충 사멸 효과)Example 3: Root-knot nematode inhibitory effect (root-knot nematode larvae killing effect)
오이('조은백다다기', 흥농종묘)를 파종하였다. 파종한 오이를 28±2℃, 16시간 광, 8시간 암조건인 생장실에서 3주간 재배하였다. 3주간 재배한 오이의 뿌리에 뿌리혹선충(Meloidogyne incognita)의 알을 500eggs/tube만큼 접종하였다. 접종 30분 후, 오이 뿌리에 부화율 억제 효과가 있었던 네오바실러스 드렌텐시스(Neobacillus drentensis) JC05 균주 배양액의 부탄올 추출물(100ppm)을 1mL씩 처리하였다. 추출물 처리 3주 후, 오이 뿌리에 있는 흙을 제거하고, 에리오글라우신 다이소듐(erioglaucine disodium) 용액(0.1mg/L)에 오이 뿌리를 30분간 침지하여 염색된 난낭의 수를 조사하였다(Ferris 등, 2012).Cucumbers ('Joeunbaekdadagi', Heungnong seedling) were sown. The sowed cucumbers were grown for 3 weeks in a growth room under conditions of 28±2℃, 16 hours of light and 8 hours of darkness. Root knot nematodes ( Meloidogyne incognita ) were inoculated into the roots of cucumbers grown for 3 weeks at 500eggs/tube. 30 minutes after inoculation, cucumber roots were treated with 1 mL of butanol extract (100 ppm) of Neobacillus drentensis JC05 strain culture, which had an effect of inhibiting hatching rate. After 3 weeks of extract treatment, soil on cucumber roots was removed, and cucumber roots were immersed in erioglaucine disodium solution (0.1 mg/L) for 30 minutes to examine the number of stained egg sacs (Ferris et al., 2012).
일반적으로, 식물의 뿌리는 선충집단을 활발하게 하거나 억제하는 복잡한 형태의 화합물을 방출하는데, 본 발명의 일 실시예에 있어서, 식물의 뿌리는 접종된 뿌리혹선충(Meloidogyne incognita) 알의 부화를 자극하고, 자극받은 상기 알에서 뿌리혹선충 유충이 나온다. 즉, 상기 추출물이 뿌리혹선충 알에 영향을 주는 것이 아닌, 부화된 뿌리혹선충 유충에 영향을 주는 것으로 해석하는 것이 바람직하다.In general, the roots of plants release complex types of compounds that activate or inhibit nematode populations. In one embodiment of the present invention, the roots of plants stimulate the hatching of inoculated root knot nematodes ( Meloidogyne incognita ) eggs and , Root-knot nematode larvae emerge from the stimulated eggs. That is, it is preferable to interpret that the extract does not affect root-knot nematode eggs, but affects hatched root-knot nematode larvae.
따라서, 실시예 3은 상기 추출물이 뿌리혹선충 유충을 사멸시킬 수 있는지를 확인할 수 있는 실험이다.Therefore, Example 3 is an experiment that can confirm whether the extract can kill the root-knot nematode larvae.
도 3은 본 발명의 일 실시예에 따라 네오바실러스 드렌텐시스(Neobacillus drentensis) JC05 균주 배양액의 부탄올 추출물을 처리한 오이 유묘에서의 뿌리혹선충 난낭의 수를 나타낸 그래프이다.3 is a graph showing the number of root-knot nematode egg sacs in cucumber seedlings treated with butanol extract of Neobacillus drentensis JC05 strain culture medium according to an embodiment of the present invention.
도 3을 참조하면, 네오바실러스 드렌텐시스(Neobacillus drentensis) JC05 균주 배양액의 부탄올 추출물이 오이 유묘에서 뿌리혹선충을 억제하는 효과가 있었다. 더 자세하게는, 상기 추출물을 처리하지 않은 대조군에서는 오이 뿌리에 생성된 난낭의 수가 평균 8.6±3.2개인 반면, 네오바실러스 드렌텐시스(Neobacillus drentensis) JC05 균주 배양액의 부탄올 추출물인 실험군에서는 오이 뿌리에 생성된 난낭의 수가 평균 4.9±1.6개로 감소하였다. 즉, 상기 부탄올 추출물이 뿌리혹선충 유충 일부를 사멸시켰고, 형성된 난낭의 수가 평균 8.6개에서 평균 4.9개로 현저히 감소했음을 확인하였다.Referring to Figure 3, Neobacillus drentensis ( Neobacillus drentensis ) The butanol extract of the JC05 strain culture medium had an effect of inhibiting root knot nematodes in cucumber seedlings. More specifically, in the control group not treated with the extract, the average number of egg sacs produced in cucumber roots was 8.6 ± 3.2, whereas in the experimental group, which was a butanol extract of Neobacillus drentensis JC05 strain culture medium, the number of egg sacs produced in cucumber roots was The number of egg sacs decreased to an average of 4.9±1.6. That is, it was confirmed that the butanol extract killed some of the root-knot nematode larvae, and the number of egg sacs formed significantly decreased from an average of 8.6 to an average of 4.9.
