In vitro and in vivo inhibitory effect of gaseous chlorine dioxide against pathogenic fungi isolated from stored sweetpotato (Ipomoea batatas L.)

이예지 Graduate School, Korea University 2018 국내석사

Sweetpotato (Ipomoea batatas Lam.) is one of the important food crops in Korea; however, postharvest fungi still cause a significant loss of quality during storage. In this study pathogenic fungi from sweetpotatoes were isolated and identified; Four fungal isolates, SP-d1, SP-f6, SP-p15, and SP-r1, were obtained from diseased sweetpotatoes from Muan, Korea. For identification of these isolates, internal transcribed spacer (ITS) region sequences were amplified using the primers ITS1 and ITS4. As a result, isolates SP-d1, SP-f6, SP-p15, and SP-r1 were identified as Diaporthe batatas (100% similarity), Fusarium oxysporum (99.10% similarity), Penicillium expansum (100% similarity), and Rhizopus oryzae (99.15% similarity), respectively. To test pathogenicity of the isolates, healthy sweetpotatoes were hole-inoculated with mycelial agar plugs. Isolates SP-d1, SP-f6, SP-p15, and SP-r1 caused symptoms on sweetpotato roots at 14, 10, 14 days, and 36 hours after inoculation, respectively. These isolates were re-isolated from the lesions to prove Koch’s postulates. In vitro and in vivo inhibitory effects of gaseous chlorine dioxide (ClO2) against isolates SP-d1 and SP-f6 were also evaluated. For in vitro tests, each fungus smeared on acidified potato dextrose agar (APDA) was treated with various ClO2 concentrations (1, 5, 10, and 20 ppm) for 0, 1, 10, 30, and 60 min. Fungal populations were decreased at the concentrations of ClO2 over the times. For isolate SP-d1, the decline of most spore development started at 30 min of 1 ppm ClO2; for isolate SP-f6, almost all spore development was inhibited at 5 ppm treatment of ClO2 for 5 min. The test fungal isolates were inhibited at the 20 ppm treatment of ClO2. For in vivo tests, spore suspension of each isolate was dropped on sweetpotato slices and treated with various ClO2 concentrations (5, 10, and 20 ppm) for 0, 