KR20220047146A - New variety of Chinese cabbage and its seeds - Google Patents
New variety of Chinese cabbage and its seeds Download PDFInfo
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- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01H—NEW PLANTS OR NON-TRANSGENIC PROCESSES FOR OBTAINING THEM; PLANT REPRODUCTION BY TISSUE CULTURE TECHNIQUES
- A01H6/00—Angiosperms, i.e. flowering plants, characterised by their botanic taxonomy
- A01H6/20—Brassicaceae, e.g. canola, broccoli or rucola
- A01H6/204—Brassica rapa
-
- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01H—NEW PLANTS OR NON-TRANSGENIC PROCESSES FOR OBTAINING THEM; PLANT REPRODUCTION BY TISSUE CULTURE TECHNIQUES
- A01H1/00—Processes for modifying genotypes ; Plants characterised by associated natural traits
- A01H1/02—Methods or apparatus for hybridisation; Artificial pollination ; Fertility
-
- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01H—NEW PLANTS OR NON-TRANSGENIC PROCESSES FOR OBTAINING THEM; PLANT REPRODUCTION BY TISSUE CULTURE TECHNIQUES
- A01H1/00—Processes for modifying genotypes ; Plants characterised by associated natural traits
- A01H1/10—Processes for modifying non-agronomic quality output traits, e.g. for industrial processing; Value added, non-agronomic traits
-
- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01H—NEW PLANTS OR NON-TRANSGENIC PROCESSES FOR OBTAINING THEM; PLANT REPRODUCTION BY TISSUE CULTURE TECHNIQUES
- A01H5/00—Angiosperms, i.e. flowering plants, characterised by their plant parts; Angiosperms characterised otherwise than by their botanic taxonomy
- A01H5/10—Seeds
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- Life Sciences & Earth Sciences (AREA)
- Health & Medical Sciences (AREA)
- Botany (AREA)
- Developmental Biology & Embryology (AREA)
- Environmental Sciences (AREA)
- Physiology (AREA)
- General Health & Medical Sciences (AREA)
- Genetics & Genomics (AREA)
- Natural Medicines & Medicinal Plants (AREA)
- Micro-Organisms Or Cultivation Processes Thereof (AREA)
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Abstract
Description
본 발명은 기능성 물질 함량이 증가되고, 항암활성이 높은 신품종 배추 및 이의 종자에 관한 것이다.The present invention relates to a new variety of Chinese cabbage having an increased content of functional substances and high anticancer activity, and seeds thereof.
배추(Brassica rapa ssp. pekinensis)는 십자화과 배추속에 속하는 식물로 중국이 원산지이다. 배추는 김치의 주원료로서 우리나라에서 가장 많이 이용되는 채소 중 하나이며, 수분 함량이 높고 섬유질과 비타민 A 및 C가 풍부하다. Chinese cabbage ( Brassica rapa ssp. pekinensis ) is a plant belonging to the genus Brassica rapa ssp. Chinese cabbage is one of the most used vegetables in Korea as the main ingredient of kimchi. It has a high water content and is rich in fiber and vitamins A and C.
배추의 잎은 나선형으로 착생되고 줄기가 단축되어 로제트 모양을 갖는다. 뿌리에서 나오는 잎은 옅은 녹색을 띠며 배추의 꽃은 짙은 황색을 띤 십자화관으로 1개의 암술 및 6개의 수술을 포함한다. 배추는 저온에서 꽃눈이 생긴 후 고온 장일이 되면 추대하는 특성을 갖는다.The leaves of Chinese cabbage are spirally epiphytic and the stem is shortened to have a rosette shape. The leaf coming out of the root is light green, and the flower of Chinese cabbage is a dark yellow cruciferous corolla, containing 1 pistil and 6 stamens. Chinese cabbage has the characteristic of budding at high temperature after flower buds are formed at low temperature.
배추의 품종은 결구 형태 또는 재배 기간에 따라 분류된다. 속이 꽉 차는 결구종은 잎이 중심부에 모이나 끝이 겹치지 않는 포합형과 양배추처럼 잎이 중앙 부위를 넘어 자라서 끝이 서로 덮이는 포피형으로 나누어진다.The varieties of Chinese cabbage are classified according to the shape of the tuber or the cultivation period. In the hyaluronic species, which is full, the leaves are gathered in the center, but the ends do not overlap, and the leaves are divided into the foreskin-type, in which the leaves grow beyond the central part and cover each other like cabbage.
또한, 배추, 양배추, 브로콜리 등에 함유된 물질인 글루코시놀레이트가 항암효과가 있음이 알려짐에 따라, 상기 글루코시놀레이트와 같은 기능성 물질이 많이 함유되고 항암활성을 지닌 신품종 개발이 진행되고 있다.In addition, as glucosinolate, a substance contained in Chinese cabbage, cabbage, broccoli, etc., is known to have anticancer effects, new varieties containing a lot of functional substances such as glucosinolate and anticancer activity are being developed.
본 발명의 목적은 글루코시놀레이트 함량이 증가되고, 항암 활성이 우수한 신품종 배추, 신품종 배추의 종자, 신품종 배추의 부분체 및 신품종 배추의 자손 식물체를 제공하는 것이다.An object of the present invention is to provide a new type of Chinese cabbage, seeds of a new type of Chinese cabbage, a part of a new type of Chinese cabbage, and progeny plants of a new type of Chinese cabbage, which have an increased glucosinolate content and have excellent anticancer activity.
본 발명의 다른 목적은 상기 신품종 배추 종자를 생산하는 방법을 제공하는 것이다.Another object of the present invention is to provide a method for producing the new variety of Chinese cabbage seeds.
본 발명의 일 실시예는 모본인 유지종 배추 및 부본인 박쵸이 배추를 교잡하여 수득된 신품종 배추종자로서, 상기 신품종 배추 종자는 수탁번호 KACC 98082P로 기탁된 것을 특징으로 하는 신품종 배추 종자이다.An embodiment of the present invention is a new variety of Chinese cabbage seeds obtained by hybridizing a parent oilseed cabbage and a second parent, Bakchoy cabbage, wherein the new variety of cabbage seed is a new variety of Chinese cabbage seed, characterized in that it is deposited under the accession number KACC 98082P.
상기 신품종 배추는 글루코시놀레이트 함량이 상기 모본 및 부본보다 증가된 것일 수 있다.The new variety of Chinese cabbage may have an increased glucosinolate content than the parent and secondary versions.
상기 모본인 유지종 배추는 노란 종피를 갖는 유전자원 YS-033(CGN06835) 계통이고, 상기 부본인 박쵸이 배추는 진갈색 종피를 갖는 유전자원 PC-099(CGN13924) 계통일 수 있다.The parent oilseed cabbage may be a genetic resource YS-033 (CGN06835) having a yellow seed coat, and the parent Bakchoy cabbage may be a genetic resource PC-099 (CGN13924) line having a dark brown seed coat.
상기 신품종 배추 종자는 서열번호 4로 표시될 수 있다.The new variety of Chinese cabbage seeds may be represented by SEQ ID NO: 4.
상기 신품종 배추 종자는 3-부테닐 이소티오시아네이트(3-Butenyl isothiocyanate), 4-펜테닐 이소티오시아네이트(4-pentenyl isothiocyanate), 2-페네틸 이소티오시아네이트(2-Phenethyl isothiocyanate) 및 설포라판(Sulforaphane)으로 이루어진 군에서 선택된 2종 이상의 함량이 부본인 박쵸이 배추보다 증가될 수 있다.The new varieties of Chinese cabbage seeds are 3-Butenyl isothiocyanate, 4-pentenyl isothiocyanate, 2-Phenethyl isothiocyanate, and The content of two or more selected from the group consisting of sulforaphane may be increased than that of Bakchoy cabbage, which is a subbonin.
상기 신품종 배추 종자는 항암 활성을 나타낼 수 있다.The new variety of Chinese cabbage seeds may exhibit anticancer activity.
상기 항암 활성은 대장암 또는 위암에 대한 항암 활성일 수 있다.The anticancer activity may be anticancer activity against colon cancer or stomach cancer.
본 발명의 다른 실시예는 상기 신품종 배추 종자로부터 수득된 신품종 배추이다.Another embodiment of the present invention is a new variety of Chinese cabbage obtained from the new variety of Chinese cabbage seeds.
본 발명의 또 다른 실시예는 상기 신품종 배추의 부분체이다.Another embodiment of the present invention is a part of the new variety of Chinese cabbage.
상기 부분체는 프로토플라스트(protoplast), 밑씨(ovule), 세포, 화분(pollen grain), 배아(embryo), 떡잎(cotyledon), 하배축(hypocotyledon), 뿌리, 분열조직(meristem), 꽃밥(anther), 줄기 및 잎으로 구성된 군에서 선택되는 1종 이상일 수 있다.The part is a protoplast, an ovule, a cell, a pollen grain, an embryo, a cotyledon, a hypocotyledon, a root, a meristem, an anther ), and may be at least one selected from the group consisting of stems and leaves.
본 발명의 또 다른 실시예는 상기 신품종 배추의 자손이다.Another embodiment of the present invention is the progeny of the new variety of Chinese cabbage.
본 발명의 또 다른 실시예는 a) 유지종 배추를 자가수분하여 모본을 수득하는 단계; b) 박쵸이 배추를 소포자배양하여 부본을 수득하는 단계; c) 상기 모본 및 부본을 교잡하여 잡종을 수득하는 단계; 및 d) 상기 잡종을 소포자배양하는 단계;를 포함하는, KACC 98082P로 기탁된 신품종 배추 종자 생산방법이다.Another embodiment of the present invention comprises the steps of: a) self-pollinating oilseed cabbage to obtain a model; b) culturing Bakchoy cabbage with small spores to obtain a copy; c) hybridizing the parent and parent to obtain a hybrid; And d) culturing the spores of the hybrid; it is a method for producing new varieties of cabbage seeds deposited as KACC 98082P, including.