상술한 바와 같이 본 발명에서는 네오바실러스 드렌텐시스(Neobacillus drentensis) JC05(수탁 번호: KACC 92340P) 균주를 이용하여 뿌리혹선충 알 부화 억제용 및 유충 사멸용 조성물을 제공함으로서, 뿌리혹선충 알 부화율을 낮출 수 있고, 뿌리혹선충에 의해 생성되는 난낭의 수를 줄일 수 있다.As described above, the present invention provides a composition for inhibiting root-knot nematode egg hatching and killing larvae using Neobacillus drentensis JC05 (accession number: KACC 92340P) strain, thereby lowering the hatching rate of root-knot nematode eggs and may reduce the number of egg sacs produced by root-knot nematodes.
이상 설명한 내용을 통해 당업자라면 본 발명의 기술 사상을 일탈하지 아니하는 범위에서 다양한 변경 및 수정이 가능함을 알 수 있을 것이다. 따라서 본 발명의 기술적 범위는 명세서의 상세한 설명에 기재된 내용으로 한정되는 것이 아니며 특허 청구의 범위에 의해 정해져야만 할 것이다.Through the above description, those skilled in the art will know that various changes and modifications are possible without departing from the spirit of the present invention. Therefore, the technical scope of the present invention is not limited to the contents described in the detailed description of the specification and should be defined by the claims.
<110> REPUBLIC OF KOREA(RURAL DEVELOPMENT ADMINISTRATION) <120> ROOT-KNOT NEMATODE EGG HATCHING RATE INHIBITION MICROORGANISM NEOBACILLUS DRENTENSIS JC05 STRAIN AND USE THEREOF <130> FP-2104018-KR <160> 1 <170> KoPatentIn 3.0 <210> 1 <211> 1414 <212> RNA <213> Unknown <220> <223> 16S rRNA of Neobacillus drentensis JC05 <400> 1 agcggcggac gggtgagtaa cacgtgggca acctgcctgt aagactggga taacaccggg 60 aaaccggtgc taataccgga taatcctttt cctctcatga ggaaaagctg aaagtcggtt 120 tcggctgaca cttacagatg ggcccgcggc gcattagcta gttggtgagg taacggctca 180 ccaaggcgac gatgcgtagc cgacctgaga gggtgatcgg ccacactggg actgagacac 240 ggcccagact cctacgggag gcagcagtag ggaatcttcc acaatggacg aaagtctgat 300 ggagcaacgc cgcgtgagcg atgaaggcct tcgggtcgta aagctctgtt gttagggaag 360 aacaagtacc ggagtaactg ccggtacctt gacggtacct aaccagaaag ccacggctaa 420 ctacgtgcca gcagccgcgg taatacgtag gtggcaagcg ttgtccggaa ttattgggcg 480 taaagcgcgc gcaggcggtc ctttaagtct gatgtgaaag cccacggctc aaccgtggag 540 ggtcattgga aactggggga cttgagtgca gaagaggaaa gcggaattcc acgtgtagcg 600 gtgaaatgcg tagagatgtg gaggaacacc agtggcgaag gcggctttct ggtctgtaac 660 tgacgctgag gcgcgaaagc gtggggagca aacaggatta gataccctgg tagtccacgc 720 cgtaaacgat gagtgctaag tgttaggggg tttccgcccc ttagtgctgc agctaacgca 780 ttaagcactc cgcctgggga gtacggccgc aaggctgaaa ctcaaaggaa ttgacggggg 840 cccgcacaag cggtggagca tgtggtttaa ttcgaagcaa cgcgaagaac cttaccaggt 900 cttgacatcc tctgacactc ctagagatag gactttcccc ttcgggggac agagtgacag 960 gtggtgcatg gttgtcgtca gctcgtgtcg tgagatgttg ggttaagtcc cgcaacgagc 1020 gcaacccttg ttcttagttg ccagcattca gttgggcact ctaaggagac tgccggtgac 1080 aaaccggagg aaggtgggga tgacgtcaaa tcatcatgcc ccttatgacc tgggctacac 1140 acgtgctaca atggatggta caaagggcag caaagccgcg aggcctagcc aatcccataa 1200 aaccattctc agttcggatt gtaggctgca actcgcctac atgaagccgg aatcgctagt 1260 aatcgcggat cagcatgccg cggtgaatac gttcccgggc cttgtacaca ccgcccgtca 1320 caccacgaga gtttgtaaca cccgaagtcg gtggggtaac cgtaaggagc cagccgccta 1380 aggtgggaca gatgattggg gtgaagtcgt aaag 1414 <110> REPUBLIC OF KOREA (RURAL DEVELOPMENT ADMINISTRATION) <120> ROOT-KNOT NEMATODE EGG HATCHING RATE INHIBITION