10, 30, and 60 min. Lesion diameters of these isolates were not significantly different at the concentrations of ClO2 over the times. However, fungal populations were decreased at the ClO2 concentrations over times. These results indicate that D. batatas, F. oxysporum, P. expansum, and R. oryzae may be significant fungal pathogens on sweetpotatoes during storage in Korea and gaseous ClO2 can inhibit the fungal development and infection on sweetpotato. This also implies that ClO2 may be applied for controlling fungal diseases during sweetpotato storage. 고구마는 한국에서 주요 식용 작물 중 하나이지만, 저장 기간 중 진균에 의해 발생하는 저장병은 고구마에 심각한 손실을 일으킨다. 본 연구에서는 고구마에서 분리한 병원균을 동정하고, 이산화염소 가스 처리가 분리된 병원균에 미치는 억제효과를 분석하였다. 무안에서 받은 병든 고구마 샘플에서 총 4개의 균주(SP-d1, SP-f6, SP-p15, SP-r1)를 분리하였다. 분리된 균주는 ITS1과 ITS4 primer를 이용하여 internal transcribed spacer (ITS) region을 증폭시켜 동정하였다. 동정 결과, 균주 SP-d1, SP-f6, SP-p15, SP-r1은 각각 Diaporthe batatas (100% similarity), Fusarium oxysporum (99.10% similarity), Penicillium expansum (100% similarity), Rhizopus oryzae (99.15% similarity)로 동정되었다. 분리된 균주의 병원성 테스트를 위하여 건강한 고구마 (‘주황미’)에 각각의 병원균의 균사를 상처접종한 결과, 균주 SP-d1, SP-f6, SP-p15, SP-r1은 각각 접종 10일, 14일, 10일, 36시간 후 고구마에 병징이 나타났다. 이 균주들은 병징으로부터 재분리됨으로써 코흐의 법칙을 충족시켰다. 다음으로, 균주 SP-d1과 SP-f6에 대한 이산화염소 가스 처리의 억제효과를 분석하였다. 먼저, in vitro 실험에서는 균주의 포자 현탁액을 각각 acidified potato dextrose agar (APDA)에 도말 후, 다양한 이산화염소 농도(1, 5, 10, 20 ppm)를 0, 1, 10, 30, 60분 동안 처리하였다. 그 결과, 가스처리의 농도와 시간이 커질수록 병원균의 밀도가 감소되었다. 균주 SP-d1에 대한 억제효과가 제일 컸고, 이 균은 1 ppm의 이산화염소 가스를 30분동안 처리하였을 때에도 포자 형성이 거의 대부분 억제되었다. 균주 SP-f6은 5 ppm의 이산화염소 가스를 5분 처리하였을 때 포자의 발달이 크게 저하되었다. 20 ppm의 이산화염소 가스를 처리하였을 때에는 두 균주 모두 생장이 억제되었다. In vivo 실험에서는 1 cm 간격으로 얇게 썬 고구마 슬라이스에 각 균주의 포자 현탁액을 10 µl씩 접종한 후, 5, 10, 20 ppm 농도의 이산화염소 가스를 0, 10, 30, 60분 처리하였다. 그 결과, 병징 길이를 평가한 값은 가스처리를 한 고구마와 하지 않은 고구마 간에 통계적으로 차이가 없었다. 하지만, 곰팡이의 밀도는 가스처리의 농도와 시간이 증가할수록 감소하였다. 이 연구의 결과를 종합해보면 D. batatas, F. oxysporum, P. expansum, R. oryzae는 수확 후 저장고구마에 중요한 병원균이라는 것을 보여주고, 이산화염소 가스 처리가 이 병원균에 의한 감염을 억제할 수 있다는 것을 알 수 있었다. 따라서, 이산화염소 가스 처리는 고구마의 저장기간 동안 저장병을 방제할 수 있는 하나의 방법이 될 수 있을 것이다.