상기 a) 단계의 자가수분은 3회 이상일 수 있다.The self-pollination in step a) may be three or more times.
본 발명에 따른 신품종 배추는 글루코시놀레이트 함량이 일반 배추보다 높아 항산화 및 항암 효과가 우수하다.The new variety of Chinese cabbage according to the present invention has a higher glucosinolate content than normal Chinese cabbage, and thus has excellent antioxidant and anticancer effects.
도 1은 신품종 배추 육종에 사용된 모본인 유지종 배추 및 부본인 박쵸이 배추의 표현형질을 나타낸 것이다.
도 2는 소포자 배양과정을 나타낸 모식도이다.
도 3은 유지종 배추 모본 및 박쵸이 배추 부본의 잡종을 소포자배양한 집단의 표현형질을 나타낸 것이다.
도 4는 배추 염기서열을 분석한 참고종인 지부 배추와 신품종 배추 육종에 사용된 모본 및 부분에 대한 글루코시놀레이트 함유량을 비교한 것이다.
도 5는 신품종 배추의 글루코시놀레이트 함유량을 나타낸 것이다.
도 6은 신품종 배추의 MYB29.1의 발현량을 나타낸 것이다.
도 7은 지부, 부본, 모본 및 신품종 배추의 MYB29.1 유전자, BraA03g003070.3C 유전자를 비교 분석한 것이다.
도 8은 부본(청경채, cheong-gyeong) 및 신품종 배추(DH-005)의 대장암 세포주 3종에 대한 세포독성 실험 결과(48시간)를 나타낸 도면이다.
도 9는 부본(청경채, cheong-gyeong) 및 신품종 배추(DH-005)의 대장암 세포주 3종에 대한 세포독성 실험 결과(72시간)를 나타낸 도면이다.
도 10은 부본(청경채, cheong-gyeong) 및 신품종 배추(DH-005)의 위암 세포주 3종에 대한 세포독성 실험 결과(48시간)를 나타낸 도면이다.
도 11은 부본(청경채, cheong-gyeong) 및 신품종 배추(DH-005)의 위암 세포주 3종에 대한 세포독성 실험 결과(72시간)를 나타낸 도면이다.1 shows the phenotypes of the parent type of oilseed cabbage and the secondary type, Bakchoy cabbage, used for breeding new varieties of cabbage.
Figure 2 is a schematic diagram showing the spore culture process.
Figure 3 shows the phenotype of a group cultured with small spores of a hybrid of a Chinese cabbage parent and Bakchoy cabbage parent.
Figure 4 is a comparison of the glucosinolate content of the parent and part used in the breeding of a new type of Chinese cabbage with the branch cabbage, which is a reference species for which the base sequence of the Chinese cabbage was analyzed.
Figure 5 shows the glucosinolate content of new varieties of Chinese cabbage.
6 shows the expression level of MYB29.1 in a new variety of Chinese cabbage.
7 is a comparative analysis of MYB29.1 gene and BraA03g003070.3C gene of branch, part, parent, and new varieties of Chinese cabbage.
8 is a view showing the cytotoxicity test results (48 hours) for three types of colorectal cancer cell lines of bubon (cheonggyeongchae, cheong-gyeong) and a new variety of Chinese cabbage (DH-005).
9 is a view showing the cytotoxicity test results (72 hours) for three types of colorectal cancer cell lines of bubon (cheonggyeongchae, cheong-gyeong) and a new variety of Chinese cabbage (DH-005).
10 is a view showing the cytotoxicity test results (48 hours) for three types of gastric cancer cell lines of bubon (cheonggyeongchae, cheong-gyeong) and a new variety of Chinese cabbage (DH-005).
11 is a view showing the cytotoxicity test results (72 hours) for three types of gastric cancer cell lines of bubon (cheonggyeongchae, cheong-gyeong) and a new variety of Chinese cabbage (DH-005).
이하, 본 발명이 속하는 기술 분야에서 통상의 지식을 가진 자가 용이하게 실시할 수 있도록 본 발명의 구체적인 실시예를 통하여 보다 상세하게 설명한다. 그러나 본 발명은 여러 가지 상이한 형태로 구현될 수 있으며 여기에서 설명하는 실시예에 한정되지 않는다.Hereinafter, specific embodiments of the present invention will be described in detail so that those of ordinary skill in the art can easily carry out the present invention. However, the present invention may be embodied in several different forms and is not limited to the embodiments described herein.
본 발명의 일 실시예는 모본인 유지종 배추 및 부본인 박쵸이 배추를 교잡하여 수득된 신품종 배추 종자로서, 상기 신품종 배추 종자의 수탁번호는 KACC 98082P이다.An embodiment of the present invention is a new type of Chinese cabbage seed obtained by hybridizing a parent oilseed cabbage and a secondary, Bakchoy cabbage, and the accession number of the new type of cabbage seed is KACC 98082P.
본 발명에서 상기 모본인 유지종 배추는 노란 종피를 갖는 유전자원 YS-033(CGN06835)일 수 있다. 상기 부본인 박쵸이 배추는 중국 배추 종 중의 하나로서, 청경채라고 불리기도 하며, 바람직하게는 진갈색 종피를 갖는 유전자원 PC-099(CGN13924) 계통일 수 있다.In the present invention, the parent oilseed cabbage may be a genetic resource YS-033 (CGN06835) having a yellow seed coat. The parent, Bakchoy cabbage, is one of Chinese cabbage species, and is also called bok choy, and may preferably be a genetic resource PC-099 (CGN13924) having a dark brown seed coat.
더욱 구체적으로 본 발명에서 사용된 모본은 상기 유지종 배추 YS-033(CGN06835)를 1회 이상, 바람직하게는 3회 자가수분한 제4세대에서 얻은 식물일 수 있으며, 본 발명에서 상기 식물은 "S4_YS-033"로 지칭될 수 있다.More specifically, the mother pattern used in the present invention may be a plant obtained from the fourth generation self-pollinating the oilseed cabbage YS-033 (CGN06835) one or more times, preferably three times, and in the present invention, the plant is " S4_YS-033".
또한 본 발명에서 사용된 부본은 상기 박쵸이 배추 PC-099(CGN13924)를 소포자배양을 통해 얻은 식물일 수 있으며, 본 발명에서 상기 식물은 "DH_PC-099"로 지칭될 수 있다.In addition, the part used in the present invention may be a plant obtained by sporophore culture of the Bakchoy cabbage PC-099 (CGN13924), and in the present invention, the plant may be referred to as "DH_PC-099".
본 발명의 신품종 배추 종자는 "S4_YS-033" 및 "DH_PC-099"를 교배하여 얻은 잡종으로부터 수득될 수 있다.The new variety of Chinese cabbage seeds of the present invention can be obtained from a hybrid obtained by crossing "S4_YS-033" and "DH_PC-099".
구체적으로 신품종 배추 종자는 상기 교배하여 얻은 잡종을 소포자배양한 식물체로부터 수득될 수 있으며, 본 발명의 상기 신품종 배추 종자 및 이의 식물체는 "유지청경채"로 명명될 수 있으며, BrYSP_DH_005로 지칭될 수 있다. Specifically, the new variety of Chinese cabbage seeds can be obtained from a plant obtained by culturing the hybrid obtained by the crossing, and the new variety of Chinese cabbage seed and its plant of the present invention may be called "oil and bok choy", and may be referred to as BrYSP_DH_005.
본 발명에 따른 신품종 배추 종자를 2020년 8월 3일자로 국립농업과학원에 기탁하였다(수탁번호: KACC 98082P).New varieties of cabbage seeds according to the present invention were deposited with the National Academy of Agricultural Sciences on August 3, 2020 (accession number: KACC 98082P).
본 발명에서 상기 "소포자배양"은 소포자를 인공배양하는 것을 말하며, 종자식물에 있어서는 화분배양이 이에 속한다. In the present invention, the term "spore culture" refers to artificial culture of small spores, and in the case of seed plants, pollen culture belongs to this.
본 발명에서 소포자 배양은 종래 공지된 방법에 따라 수행될 수 있다(J Plant Biotechnol, 41:116-122,2014, Seo, M. S. et al., 및 한국국제농업개발학회지, 24(4): 522-528, 서미숙 외).In the present invention, spore culture can be performed according to a conventionally known method (J Plant Biotechnol, 41: 116-122, 2014, Seo, M. S. et al., and Korean Journal of International Agricultural Development, 24(4): 522- 528, Seo Mi-sook et al.).
본 발명에서 상기 신품종 배추 종자는 서열번호 4로 표시될 수 있다.In the present invention, the new variety of Chinese cabbage seeds may be represented by SEQ ID NO: 4.
본 발명의 다른 실시예는 상기 수탁번호 KACC 98082P인 신품종 배추 종자로부터 수득된 신품종 배추일 수 있다.Another embodiment of the present invention may be a new type of Chinese cabbage obtained from the seed of the new type of cabbage, which is the accession number KACC 98082P.
본 발명에 따른 상기 신품종 배추는 글루코시놀레이트 함량이 상기 모본 및 부본보다 증가될 수 있고, 이에 따라 일반배추(지부배추)보다 글루코시놀레이트 함량이 높을 수 있다. 구체적으로, 상기 신품종 배추는 글루코시놀레이트 중 글루코라파닌 및 글루코나스터틴의 함량이 높아 기능성 품종 및 육종재료로의 활용도가 높다.In the new variety of Chinese cabbage according to the present invention, the glucosinolate content may be increased compared to the parent and secondary versions, and accordingly, the glucosinolate content may be higher than that of general Chinese cabbage (branch cabbage). Specifically, the new variety of Chinese cabbage has a high content of glucoraphanin and gluconastatin among glucosinolates, so that it has high utility as a functional variety and breeding material.