MICROORGANISM NEOBACILLUS DRENTENSIS JC05 STRAIN AND USE THEREOF <130> FP-2104018-KR <160> 1 <170> KoPatentIn 3.0 <210> 1 <211> 1414 <212> RNA <213> unknown <220> <223> 16S rRNA of Neobacillus drentensis JC05 <400> 1 agcggcggac gggtgagtaa cacgtgggca acctgcctgt aagactggga taacaccggg 60 aaaccggtgc taataccgga taatcctttt cctctcatga ggaaaagctg aaagtcggtt 120 tcggctgaca cttacagatg ggcccgcggc gcattagcta gttggtgagg taacggctca 180 ccaaggcgac gatgcgtagc cgacctgaga gggtgatcgg ccacactggg actgagacac 240 ggcccagact cctacggggag gcagcagtag ggaatcttcc acaatggacg aaagtctgat 300 ggagcaacgc cgcgtgagcg atgaaggcct tcgggtcgta aagctctgtt gttagggaag 360 aacaagtacc ggagtaactg ccggtacctt gacggtacct aaccagaaag ccacggctaa 420 ctacgtgcca gcagccgcgg taatacgtag gtggcaagcg ttgtccggaa ttatattgggcg 480 taaagcgcgc gcaggcggtc ctttaagtct gatgtgaaag cccacggctc aaccgtggag 540 ggtcattgga aactggggga cttgagtgca gaagaggaaa gcggaattcc acggttagcg 600 gtgaaatgcg tagagatgtg gaggaacacc agtggcgaag gcggctttct ggtctgtaac 660 tgacgctgag gcgcgaaagc gtggggagca aacaggatta gataccctgg tagtccacgc 720 cgtaaacgat gagtgctaag tgttaggggg tttccgcccc ttagtgctgc agctaacgca 780 ttaagcactc cgcctgggga gtacggccgc aaggctgaaa ctcaaaggaa ttgacggggg 840 cccgcacaag cggtggagca tgtggtttaa ttcgaagcaa cgcgaagaac cttaccaggt 900 cttgacatcc tctgacactc ctagagatag gactttcccc ttcgggggac agagtgacag 960 gtggtgcatg gttgtcgtca gctcgtgtcg tgagatgttg ggttaagtcc cgcaacgagc 1020 gcaacccttg ttcttagttg ccagcattca gttgggcact ctaaggagac tgccggtgac 1080 aaaccggagg aaggtgggga tgacgtcaaa tcatcatgcc ccttatgacc tgggctacac 1140 acgtgctaca atggatggta caaagggcag caaagccgcg aggcctagcc aatcccataa 1200 aaccattctc agttcggatt gtaggctgca actcgcctac atgaagccgg aatcgctagt 1260 aatcgcggat cagcatgccg cggtgaatac gttcccgggc cttgtacaca ccgcccgtca 1320 caccacgaga gtttgtaaca cccgaagtcg gtggggtaac cgtaaggagc cagccgccta 1380 aggtggggaca gatgattggg gtgaagtcgt aaag 1414
Claims (10)
상기 균주는 서열번호 1로 표시되는 염기서열의 16S rRNA를 갖는 것을 특징으로 하는 네오바실러스 드렌텐시스(Neobacillus drentensis) JC05(수탁 번호: KACC 92340P) 균주.According to claim 1,
The strain is characterized by having 16S rRNA of the nucleotide sequence represented by SEQ ID NO: 1 Neobacillus drentensis ( Neobacillus drentensis ) JC05 (accession number: KACC 92340P) strain.
상기 균주의 배양액은 부탄올에 의해 추출되는 뿌리혹선충 알 부화 억제용 조성물.According to claim 3,
The culture solution of the strain is a composition for inhibiting root-knot nematode egg hatching extracted by butanol.
상기 균주의 배양액은 부탄올에 의해 추출되는 뿌리혹선충 알 부화 억제 방법.According to claim 5,
Root-knot nematode egg hatching inhibition method in which the culture solution of the strain is extracted by butanol.
상기 식물은 토마토 또는 오이인 뿌리혹선충 알 부화 억제 방법.According to claim 5,
The method of inhibiting egg hatching of root knot nematodes in which the plant is tomato or cucumber.
상기 처리하는 단계에 있어서,
상기 균주, 상기 배양액, 및 상기 조성물을 상기 식물 뿌리에 접종 방법으로 처리하는 것을 포함하는 뿌리혹선충 알 부화 억제 방법.According to claim 5,
In the processing step,
A method for inhibiting hatching of root-knot nematode eggs, comprising treating the strain, the culture medium, and the composition with the method of inoculating the plant roots.
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