Morphological, biochemical, and molecular characterization of sweetpotato (Ipomoea batatas (L.) Lam.) germplasms collected in Korea

Nie, Hualin 서울시립대학교 일반대학원 2020 국내박사

고구마(Ipomoea batatas (L.) Lam.)는 메꽃과에 속하는 육배체의 괴근 작물이다. 이는 쌀, 밀, 옥수수, 감자, 보리, 카사바 다음으로 세계에서 일곱 번째로 가장 중요한 식량작물이며, 개발도상국에서는 다섯 번째로 중요한 작물이다. 한국 고구마의 유전자 및 표현형 정보가 부족하여 품종 관리자, 육종가, 유전학자, 원예학자 및 식물 병리학자들이 어려움을 안고 있다. 본 연구에서는 한국에서 채취한 66 개의 고구마 생식질을 이용하여 분자적 표지, 형태학적 연구, 생화학적 분석을 통해 유전다양성과 대사다양성에 대해 평가되었다. 형태학적 특성을 연구한 결과로 저장뿌리의 착색이 매우 중요한 역할을 하고 있다. 66 개의 고구마 생식질 중에서 총 39,424 개의 단일염기다형성(SNP)이 genotyping by sequencing (GBS) 방법을 사용하여 선발되었다. Neighbor-Joining 클러스터 분석 및 집단구성에 의하면 39,424 개의 SNP에서 생식질을 다섯 개의 그룹으로 분리했다. 유전적 다양성과 집단구성에서 육종계통, 지리적 영역 및 착색 유형과 밀접한 관련이 있었음을 보여주었다. 기존 조건에 생화학적 수준에서 유전자형 및 저장근 색깔 간의 유의한 상관관계를 찾기 위해 생화학적 분석, 즉 탄수화물 및 착색 조사를 수행하였다. 당 함량 최대치는 황색 고구마 및 유전자 그룹 3에 속한 개체들에서 관찰되었으며, 카로티노이드 최대치는 주황색 고구마 및 유전자 그룹 5에 속한 개체들에서 관찰되었다. 더 깊은 연구를 위해 전분 대사에 관여하는 중요한 유전자 및 단백질이 조사되었다. ADP-glucose pyrophosphorylase (AGPase) 유전자는 전분 대사에 매우 중요한 유전자로 밝혀졌으며 유전자와 단백질의 여러 기작에 작용했을 것으로 예상된다. 이를 통해 한국 고구마의 유전자 다양성, 대사 다양성 및 전분 대사 관련 유전자에 대한 포괄적인 연구를 제시했다. 본 연구에서 개발한 대사 산물 측정 방법과 전분 대사 관련 유전자 분석 방법은 고구마 육종에 중요한 방향을 제시하고 대사와 유전자 다양성 간의 상호관계를 분석하는 새로운 방법을 제공하였다. 본 연구는 고구마 품종 특성의 평가 및 개선에 큰 잠재력을 가지고 있으며 고구마의 다양성 평가에 전반적인 연구 이행을 향한 첫 단계다. Sweetpotato (Ipomoea batatas (L.) Lam.) is a hexaploid (2x = 6n = 90) tuber crop belonging to the family convolvulaceae. The species is the world’s seventh most important food crop only after rice, wheat, maize, potato, barley, and cassava. Korean sweetpotato germplasm resources generally lack genetic information, which poses challenges for germplasm managers, breeders, geneticists, horticulturalists, and plant pathologists. In this study, 66 sweetpotato germplasms collected in Korea were evaluated for genetic diversity and metabolic diversity using molecular markers, morphological investigation, and biochemical analysis. Morphological traits were studied and were highly significant for storage root flesh color. Among the 66 sweetpotato germplasms, 39,424 single nucleotide polymorphisms (SNPs) were selected using genotyping by sequencing (GBS). Neighbor-Joining cluster analysis and population structure based on the 39,424 SNPs separated the germplasm into five groups. Genetic diversity and population structure were highly associated with breeding lineage, geographic region, and color-fleshed type. Biochemical analysis, viz. carbohydrate and pigmentation, was performed to find out the significant correlation with the genotype and the type of storage root flesh color at a biochemical level under given conditions. Maximum total sugars were observed in the yellow-fleshed germplasms and the genetic group three germplasms, whereas maximum total carotenoid was observed in the orange-fleshed germplasms and the genetic group five germplasms. Important genes and proteins involved in starch metabolism were also studied. ADP-glucose pyrophosphorylase (AGPase) genes were proved to be very important genes for starch metabolism and probably functioned in several mechanisms of genes and proteins. This study presents a comprehensive research on the genetic diversity, metabolic diversity, and the starch metabolism-related gene of Korean sweetpotato. Assessment of SNPs markers and metabolite, developed in this study, have great potential in marker-assisted breeding, which can be used to characterize Korean sweetpotato. Analysis of the starch metabolism-related genes provided a significant direction for sweetpotato breeding and a new method for analysis of the relationships between metabolic and genetic diversity. Furthermore, this study is the first step toward the implementation of quantitative research on the metabolic diversity of sweetpotato, which has great potential for the evaluation and improvement of sweetpotato cultivars.

Manipulation of Lipophilic Antioxidants to Enhance Abiotic Stress Tolerance and Nutritional Quality in Sweetpotato