또한, 본 발명의 신품종 배추는 모본인 유지종 배추의 특징인 높은 글루코시놀레이트 함량과 부본인 박쵸이 배추의 특징인 넓은 엽형을 모두 가진 품종일 수 있다.In addition, the new variety of Chinese cabbage of the present invention may be a variety having both a high glucosinolate content, which is a characteristic of an oily cabbage, which is a parent, and a broad leaf shape, which is a characteristic of a parent, Bakchoy cabbage.
또한, 본 발명의 신품종 배추는 3-부테닐 이소티오시아네이트(3-Butenyl isothiocyanate), 4-펜테닐 이소티오시아네이트(4-pentenyl isothiocyanate), 2-페네틸 이소티오시아네이트(2-Phenethyl isothiocyanate) 및 설포라판(Sulforaphane)으로 이루어진 군에서 선택된 2종 이상의 함량이 부본인 박쵸이 배추보다 증가될 수 있다.In addition, the new Chinese cabbage of the present invention is 3-Butenyl isothiocyanate, 4-pentenyl isothiocyanate, 2-Phenethyl isothiocyanate (2-Phenethyl). The content of two or more selected from the group consisting of isothiocyanate) and sulforaphane may be increased than that of Bakchoi Chinese cabbage.
상기 3-부테닐 이소티오시아네이트(3-Butenyl isothiocyanate), 4-펜테닐 이소티오시아네이트(4-pentenyl isothiocyanate), 2-페네틸 이소티오시아네이트(2-Phenethyl isothiocyanate) 및 설포라판(Sulforaphane)은 글루코시놀레이트의 가수분해 산물일 수 있으며, 상기 가수분해는 가수분해효소(Myrosinase)에 의해 수행될 수 있다.The 3-Butenyl isothiocyanate, 4-pentenyl isothiocyanate, 2-Phenethyl isothiocyanate, and Sulforaphane may be a hydrolysis product of glucosinolate, and the hydrolysis may be performed by a hydrolase (Myrosinase).
상기 3-부테닐 이소티오시아네이트(3-Butenyl isothiocyanate), 4-펜테닐 이소티오시아네이트(4-pentenyl isothiocyanate), 2-페네틸 이소티오시아네이트(2-Phenethyl isothiocyanate) 및 설포라판(Sulforaphane)은 항암 효과를 나타낼 수 있고, 이에 따라 상기 신품종 배추 종자는 항암 활성을 나타낼 수 있다.The 3-Butenyl isothiocyanate, 4-pentenyl isothiocyanate, 2-Phenethyl isothiocyanate, and Sulforaphane may exhibit an anticancer effect, and thus the new variety of Chinese cabbage seeds may exhibit anticancer activity.
상기 항암 활성은 위암 또는 대장암에 대한 항암 활성일 수 있다. 본 발명의 일 실시예에서는 대장암 세포주 (HCT116, HT29, SW 480), 위암 세포주 (AGS, SNU-1, SNU-16)를 이용하여 신품종 배추의 항암활성을 확인하였다.The anticancer activity may be anticancer activity against gastric cancer or colon cancer. In an embodiment of the present invention, the anticancer activity of new varieties of Chinese cabbage was confirmed using colon cancer cell lines (HCT116, HT29, SW 480) and gastric cancer cell lines (AGS, SNU-1, SNU-16).
본 발명의 또 다른 실시예는 신품종 배추의 부분체 또는 이로부터 유래된 자손 식물체이다.Another embodiment of the present invention is a part of a new variety of Chinese cabbage or a progeny plant derived therefrom.
상기 부분체는 프로토플라스트(protoplast), 밑씨(ovule), 세포, 화분(pollen grain), 배아(embryo), 떡잎(cotyledon), 하배축(hypocotyledon), 뿌리, 분열조직(meristem), 꽃밥(anther), 줄기, 또는 잎일 수 있다. 상기 유래된 자손 식물체는 종자(seed), 또는 조직 배양물을 포함한다. 상기 배추 식물체로부터 수득되는 잎 및/또는 줄기는 김치, 나물, 샐러드 등 식재료로서 다양하게 이용될 수 있다. 상기 배추 식물체로부터 수득되는 종자 및/또는 세포는 다른 배추 개체, 계통, 또는 품종을 육성하는데 이용될 수 있다.The part is a protoplast, an ovule, a cell, a pollen grain, an embryo, a cotyledon, a hypocotyledon, a root, a meristem, an anther ), stem, or leaf. The derived progeny plants include seeds, or tissue cultures. The leaves and/or stems obtained from the Chinese cabbage plant can be used in various ways as ingredients such as kimchi, greens, and salad. The seeds and/or cells obtained from the Chinese cabbage plant can be used to grow other Chinese cabbage individuals, lines, or varieties.
본 발명의 또 다른 실시예는 a) 유지종 배추를 자가수분하여 모본을 수득하는 단계; b) 박쵸이 배추를 소포자배양하여 부본을 수득하는 단계; c) 상기 모본 및 부본을 교잡하여 잡종을 수득하는 단계; 및 d) 상기 잡종을 소포자배양하는 단계;를 포함하고, 수탁번호 KACC 98082P로 기탁된 것을 특징으로 하는 신품종 배추 종자 생산방법이다.Another embodiment of the present invention comprises the steps of: a) self-pollinating oilseed cabbage to obtain a model; b) culturing Bakchoy cabbage with small spores to obtain a copy; c) hybridizing the parent and parent to obtain a hybrid; And d) the step of culturing the small spores of the hybrid; it is a method for producing new varieties of cabbage seeds, characterized in that it is deposited under the accession number KACC 98082P.
상기 a) 단계에서 자가수분은 3회 이상일 수 있다.The self-pollination in step a) may be three or more times.
이하, 본 발명에 따른 구체적인 실시예를 들어 설명한다.Hereinafter, specific examples according to the present invention will be described.
실시예 1 : 소포자배양을 통한 신품종 배추 유지청경채 고정집단 육성Example 1: Cultivation of a fixed group of new varieties of Chinese cabbage with small spore culture
노란종피를 갖는 유전자원 YS-033(CGN06835) 유지종 배추를 3회 자가수분하여 얻은 제4세대 S4_YS-033을 육성하고 모본으로 활용하였으며, 계통 이름을 "LP08"로 명명하였다. 진갈색종피를 갖는 유전자원 PC-099(CGN13924) 박쵸이 배추를 소포자배양하여 DH_PC-099를 육성하고 부본으로 활용하였으며, 계통 이름을 "LP21"로 명명하였다(도 1). Genetic resource YS-033 (CGN06835) with yellow seed coat The 4th generation S 4 _YS-033 obtained by self-pollination of
상기 모본 및 부본을 교잡하여 얻은 미숙한 꼬투리(1.9~3.2mm)에서 고정된 식물체를 소포자 배양을 수행하여 2014년에서 2015년 까지, 유지청경채 고정집단 162계통을 육성하였으며(도 2), 소포자 배양은 재분화하기까지 16주 정도 소요되며, 종래 공지된 방법을 이용하였다(J Plant Biotechnol, 41:116-122,2014, Seo, M. S. et al., 및 한국국제농업개발학회지, 24(4): 522-528, 서미숙 외).From 2014 to 2015, from 2014 to 2015, 162 strains of a fixed group of oilseed bok choy were cultivated by culturing small spores in immature pods (1.9 to 3.2 mm) obtained by hybridizing the parent and parent. It takes about 16 weeks to redifferentiate, and a conventionally known method was used (J Plant Biotechnol, 41:116-122,2014, Seo, M. S. et al., and Journal of the Korean Society for International Agricultural Development, 24(4): 522) -528, Seo Mi-sook et al.).
구체적으로, 유지배추(LP08)와 박쵸이배추(LP21)를 교잡한 F1 종자를 파종하여 온실에서 3주간 키운 후, 4℃에서 24시간 광 조건으로 35일간 저온 처리하여 다시 온실에서 꽃분화를 유도하였다. 개화기 전 미숙 꽃봉오리의 회뢰를 수집하였다. 미숙 꽃봉오리의 크기는 암술머리보다 수술의 길이가 2mm 덜 자란 시기의 화뢰들을 채취하였다(화뢰 크기 1.9~3.2mm). 채취한 화뢰들은 1% 락스 용액에서 15분간 소독 후 멸균수로 3회 세정하여 무균상태로 만들었다. 소독한 화뢰들을 막자사발에 넣고 13% sucrose가 첨가된 B5 엑체배지를 화뢰 한 개 당 1ml씩 첨가하여 막자를 이용하여 부드럽게 갈아주었다. 나일론 망에 용액을 여과한 후 여과된 용약을 1000rpm에서 3분간 원심분리하고 상등액을 제거하였다. 동일량의 B5 액체 배지를 첨가하여 섞고, 1000rpm에서 3분간 원심분리 과정을 2회 더 하였다. 상등액을 제거하고 용기바닥에 있는 소포자들에 대하여 NLN 액체 배지를 첨가한 후 agarose 활성탄 용액을 혼합하였다. 소포자들은 32℃, 2일간 암배양한 후 25℃ 배양실에서 약 15일간 암배양하여 소포자로부터 배 발생을 유도하였다. 소포자에서 유도된 배들은 25℃배양기에서 30rpm으로 24시간 명조건에서 현탁배양한 후 MS 재분화 배지에 치상하여 재분화를 유도하였다. 재분화 유식물체는 배수성 검증을 위하여 어린잎의 일부를 채취한 후 DAPI 염색한 후 배수성 검증기(flow cytometry)를 이용하여 배수성 검증을 실시한다. 배수성 검증 결과, 이배체(2n)로 확인된 유식물체는 온실에서 재배하고, 뇌수분을 통해 종자를 확보하였다.Specifically, F1 seeds obtained by hybridizing oilseed cabbage (LP08) and Korean cabbage (LP21) were sown and grown for 3 weeks in a greenhouse. did Circumferentials of immature flower buds were collected before flowering. For the size of immature flower buds, flower buds were collected when the stamens were 2 mm shorter than the stigma (the size of the stigmas was 1.9 to 3.2 mm). The collected pyrotechnics were sterilized in 1% lacquer solution for 15 minutes and then washed three times with sterile water to make them sterile. Put the sterilized fireballs in a mortar, add 1ml of B5 broth to which 13% sucrose is added, and grind gently using a pestle. After filtering the solution through a nylon net, the filtered solution was centrifuged at 1000 rpm for 3 minutes, and the supernatant was removed. The same amount of B5 liquid medium was added and mixed, and the centrifugation process was further performed twice at 1000 rpm for 3 minutes. After removing the supernatant and adding NLN liquid medium to the spores at the bottom of the vessel, agarose activated carbon solution was mixed. The vesicles were cultured in the dark at 32°C for 2 days, and then incubated in a culture room at 25°C for about 15 days to induce embryogenesis from the small spores. Embryos induced from spores were suspended in a 25 ℃ incubator at 30 rpm under light conditions for 24 hours, and then re-differentiated by dentition in MS re-differentiation medium. For the redifferentiated plant, a part of young leaves is collected for verification of ploidy, dyed with DAPI, and then verified for ploidy using flow cytometry. As a result of the verification of ploidy, the seedlings identified as diploid (2n) were grown in a greenhouse, and seeds were secured through brain pollination.
종자를 채종한 각 계통에 대한 유전특성 등을 seedling 후 3~6주간 조사하였다. 소포자배양은 어린화분(n)을 배양하고, 배양과정 중에서 식물 스스로 안정화되면서 2n 체세포를 가짐으로 유전자 하나 하나에 대한 동형접합되므로 계통 한 개체 한 개체가 똑같은 형질을 나타내었다.The genetic characteristics of each line from which the seeds were harvested were investigated 3 to 6 weeks after seedling. In small spore culture, young pollen (n) is cultured, and the plant itself is stabilized during the cultivation process and has 2n somatic cells, so it is homozygous for each gene.
또한, 모본인 유지종 배추 엽형은 무엽형이고, 부본인 빅쵸이 배추 엽형은 결각이 없는 타원형 모양의 엽형이었으나, 신품종 배추인 유지청경채의 엽형은 162 계통간 매우 다양하게 나타났다(도 3).In addition, the leaf type of the parent oilseed cabbage leaf type was leafless, and the parent oil type cabbage leaf type had an oval leaf type without indentation, but the leaf type of the new type of cabbage, oilseed bok choy, was very diverse among 162 strains (FIG. 3).
실시예 2 : 글루코시놀레이트 함량 분석을 통한 유지청경채 선발Example 2: Selection of oily bok choy through glucosinolate content analysis
배추 종간 잡종(F1)의 소포자배양에 의한 dubled haploid 집단 66개 식물체 생성에 대하여 보고한 바 있다(J Plant Biotechnol 44:35-41, 2017, 서미숙 외). 최종적으로 유지청경채 162 계통을 육성하였다.It has been reported on the generation of 66 plants of a dubled haploid population by vesicle culture of a Chinese cabbage interspecies hybrid (F1) (J Plant Biotechnol 44:35-41, 2017, Misook Seo et al.). Finally, 162 fermented bok choy lines were bred.
배추 전체 게놈 염기서열 분석한 지부배추(Chinese cabbage, Chiffu, 결구배추)와 양친에 대한 글루코시놀레이트 합성량을 분석하였다. 배추 글루코시놀레이트 합성에 관련하여 11개의 서브물질이 분석되었으며, 배추의 대표적인 항산화물질은 글루코라파닌 및 글루코나스터틴이다. The amount of glucosinolate synthesis was analyzed for Chinese cabbage (Chiffu, Chinese cabbage) and parental sequencing of the entire genome of Chinese cabbage. 11 sub-materials related to the glucosinolate synthesis of Chinese cabbage were analyzed, and representative antioxidants of Chinese cabbage are glucoraphanin and gluconastatin.
또한 서브물질 중 프로고이트린은 같은 배추과 유채의 경우, 소 사료로 활용했을 때 갑상선암을 유발할 수 있다고 발표된 바 있어, 프로고이트린의 함량은 낮고, 두 개의 항산화 및 항암 물질의 함량이 높은 재조합 개체를 선발을 목표로 양친(모본 및 부본) 및 유지청경채 162 계통에 대한 글루코시놀레이트 함유량을 분석하였으며, 글루코시놀레이트 총 함유량과 11개 서브물질의 함유량을 분석한 결과를 도 4 및 도 5에 나타내었다.In addition, it has been announced that progoitrin among sub-materials can cause thyroid cancer when used as cattle feed in the same Chinese cabbage and rapeseed. The glucosinolate content was analyzed for the parents (mother and secondary) and 162 fermented bok choy lines with the aim of selecting shown in
분석 결과, 지부배추는 글루코시놀레이트 총 합성량이 8.39±0.69이었고, 모본 "LP08"은 47.55±3.66, 부본 박쵸이배추 "LP21"은 10.76±0.36으로 분석되었다. As a result of the analysis, the total amount of glucosinolate synthesis was 8.39±0.69 for Chinese cabbage, 47.55±3.66 for “LP08”, and 10.76±0.36 for “LP21” in Korean cabbage.
분석된 11개의 서브물질 중 글루코나스터틴은 지부배추의 함량이 0.14±0.09 보다 "LP08"이 0.47±0.14로 3.4배, "LP21"은 0.81±0.09로 5.8배 높았다. 글루코라파닌은 브로콜리 항산화물질인 설포라판의 전구물질로서, 지부배추 및 부본에는 검출되지 않았고, 모본에서만 0.16±0.05 검출되었다(도 4).Among the 11 sub-materials analyzed, the content of Chinese cabbage was 3.4 times higher in "LP08" than 0.14 ± 0.09, 0.47 ± 0.14, and 5.8 times higher in "LP21", 0.81 ± 0.09. As a precursor of sulforaphane, which is an antioxidant in broccoli, glucoraphanin was not detected in Chinese cabbage and Chinese cabbage, but was detected only in the mother plant by 0.16±0.05 ( FIG. 4 ).
이들 신품종 배추인 유지청경채 고정집단의 표현형질로 엽형은 무엽형, 배추엽형을 보이고, 모용(털)이 발달하지 않을 뿐 아니라, 측아가 많이 발달하여 생육 40일 경이면 쌈으로 활용할 수 있는 중간엽형이 50여 장에서 100여 장이 발달하는 쌈채소의 모습을 가지고 있었다.As the phenotype of the fixed group of these new varieties of Chinese cabbage, oilseed bok choy, the leaf type shows a leafless type and a cabbage leaf type, and the hair (hair) does not develop, and the lateral buds are developed a lot, so it is a mesenchymal type that can be used as a wrap around 40 days of growth. It had the appearance of ssam vegetables, which developed from 50 to 100 sheets.
상기 선발된 유지청경채 중 계통번호 005번은 일반배추에서는 거의 검출되지 않은 글루코라파닌 함량이 1.23±0.18로, 모본으로 이용한 유지배추보다 12배 높았으며, 이는 양배추종인 브로콜리 우수계통이 보통 1.1 umoles g 보다 더 높은 함량이다. 또한, 005번은 글루코시놀레이트 함유량이 44.12±2.86으로 일반배추보다 5배 높아 육종 모본으로 활용했을 때 기능성 품종을 개발할 수 있는 좋은 육종재료임을 확인할 수 있었으며, 배추엽형으로 쌈채소로도 이용 가능함을 확인하였다(도 5).Among the selected oil and bok choy, line No. 005 had a glucoraphanin content of 1.23±0.18, which was hardly detected in common cabbage, which was 12 times higher than that of the oily cabbage used as a parent, which is higher than that of the excellent line of broccoli, a cabbage species, usually 1.1 umoles g. higher content. In addition, No. 005 has a glucosinolate content of 44.12±2.86, which is 5 times higher than that of general cabbage, confirming that it is a good breeding material for developing functional varieties when used as a breeding model. was done (FIG. 5).
실시예 3 : 소포자 배양을 통한 유지청경재 BrYSP_DH_005 식물체의 유전자 발현 및 염기서열 분석Example 3: Analysis of gene expression and sequencing of BrYSP_DH_005 plants of oilseed bok choy by culturing small spores
실시예 2에서 선발된 계통번호 005번인 BrYSP_DH_005(DH005) 식물체에 대한 글루코시놀레이트 합성 관련 유전자들에 대한 발현량 분석을 수행하였다. 유전자 정량분석은 RNAeasy® Plant Mini Kit(Quiagen, Germany)에서 제공하는 방법에 따라 BrYSP_DH_005 식물체의 잎 100mg에서 RNA를 분리하였고, RNA 2μg에 DNAase I (Recombinant DNAase I, Takar, Shiga, Japan)을 처리하고 cDNA를 합성하였다(AmfiRivert II cDNA Synthesis Master Mix, GenDEOPT, Massachusetts, USA). 정량분석 PCR은 10배 희석한 cDNA를 사용하였다. 발현 대조구 유전자는 배추 GAPDH에 대한 상대적 발현량을 분석하였고, 3번 반복에 대한 평균값으로 분석하였다.Expression levels of glucosinolate synthesis-related genes were analyzed for the BrYSP_DH_005 (DH005) plant, which is phylogenetic No. 005, selected in Example 2. For gene quantitative analysis, RNA was isolated from 100 mg of leaves of BrYSP_DH_005 plants according to the method provided by RNAeasy ® Plant Mini Kit (Quiagen, Germany), and 2 μg of RNA was treated with DNAase I (Recombinant DNAase I, Takar, Shiga, Japan) and cDNA was synthesized (AmfiRivert II cDNA Synthesis Master Mix, GenDEOPT, Massachusetts, USA). For quantitative PCR, cDNA diluted 10-fold was used. Expression control gene was analyzed relative to Chinese cabbage GAPDH, and analyzed as the average value for 3 repetitions.
실시예 2에서 선발된 계통번호 005번인 BrYSP_DH_005 식물체의 경우, 배추 글루코시놀레이트 합성에 관여하는 여러 개 유전자 중에서 두 개 copies가 있는 MYB29.1 유전자의 발현량이 아주 높은 것을 알 수 있었다 (도 6).In the case of the BrYSP_DH_005 plant, which is phylogenetic No. 005, selected in Example 2, it was found that the expression level of the MYB29.1 gene having two copies among several genes involved in the synthesis of Chinese cabbage glucosinolate was very high (FIG. 6) .
실시예 2에서 선발된 계통번호 005번인 BrYSP_DH_005 식물체와 양친으로 이용한 배추 유전체를 염기서열 분석하였다. 양친으로 활용한 모본 "LP08"과 부본인 박쵸이배추 "LP21"과 실시예 2에서 선발된 계통번호 005번인 "BrYSP_DH_005" 배추의 게노믹 DNA를 분리하고 350bp 크기의 라이브러리를 작성하였다. 양 말단에 대하여 150bp 서열을 판독하였다. The genome sequence of the BrYSP_DH_005 plant, phylogenetic No. 005 selected in Example 2, and the Chinese cabbage genome used as the parent was sequenced. The genomic DNA of the parent “LP08” used as the parent and the parent “LP21” and the secondary “BrYSP_DH_005” cabbage selected in Example 2, phylogenetic No. 005, was isolated, and a 350 bp-sized library was prepared. 150bp sequences were read for both ends.
모본에 대해서 7억7천5백개, 부본에 대해서는 7억4백만개 리드에서 각 각 78.2 Gb와 71.1 Gb 서열을 획득하였다. BrYSP_DH_005 배추는 1억3천6백만개 리드에 대하여 13.2Gb의 서열을 확보하였다. 배추레퍼런스인 '지부'배추는 결구하는 배추로서 지금까지 3회 유전체 조립 서열을 개선하였고, 최종 보고된 '지부' 시퀀스는 B. rapa v3.0이다(Horticulture Research 5: 50, 2018, Zhang et al.).78.2 Gb and 71.1 Gb sequences were obtained from 775 million reads for the parent and 74 million reads for the minor, respectively. BrYSP_DH_005 Chinese cabbage secured a sequence of 13.2 Gb for 136 million reads. The 'Jibu' cabbage, which is a reference of Chinese cabbage, has improved the genome assembly sequence three times so far as a Chinese cabbage, and the final reported 'Jibu' sequence is B. rapa v3.0 (Horticulture Research 5: 50, 2018, Zhang et al. .).
도 7은 서열번호 1로 표시되는 지부의 MYB29.1유전자(BraA03g003070.3C), 서0열번호 2로 표시되는 부본 LP21의 MYB29.1유전자(BraA03g003070.3C), 서열번호 3으로 표시되는 모본 LP08의 MYB29.1유전자(BraA03g003070.3C) 및 서열번호 4로 표시되는 신품종 배추 DH005의 MYB29.1유전자(BraA03g003070.3C)를 비교 분석한 것이다.7 shows the MYB29.1 gene ( BraA03g003070.3C ) of the branch represented by SEQ ID NO: 1, the MYB29.1 gene ( BraA03g003070.3C ) of the sub - LP21 represented by SEQ ID NO: 2, and model LP08 represented by SEQ ID NO: 3 of MYB29.1 gene ( BraA03g003070.3C ) and MYB29.1 gene ( BraA03g003070.3C ) of the new Chinese cabbage DH005 represented by SEQ ID NO: 4 were comparatively analyzed.
배추 글루코시놀레이트 합성 관련유전자 10개, 32개 loci를 비교 분석하였고, 배추 레퍼런스와 다른 시퀀스를 보이는 여러 개의 유전자를 탐색할 수 있었고, 정량 PCR 분석으로 확인한 MYB29.1 유전자의 한 개 copy에서도 다른 서열을 확인하였다. 도 7을 살펴보면, 배추 MYB29.1 유전자의 한 개 copy인 BraA03g003070.3C유전자에서 글루코시놀레이트 합성량이 낮은 지부 레퍼런스와 박쵸이 배추 LP21과 비교하여 글루코시놀레이트 합성량이 높은 모본배추와 BrYSP_DH_005 식물체는 유전자 3개 bp 삽입('CTA') 및 161개 SNPs (Single Nucleotide polymorphisms)가 분석되었다. 또한 높은 글루코시놀레이트 합성하는 모계 LP08과 BrYSP_DH_005 유지청경채 배추는 SNP 서열 변이에 의해서 조기 종결 코돈이('TAG') 생성되었다(도 7).10 and 32 loci related to glucosinolate synthesis in Chinese cabbage were compared and analyzed, and several genes showing different sequences from the reference of Chinese cabbage were searched. The sequence was confirmed. Referring to FIG. 7, in the BraA03g003070.3C gene, which is one copy of the Chinese cabbage MYB29.1 gene, the parental Chinese cabbage and the BrYSP_DH_005 plant showed a high glucosinolate synthesis level compared to the branch reference with low glucosinolate synthesis and Bakchoy cabbage LP21. A 3 bp gene insertion ('CTA') and 161 SNPs (Single Nucleotide polymorphisms) were analyzed. In addition, early stop codon ('TAG') was generated by SNP sequence mutation in maternal LP08 and BrYSP_DH_005 oilseed bok choy cabbage, which synthesize high glucosinolate (FIG. 7).
실시예 4 : 글루코시놀레이트 가수분해 산물 함량 확인Example 4: Confirmation of glucosinolate hydrolysis product content
실시예 2에서 선발된 계통번호 005번인 BrYSP_DH_005(DH005) 식물체와 부본인 박쵸이 배추를 동결 건조하고, 동결 건조된 시료 분말 50mg을 1mL 증류수와 함께 1.5mL-마이크로 원심분리튜브에 현탁하고, 1mL 디클로로메탄을 첨가한 후, 실온에서 24시간 동안 빛이 없는 상태에서 내인성 미로시나아제를 첨가하여 가수분해를 진행한다.The BrYSP_DH_005 (DH005) plant, which is phylogenetic No. 005 (DH005) selected in Example 2, was freeze-dried, and 50 mg of the freeze-dried sample powder was suspended in a 1.5mL-microcentrifuge tube with 1mL distilled water, and 1mL dichloro After methane is added, hydrolysis is carried out by adding endogenous myrosinase in the absence of light at room temperature for 24 hours.
이후, 샘플을 12,000 x g에서 4분 동안 원심 분리하고 하부 유기층을 수집한다. 자동 샘플러(AOC20s, Shimadzu, Japan)와 모세관 컬럼(30 m × 0.32 mm × 0.25 μm DB-5, Agilent Technologies)이 장착 된 MS 검출기(GCMS-QP 2020NX, Shimadzu, Japan)에 연결된 가스 크로마토 그래피(GC, Nexis GC-2030, Shimadzu, Japan)를 사용하여 글루코시놀레이트 가수 분해 생성물을 측정하고, 이에 대한 결과를 하기 표 2에 나타내었다.Then, the sample is centrifuged at 12,000 x g for 4 min and the lower organic layer is collected. Gas chromatography (GC) connected to an MS detector (GCMS-QP 2020NX, Shimadzu, Japan) equipped with an autosampler (AOC20s, Shimadzu, Japan) and a capillary column (30 m × 0.32 mm × 0.25 μm DB-5, Agilent Technologies). , Nexis GC-2030, Shimadzu, Japan) was used to measure glucosinolate hydrolysis products, and the results are shown in Table 2 below.
이때, 디클로로메탄 1㎕ 추출물의 샘플을 1:1의 분할 비율로 GC-MS에 주입하였다. 초기 온도를 40℃로 2분 동안 유지 한 후, 오븐 온도를 15℃/min로 320℃까지 증가시켰다. 주입기 및 검출기 온도는 각각 260℃ 와 320℃로 설정하였고, 헬륨 운반 가스의 유속은 1.2mL/min로 설정하였다.At this time, a sample of 1 μl of dichloromethane extract was injected into GC-MS at a split ratio of 1:1. After maintaining the initial temperature at 40°C for 2 minutes, the oven temperature was increased to 320°C at 15°C/min. The injector and detector temperatures were set at 260°C and 320°C, respectively, and the flow rate of the helium carrier gas was set at 1.2 mL/min.
상기 표 2를 참조하면, 본 발명에 따른 신품종 배추의 경우, 부본과 비교할 때, 3-부테닐 이소티오시아네이트(3-Butenyl isothiocyanate), 4-펜테닐 이소티오시아네이트(4-pentenyl isothiocyanate), 2-페네틸 이소티오시아네이트(2-Phenethyl isothiocyanate) 및 설포라판(Sulforaphane)의 함량이 모두 현저하게 높은 것을 확인할 수 있다.Referring to Table 2, in the case of the new Chinese cabbage according to the present invention, when compared to the bobon, 3-Butenyl isothiocyanate, 4-pentenyl isothiocyanate (4-pentenyl isothiocyanate) , it can be seen that the content of 2-phenethyl isothiocyanate and sulforaphane are all remarkably high.
실시예 5 : 항암 활성 확인Example 5: Confirmation of anticancer activity
1. 실험 준비1. Experimental Preparation
배추 시료 전처리 과정을 위해 동결건조 상태 신품종 배추(DH-005) 및 부본 배추 시료(청경채, cheong-gyeong) 500mg을 각각 세포 배양 때 사용된 RPMI-1640 배양 배지 25mL를 넣고 초음파 추출을 1시간 동안 진행하였다. 이후, 시료에 남아있는 debris 등을 제거하기 위하여 0.45μm membrane 필터를 사용하여 여과하여 시료 추출물을 제조하였다.For the preparation of the Chinese cabbage sample, add 25 mL of RPMI-1640 culture medium used for cell culture of 500 mg of freeze-dried new Chinese cabbage (DH-005) and cheong-gyeong, respectively, and perform ultrasonic extraction for 1 hour. did Thereafter, to remove debris, etc. remaining in the sample, a sample extract was prepared by filtration using a 0.45 μm membrane filter.
시료 첨가 전날에 Hemocytometer or cell counter(logos biosystem)를 이용하여 하기 표 3의 대장암 및 위암 세포의 성장 속도를 감안하여 웰(well)당 5x103~2x104의 세포를 50㎕의 RPMI 1640 배양 배지를 이용하여 배양하였다. 이때, 세포가 생존 및 배양될 수 있는 적정 pH(7.4)를 유지하고, 세포의 호흡을 위한 완충재로 배양기 내부의 공기를 CO2 5%로 유지하며 온도는 37℃로 설정하였다. 각각의 세포 배양 시 KTCC의 세포 배양 설명서에 적힌 배양에 적합한 RPMI 1640 media에 Fetal bovine serum 10%를 첨가하여 배양용액으로 사용하였다.On the day before sample addition, using a hemocytometer or cell counter (logos biosystem), 5x10 3 ~ 2x10 4 cells per well in consideration of the growth rates of colorectal cancer and gastric cancer cells in Table 3 below were added to 50 μl of RPMI 1640 culture medium. was cultured using At this time, an appropriate pH (7.4) for cell survival and culture was maintained, and the air inside the incubator was maintained at 5% CO 2 as a buffer for cell respiration, and the temperature was set to 37°C. For each cell culture, 10% of Fetal bovine serum was added to RPMI 1640 media suitable for culture written in the cell culture manual of KTCC and used as a culture solution.
이후, 대장암 및 위암 세포에 시료 추출물을 농도 별로 처리(mg/ml - 10, 5, 2.5, 1.25, 0.625, 0.313, 0.156, 0.078, 0.039)하였다.Thereafter, the sample extracts were treated for each concentration of colon cancer and gastric cancer cells (mg/ml - 10, 5, 2.5, 1.25, 0.625, 0.313, 0.156, 0.078, 0.039).
(Colon cancer)colorectal cancer
(Colon cancer)
2. HT29
3. SW 4801. HCT116
2. HT29
3. SW 480
colorectal adenocarcinoma
Dukes' type B, colorectal adenocarcinomacolorectal carcinoma
colorectal adenocarcinoma
Dukes' type B, colorectal
2. SNU-1
3. SNU-161. AGS
2. SNU-1
3. SNU-16
gastric carcinoma
gastric carcinomagastric adenocarcinoma
gastric carcinoma
gastric carcinoma
2. 세포독성 실험(MTT assay)결과2. Cytotoxicity test (MTT assay) results
시료 추출물을 농도 별로 처리한 후, MTT[3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide] 용액(2mg/ml, Sigma-Aldrich)은 20㎕씩 첨가하여 37℃에서 2시간 배양하였다. After each concentration of the sample extract was treated, 20 μl of MTT [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide] solution (2mg/ml, Sigma-Aldrich) was added at 37°C. incubated for 2 hours.
MTT의 기본원리는 노란색의 수용성 MTT tetrazolium을 세포에 처리하면 대사 과정이 온전히 살아있는 세포의 미토콘드리아 탈수소효소에 의해 tetrazolium의 ring 구조가 끊어지면서 청자색을 띠는 비수용성의 MTT formazan 결정으로 환원되는 것이며, formazan을 MTT lysis Buffer로 녹인 색소의 발색 되는 정도를 암세포 생장 억제율로 나타낸다. The basic principle of MTT is that when yellow water-soluble MTT tetrazolium is treated in cells, the metabolic process is completely reduced to blue-violet water-soluble MTT formazan crystals as the ring structure of tetrazolium is cut by mitochondrial dehydrogenase of living cells. The degree of color development of the pigment dissolved in MTT lysis buffer is expressed as the rate of inhibition of cancer cell growth.
Dimethylformamide, SDS(sodium dodecyl sulfate)가 포함된 MTT lysis Buffer 100㎕/well 첨가 후 배양기에 넣고 다음 날 ELISA reader (Molecular Devices)를 이용해 570nm에서 흡광도를 측정하였다. After adding 100 μl/well of MTT lysis buffer containing dimethylformamide and sodium dodecyl sulfate (SDS), it was placed in an incubator and absorbance was measured at 570 nm using an ELISA reader (Molecular Devices) the next day.
이에 대한 결과를 도 8 및 9에 나타내었고, 이후 무처리군과 대비해 세포 생존 및 사멸도를 확인하여 배추시료 추출물의 48시간, 72시간의 항암 활성 농도(IC50, mg/mL)를 하기 표 4 및 표 5에 나타내었다.The results are shown in Figures 8 and 9, and then, compared to the untreated group, the cell viability and apoptosis were confirmed, and the anticancer activity concentration (IC50, mg/mL) of the Chinese cabbage sample extract for 48 hours and 72 hours (IC50, mg/mL) was shown in Table 4 and Table 5.
도 8은 부본(청경채, cheong-gyeong) 및 신품종 배추(DH-005)의 대장암 세포주 3종에 대한 세포독성 실험 결과(48시간)를 나타낸 도면, 도 9는 부본(청경채, cheong-gyeong) 및 신품종 배추(DH-005)의 대장암 세포주 3종에 대한 세포독성 실험 결과(72시간)를 나타낸 도면, 도 10은 부본(청경채, cheong-gyeong) 및 신품종 배추(DH-005)의 위암 세포주 3종에 대한 세포독성 실험 결과(48시간)를 나타낸 도면, 도 11은 부본(청경채, cheong-gyeong) 및 신품종 배추(DH-005)의 위암 세포주 3종에 대한 세포독성 실험 결과(72시간)를 나타낸 도면이다.8 is a view showing the cytotoxicity test results (48 hours) for three types of colorectal cancer cell lines of bubon (cheonggyeongchae, cheong-gyeong) and new cultivar Chinese cabbage (DH-005), FIG. And a diagram showing the cytotoxicity test results (72 hours) for three types of colorectal cancer cell lines of new Chinese cabbage (DH-005), FIG. A diagram showing the cytotoxicity test results for three types (48 hours), and FIG. 11 is a cytotoxicity test result for three gastric cancer cell lines of bubon (cheonggyeongchae, cheong-gyeong) and new varieties of Chinese cabbage (DH-005) (72 hours). is a diagram showing
상기 표 4 및 표 5과 도 8 내지 도 11을 참조하면, 신품종 배추(DH-005)가 모두 부본(청경채)보다 높은 활성을 보이고 있으며, 이로부터 낮은 농도에서도 암세포주의 성장을 억제하는 것을 확인할 수 있으며, 이로부터 신품종 배추의 항암활성을 확인할 수 있다.Referring to Tables 4 and 5 and FIGS. 8 to 11, it can be seen that the new variety of Chinese cabbage (DH-005) all showed higher activity than that of bok choy (bok choy), and from this, it was confirmed that it inhibited the growth of cancer cell lines even at low concentrations. And from this, it can be confirmed the anticancer activity of the new variety of Chinese cabbage.
이를 통해, 본 발명은 글루코시놀레이트 함량이 증가되고, 항암 활성이 우수한 신품종 배추, 신품종 배추의 종자, 신품종 배추의 부분체 및 신품종 배추의 자손 식물체를 제공할 수 있고, 신품종 배추 종자를 생산하는 방법을 제공할 수 있다.Through this, the present invention can provide a new type of Chinese cabbage, a seed of a new type of Chinese cabbage, a part of a new type of Chinese cabbage, and a progeny of a new type of Chinese cabbage, with an increased glucosinolate content and excellent anticancer activity, and to produce a new type of cabbage seed method can be provided.
이상으로 본 발명의 바람직한 실시예를 상세하게 설명하였다. 본 발명의 설명은 예시를 위한 것이며, 본 발명이 속하는 기술분야에서 통상의 지식을 가진 자는 본 발명의 기술적 사상이나 필수적인 특징을 변경하지 않고서 다른 구체적인 형태 쉽게 변형이 가능하다는 것을 이해할 수 있을 것이다.The preferred embodiment of the present invention has been described in detail above. The description of the present invention is for illustration, and those of ordinary skill in the art to which the present invention pertains will understand that other specific forms can be easily modified without changing the technical spirit or essential features of the present invention.
따라서, 본 발명의 범위는 상세한 설명보다는 후술하는 특허청구범위에 의하여 나타내어지며, 특허청구범위의 의미, 범위 및 그 균등 개념으로부터 도출되는 모든 변경 또는 변형된 형태가 본 발명의 범위에 포함되는 것으로 해석되어야 한다.Therefore, the scope of the present invention is indicated by the claims to be described later rather than the detailed description, and all changes or modifications derived from the meaning, scope, and equivalent concept of the claims are interpreted as being included in the scope of the present invention. should be
<110> REPUBLIC OF KOREA(MANAGEMENT : RURAL DEVELOPMENT ADMINISTRATION) <120> New variety of Chinese cabbage and its seeds <130> DP20210186 <150> KR 10-2020-0130241 <151> 2020-10-08 <160> 4 <170> KoPatentIn 3.0 <210> 1 <211> 1083 <212> DNA <213> Artificial Sequence <220> <223> Chiffu-MYB29.1 <400> 1 atggagaaga cgtcgaagaa gctatttatt ttcaatctct gcatcaattt tggtatttta 60 gtaacaagaa gatgcaacgc aactacatac tttgtgggag acacgtccgg ttgggacata 120 agctccgatc ttgaatcttg gcctttaggc aagcgattct ctgtcggtga tgttctaatg 180 ttccaatact catcgtcgca tagtgtctac gaagtggcga aagacaactt ccaaagctgc 240 aacactacgg accccatccg tacgttcaca aatgggaaca cgaccgttgc tctgtccaaa 300 ccgggagacc ggttctttgt atgtggtaac cggctccatt gctttgctgg tatgaggcta 360 caagtcaatg tccaaggcaa tggcccatct ccggcccctg tgggcgctcc acgggccgcc 420 cccgcaggaa ttctccagcc atcttctaaa aagaataacc ctgcaaccgg ggttgctagc 480 tcggctgccc atattggtgg ccgcggtctg aggggcaaaa acttagccat gaatctttct 540 gctatcactc agggaaacga caatgacgcc ccggcgccgg gtaacgaatt tgcacagttc 600 gccgctggat gcttctgggg cgtggagctg gcgtttcaga gagtctccgg tgtgactcat 660 acggaggttg gatacaccca agggctcctc cacaatcctt catacgagga tgtctgcacg 720 aacacaacga accatgcaga ggttgtcagg gttcaatatg atcccaaaga gtgcaatttt 780 gagtctctgc ttgatgtctt ctggtctaga catgacccca ccaccttgaa tcgccaggga 840 aaagatgtgg gaacccaata cagatcaggg atatacttct acacacctga gcaggagaaa 900 caagcggcag agtcaatgga acgtcaccag caacaaatgg agagtaagat catgactgag 960 attctaccag ctaagaaatt ctacagagct gaggagtatc atcagcagta cctgtcaaag 1020 ggtggacagt cctgtggcat agcctgtaat agtccactca tgtgcagcgc tgctacggct 1080 taa 1083 <210> 2 <211> 1083 <212> DNA <213> Artificial Sequence <220> <223> LP21-MYB29.1 <400> 2 atggagaaga cgtcgaagaa gctatttatt ttcaatctct gcatcatttt tggtatttta 60 gtaacaagaa gatgcaacgc aactacatac tttgtgggag acacgtccgg ttgggacata 120 agctccgatc ttgaatcttg gcctttaggc aagcgattct ctgtcggtga tgttctaatg 180 ttccaatact catcgacgca tagtgtctac gaagtggcga aagacaactt ccaaagctgc 240 aacactacgg accccatccg tacgttcaca aatgggaaca cgaccgttgc tctgtccaaa 300 ccgggagacc ggttctttgt atgtggtaac cggctccatt gctttgctgg tatgaggcta 360 caagtcaatg tccaaggcaa tggcccatct ccggcccctg tgggcgctcc acgggccgcc 420 cccgcaggaa ttctccagcc atcttctaaa aagaataacc ctgcaaccgg ggttgctagc 480 tcggctgccc atattggtgg ccgcggtctg aggggcaaaa acttagccat gaatctttct 540 gctatcactc agggaaacga caatgacgcc ccggcgccgg gtaatgaatt tgcacagttc 600 gccgctggat gcttctgggg cgtggagctg gcgtttcaga gagtctccgg tgttactcat 660 acggaggttg gatacaccca agggttcctc cacaatcctt catacgagga tgtctgcacg 720 aacacaacga accatgcaga ggttgtcagg gttcagtatg atcctaaaga gtgcaatttt 780 gagtctctgc ttgatgtctt ctggtctaga catgacccca ccaccttgaa tcgccaggga 840 aaagatgtgg gaacccaata cagatcaggg atatacttct acacacctga gcaggagaaa 900 caagcggcag agtcaatgga acgtcaccag caacaaatgg agagtaagat catgactgag 960 attctaccag ctaagaaatt ctacagagct gaggagtatc atcagcagta cctgtcaaag 1020 ggtggacagt cctgtggcat agcctgtaat agtccactca tgtgcagcgc tgctacggct 1080 taa 1083 <210> 3 <211> 1031 <212> DNA <213> Artificial Sequence <220> <223> LP08-MYB29.1 <400> 3 atggggaaga cgtcgacgat tctatttcta ttctatctct gtatcatctt tgggatttca 60 gtgataacaa gatgcaatgc aacgacatac tttgtgggag atacctctgg ttgggacata 120 agctccgatc ttgaatcctg gactttaggc aagagattct ctgttggtga tgttctaatg 180 ttccaatact catcgacgca tagtgtctac gaagtggcaa aagacaactt ccaaagctgc 240 aactctacag acccgatccg tactttcaca aatgggaaca cgaccgtagc tctgtccaaa 300 ccgggagaca ggttctttct ctgcggtaat cgtctccatt gcttcgctgg tatgaggcta 360 caagtcaatg tcgaaggcaa tggcccatct ccgtcccctg tgggagctcc cggagctgct 420 cccgtaggaa ttcttcaacc gtcttctaag aagaataacc ctccgaccgg cgttgctact 480 agctcggctc cccatgttgg tggctgcggt ggaagggtta aaaaattggt tacggatccg 540 tctccgatcg ctcaaggacc cgacgacgac tctccggcgc cgggaaacca ctttgcccaa 600 ttcgccgccg gatgcttctg gggcgtggag cttgcgtttc agaggctccc gggggtgact 660 cagacagagg ttggatacac acaagggatc acccacaatc cttcgtacga ggacgtctgt 720 tccgaaacca cgggacacgc cgaggtcgtc agggttcagt acgatcccaa agactgcacc 780 tttgagtctc tccttgactt gttctggtcc aggcatgatc ccaccacctt gaatcgccag 840 gggaatgatg tgggaactcg ataccgatct gggatctact tctacacacc agagcaggag 900 aaactagcgc gggagtcact agaacgtcac cagcaaaaaa tggagaggaa gatcatgact 960 gagatcttgc ctgctaagaa attctacaga gccgaggagc agcagcagca gtatctgtcc 1020 aagggtggaa g 1031 <210> 4 <211> 1031 <212> DNA <213> Artificial Sequence <220> <223> DH005-MYB29.1 <400> 4 atggggaaga cgtcgacgat tctatttcta ttctatctct gtatcatctt tgggatttca 60 gtgataacaa gatgcaatgc aacgacatac tttgtgggag atacctctgg ttgggacata 120 agctccgatc ttgaatcctg gactttaggc aagagattct ctgttggtga tgttctaatg 180 ttccaatact catcgacgca tagtgtctac gaagtggcaa aagacaactt ccaaagctgc 240 aactctacag acccgatccg tactttcaca aatgggaaca cgaccgtagc tctgtccaaa 300 ccgggagaca ggttctttct ctgcggtaat cgtctccatt gcttcgctgg tatgaggcta 360 caagtcaatg tcgaaggcaa tggcccatct ccgtcccctg tgggagctcc cggagctgct 420 cccgtaggaa ttcttcaacc gtcttctaag aagaataacc ctccgaccgg cgttgctact 480 agctcggctc cccatgttgg tggctgcggt ggaagggtta aaaaattggt tacggatccg 540 tctccgatcg ctcaaggacc cgacgacgac tctccggcgc cgggaaacca ctttgcccaa 600 ttcgccgccg gatgcttctg gggcgtggag cttgcgtttc agaggctccc gggggtgact 660 cagacagagg ttggatacac acaagggatc acccacaatc cttcgtacga ggacgtctgt 720 tccgaaacca cgggacacgc cgaggtcgtc agggttcagt acgatcccaa agactgcacc 780 tttgagtctc tccttgactt gttctggtcc aggcatgatc ccaccacctt gaatcgccag 840 gggaatgatg tgggaactcg ataccgatct gggatctact tctacacacc agagcaggag 900 aaactagcgc gggagtcact agaacgtcac cagcaaaaaa tggagaggaa gatcatgact 960 gagatcttgc ctgctaagaa attctacaga gccgaggagc agcagcagca gtatctgtcc 1020 aagggtggaa g 1031 <110> REPUBLIC OF KOREA (MANAGEMENT : RURAL DEVELOPMENT ADMINISTRATION) <120> New variety of Chinese cabbage and its seeds <130> DP20210186 <150> KR 10-2020-0130241 <151> 2020-10-08 <160> 4 <170> KoPatentIn 3.0 <210> 1 <211> 1083 <212> DNA <213> Artificial Sequence <220> <223> Chiffu-MYB29.1 <400> 1 atggagaaga cgtcgaagaa gctatttatt ttcaatctct gcatcaattt tggtatttta 60 gtaacaagaa gatgcaacgc aactacatac tttgtgggag acacgtccgg ttgggacata 120 agctccgatc ttgaatcttg gcctttaggc aagcgattct ctgtcggtga tgttctaatg 180 ttccaatact catcgtcgca tagtgtctac gaagtggcga aagacaactt ccaaagctgc 240 aacactacgg accccatccg tacgttcaca aatgggaaca cgaccgttgc tctgtccaaa 300 ccgggagacc ggttctttgt atgtggtaac cggctccatt gctttgctgg tatgaggcta 360 caagtcaatg tccaaggcaa tggcccatct ccggcccctg tgggcgctcc aggggccgcc 420 cccgcaggaa ttctccagcc atcttctaaa aagaataacc ctgcaaccgg ggttgctagc 480 tcggctgccc atattggtgg ccgcggtctg aggggcaaaa acttagccat gaatctttct 540 gctatcactc agggaaacga caatgacgcc ccggcgccgg gtaacgaatt tgcacagttc 600 gccgctggat gcttctgggg cgtggagctg gcgtttcaga gagtctccgg tgtgactcat 660 acggaggttg gatacaccca agggctcctc cacaatcctt catacgagga tgtctgcacg 720 aacacaacga accatgcaga ggttgtcagg gttcaatatg atcccaaaga gtgcaatttt 780 gagtctctgc ttgatgtctt ctggtctaga catgacccca ccaccttgaa tcgccaggga 840 aaagatgtgg gaacccaata cagatcaggg atatacttct acacacctga gcaggagaaa 900 caagcggcag agtcaatgga acgtcaccag caacaaatgg agagtaagat catgactgag 960 attctaccag ctaagaaatt ctacagagct gaggagtatc atcagcagta cctgtcaaag 1020 ggtggacagt cctgtggcat agcctgtaat agtccactca tgtgcagcgc tgctacggct 1080 taa 1083 <210> 2 <211> 1083 <212> DNA <213> Artificial Sequence <220> <223> LP21-MYB29.1 <400> 2 atggagaaga cgtcgaagaa gctatttatt ttcaatctct gcatcatttt tggtatttta 60 gtaacaagaa gatgcaacgc aactacatac tttgtgggag acacgtccgg ttgggacata 120 agctccgatc ttgaatcttg gcctttaggc aagcgattct ctgtcggtga tgttctaatg 180 ttccaatact catcgacgca tagtgtctac gaagtggcga aagacaactt ccaaagctgc 240 aacactacgg accccatccg tacgttcaca aatgggaaca cgaccgttgc tctgtccaaa 300 ccgggagacc ggttctttgt atgtggtaac cggctccatt gctttgctgg tatgaggcta 360 caagtcaatg tccaaggcaa tggcccatct ccggcccctg tgggcgctcc aggggccgcc 420 cccgcaggaa ttctccagcc atcttctaaa aagaataacc ctgcaaccgg ggttgctagc 480 tcggctgccc atattggtgg ccgcggtctg aggggcaaaa acttagccat gaatctttct 540 gctatcactc agggaaacga caatgacgcc ccggcgccgg gtaatgaatt tgcacagttc 600 gccgctggat gcttctgggg cgtggagctg gcgtttcaga gagtctccgg tgttactcat 660 acggaggttg gatacaccca agggttcctc cacaatcctt catacgagga tgtctgcacg 720 aacacaacga accatgcaga ggttgtcagg gttcagtatg atcctaaaga gtgcaatttt 780 gagtctctgc ttgatgtctt ctggtctaga catgacccca ccaccttgaa tcgccaggga 840 aaagatgtgg gaacccaata cagatcaggg atatacttct acacacctga gcaggagaaa 900 caagcggcag agtcaatgga acgtcaccag caacaaatgg agagtaagat catgactgag 960 attctaccag ctaagaaatt ctacagagct gaggagtatc atcagcagta cctgtcaaag 1020 ggtggacagt cctgtggcat agcctgtaat agtccactca tgtgcagcgc tgctacggct 1080 taa 1083 <210> 3 <211> 1031 <212> DNA <213> Artificial Sequence <220> <223> LP08-MYB29.1 <400> 3 atggggaaga cgtcgacgat tctatttcta ttctatctct gtatcatctt tgggatttca 60 gtgataacaa gatgcaatgc aacgacatac tttgtgggag atacctctgg ttgggacata 120 agctccgatc ttgaatcctg gactttaggc aagagattct ctgttggtga tgttctaatg 180 ttccaatact catcgacgca tagtgtctac gaagtggcaa aagacaactt ccaaagctgc 240 aactctacag acccgatccg tactttcaca aatgggaaca cgaccgtagc tctgtccaaa 300 ccgggagaca ggttctttct ctgcggtaat cgtctccatt gcttcgctgg tatgaggcta 360 caagtcaatg tcgaaggcaa tggcccatct ccgtcccctg tgggagctcc cggagctgct 420 cccgtaggaa ttcttcaacc gtcttctaag aagaataacc ctccgaccgg cgttgctact 480 agctcggctc cccatgttgg tggctgcggt ggaagggtta aaaaattggt tacggatccg 540 tctccgatcg ctcaaggacc cgacgacgac tctccggcgc cgggaaacca ctttgcccaa 600 ttcgccgccg gatgcttctg gggcgtggag cttgcgtttc agaggctccc gggggtgact 660 cagacagagg ttggatacac acaagggatc acccacaatc cttcgtacga ggacgtctgt 720 tccgaaacca cgggacacgc cgaggtcgtc agggttcagt acgatcccaa agactgcacc 780 tttgagtctc tccttgactt gttctggtcc aggcatgatc ccaccacctt gaatcgccag 840 gggaatgatg tgggaactcg ataccgatct gggatctact tctacacacc agagcaggag 900 aaactagcgc gggagtcact agaacgtcac cagcaaaaaa tggagaggaa gatcatgact 960 gagatcttgc ctgctaagaa attctacaga gccgaggagc agcagcagca gtatctgtcc 1020 aagggtggaa g 1031 <210> 4 <211> 1031 <212> DNA <213> Artificial Sequence <220> <223> DH005-MYB29.1 <400> 4 atggggaaga cgtcgacgat tctatttcta ttctatctct gtatcatctt tgggatttca 60 gtgataacaa gatgcaatgc aacgacatac tttgtgggag atacctctgg ttgggacata 120 agctccgatc ttgaatcctg gactttaggc aagagattct ctgttggtga tgttctaatg 180 ttccaatact catcgacgca tagtgtctac gaagtggcaa aagacaactt ccaaagctgc 240 aactctacag acccgatccg tactttcaca aatgggaaca cgaccgtagc tctgtccaaa 300 ccgggagaca ggttctttct ctgcggtaat cgtctccatt gcttcgctgg tatgaggcta 360 caagtcaatg tcgaaggcaa tggcccatct ccgtcccctg tgggagctcc cggagctgct 420 cccgtaggaa ttcttcaacc gtcttctaag aagaataacc ctccgaccgg cgttgctact 480 agctcggctc cccatgttgg tggctgcggt ggaagggtta aaaaattggt tacggatccg 540 tctccgatcg ctcaaggacc cgacgacgac tctccggcgc cgggaaacca ctttgcccaa 600 ttcgccgccg gatgcttctg gggcgtggag cttgcgtttc agaggctccc gggggtgact 660 cagacagagg ttggatacac acaagggatc acccacaatc cttcgtacga ggacgtctgt 720 tccgaaacca cgggacacgc cgaggtcgtc agggttcagt acgatcccaa agactgcacc 780 tttgagtctc tccttgactt gttctggtcc aggcatgatc ccaccacctt gaatcgccag 840 gggaatgatg tgggaactcg ataccgatct gggatctact tctacacacc agagcaggag 900 aaactagcgc gggagtcact agaacgtcac cagcaaaaaa tggagaggaa gatcatgact 960 gagatcttgc ctgctaagaa attctacaga gccgaggagc agcagcagca gtatctgtcc 1020 aagggtggaa g 1031
Claims (13)
A new variety of Chinese cabbage seeds deposited under the accession number KACC 98082P, obtained by hybridizing the parent oilseed cabbage and the parent, Bakchoy cabbage.
상기 신품종 배추는 글루코시놀레이트 함량이 상기 모본 및 부본보다 증가된 것을 특징으로 하는 신품종 배추 종자.
The method of claim 1,
The new variety of Chinese cabbage seeds is a new variety of Chinese cabbage seeds, characterized in that the glucosinolate content is increased than that of the parent and secondary versions.
상기 모본인 유지종 배추는 노란 종피를 갖는 유전자원 YS-033(CGN06835) 계통이고, 상기 부본인 박쵸이 배추는 진갈색 종피를 갖는 유전자원 PC-099(CGN13924) 계통인 것을 특징으로 하는 신품종 배추 종자.
The method of claim 1,
The parent oilseed cabbage is a genetic resource YS-033 (CGN06835) having a yellow seed coat, and the parent Bakchoy cabbage is a genetic resource PC-099 (CGN13924) line having a dark brown seed coat. .
상기 신품종 배추 종자는 서열번호 4로 표시되는 것을 특징으로 하는 신품종 배추 종자.
The method of claim 1,
The new variety of Chinese cabbage seeds is a new variety of Chinese cabbage seeds, characterized in that represented by SEQ ID NO: 4.
상기 신품종 배추 종자는 3-부테닐 이소티오시아네이트(3-Butenyl isothiocyanate), 4-펜테닐 이소티오시아네이트(4-pentenyl isothiocyanate), 2-페네틸 이소티오시아네이트(2-Phenethyl isothiocyanate) 및 설포라판(Sulforaphane)으로 이루어진 군에서 선택된 2종 이상의 함량이 부본인 박쵸이 배추보다 증가된 것을 특징으로 하는 신품종 배추 종자.
The method of claim 1,
The new varieties of Chinese cabbage seeds are 3-Butenyl isothiocyanate, 4-pentenyl isothiocyanate, 2-Phenethyl isothiocyanate, and A new type of Chinese cabbage seed, characterized in that the content of two or more selected from the group consisting of Sulforaphane is increased than that of Bakchoy cabbage.
상기 신품종 배추 종자는 항암 활성을 나타내는 것을 특징으로 하는 신품종 배추 종자.
The method of claim 1,
The new type of Chinese cabbage seed is a new type of Chinese cabbage seed, characterized in that it exhibits anticancer activity.
상기 항암 활성은 대장암 또는 위암에 대한 항암 활성인 것을 특징으로 하는 신품종 배추 종자.
7. The method of claim 6,
The anticancer activity is a new variety of Chinese cabbage seeds, characterized in that the anticancer activity against colon cancer or stomach cancer.
A new variety of Chinese cabbage obtained from the seed of any one of claims 1 to 7.
A part of the new variety of Chinese cabbage according to claim 8.
10. The method according to claim 9, wherein the part is a protoplast, an ovule, a cell, a pollen grain, an embryo, a cotyledon, a hypocotyledon, a root, a meristem ( meristem), anther, a part of a new type of Chinese cabbage, characterized in that at least one selected from the group consisting of stems and leaves.
The descendants of the new variety of Chinese cabbage of claim 8.
b) 박쵸이 배추를 소포자배양하여 부본을 수득하는 단계;
c) 상기 모본 및 부본을 교잡하여 잡종을 수득하는 단계; 및
d) 상기 잡종을 소포자배양하는 단계;
를 포함하는, 수탁번호 KACC 98082P로 기탁된 신품종 배추 종자의 생산방법.
a) self-pollination of oilseed cabbage to obtain a parent;
b) culturing Bakchoy cabbage with small spores to obtain a copy;
c) hybridizing the parent and parent to obtain a hybrid; and
d) culturing the hybrid with vesicles;
A method for producing new varieties of cabbage seeds deposited with accession number KACC 98082P, including a.
상기 a) 단계의 자가수분은 3회 이상인 것을 특징으로 하는 신품종 배추 종자의 생산방법.13. The method of claim 12,
The self-pollination of step a) is a method of producing a new variety of Chinese cabbage seeds, characterized in that 3 or more times.
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