김소은 과학기술연합대학원 대학교 2022 국내박사

The dramatic increase of global population combined with rapid industrialization has led to great strains on global food, nutrition and energy supplies. According to estimates of the Food and Agriculture Organization of the United Nations (UN-FAO), the world population will be more than 9.7 billion by 2050 and 11 billion by 2100. FAO anticipate that global food and energy requirements in 2050 will exceed current needs by more than 1.7-fold and 3.5-fold, respectively. In order to solve these global issues such as food and nutrition security in the face of climate change and aging society, it is necessary to develop food crops that are resistant to environmental stress with enhanced essential nutrients. Sweetpotato [Ipomoea batatas (L.) Lam] is the sixth most important starch crop in the world and has rich source of starch, low molecular weight (LMW) antioxidants, dietary fiber and potassium. High levels of LMW antioxidants are an important for both the plant protection to environmental stress and nutritional merits for human being as anti-aging and anti-disease agents. LMW antioxidants in plants can be divided into lipid-soluble antioxidants such as carotenoids and tocopherols (vitamin E) and water-soluble antioxidants such as ascorbic acid (vitamin C), glutathione and polyphenols. In previous study, sweetpotato IbOr protein isolated form an orange-fleshed sweetpotato, with strong holdase chaperone activity, protects a key enzyme, phytoene synthase (IbPSY), in the carotenoid biosynthetic pathway and stabilizes a photosynthetic component, oxygen-evolving enhancer protein 2-1 (IbPsbP), under heat and oxidative stresses in plants. In addition, previously, five tocopherol biosynthetic genes were isolated and investigated their expression levels in leaves under drought, salt, and oxidative stresses. In this study, gene manipulation of lipid-soluble antioxidants such as carotenoids and tocopherols was conducted to develop transgenic sweetpotato plants with enhanced abiotic stress tolerance and nutrition quality through plant biotechnology. The major results are as follows. 1) Site-directed mutagenesis of the IbOr gene, leading to Arg to His substitution at position 96 in the IbOr protein (IbOr-R96H), was performed and IbOr-R96H transgenic sweetpotato calli and plants using a white-fleshed sweetpotato cultivar, Xushu 29, were successfully generated. Transgenic sweetpotato calli overexpressing IbOr-R96H with a dark-orange color exhibited 13.3-fold higher total carotenoid contents and 39.3-fold higher levels of β-carotene than those in IbOr-WT calli with a light-yellow color. Interestingly, transgenic calli overexpressing IbOr-R96H showed increased tolerance to salt (150 mM NaCl) and heat (47°C) stresses. In addition, storage roots of transgenic sweetpotato plants overexpressing IbOr-WT and IbOr-R96H exhibited a light-yellow color and light-orange color, respectively. The total carotenoid contents and β-carotene content of IbOr-R96H storage roots were up to 6.1- and 99.3-fold higher, respectively, than those of IbOr-WT storage roots. Particularly, the β-carotene content of IbOr-R96H storage roots was up to 186.2-fold higher than that of NT storage roots. In addition, leaf discs excised from IbOr-R96H plants showed greater tolerance to heat stress (47°C) than NT and IbOr-WT plants. 2) To further analyze the regulatory mechanisms of IbOr, through yeast two hybrid (Y2H) screening, I found several potential candidates interacting with IbOr protein. I conducted to elucidate the mechanism for regulating environmental stress resistance in relation to the interaction between IbOr and chlorophyll a,b-binding protein (CAB) which is one of the proteins constituting the photosystem. Sweetpotato CAB protein (IbCAB) were isolated and confirmed interaction with IbOr-WT and IbOr-R96H protein using co-immunoprecipitation (Co-IP) and bimolecular fluorescence complementation (BiFC) assay. In order to resolve the regulatory mechanism through this interaction, IbCAB expression pattern analysis under various environmental stress conditions and light-related stress tolerance assay are under investigation. 3) Previously, isolation of 4-hydroxyphenylpyruvate dioxygenase (IbHPPD) and tocopherol cyclase (IbTC) genes in tocopherol biosynthesis pathway has been conducted. Transgenic sweetpotato plants overexpressing IbHPPD (referred as HP plants) and IbTC (referred as TC plants) were generated to understand the function of tocopherol biosynthetic genes in sweetpotato. Compared with non-transgenic (NT) plants, HP plants exhibited enhanced tolerance to multiple environmental stresses, including salt, drought, and oxidative stresses. In addition, HP plants showed increased tolerance to the herbicide sulcotrione, which is involved in the inhibition of the HPPD. Interestingly, under dehydrated condition, HP plants displayed an elevated α-tocopherol content to 19–27% in leaves compared with NT plants. Additionally, the content of α-tocopherol was 1.6–3.3-fold higher in TC leaves than in NT leaves. Compared with NT plants, TC plants showed enhanced tolerance to multiple environmental stresses, including salt, drought, and oxidative stresses. In conclusion, (1) my results demonstrate that overexpression IbOr-R96H in sweetpotato exhibited enhanced production of carotenoids and environmental stress tolerance and will facilitate the development of new cultivars with enhanced nutritional content and environmental stress tolerance. (2) I anticipate that site-specific mutagenesis of IbOr combined with CRISPR-Cas9-mediated base-editing techniques could be an effective tool for the improvement of carotenoid contents in various plant species. (3) It has been suggested that the interaction of IbOr and IbCAB protein plays an important role in photosynthesis. (4) In addition, IbHPPD and IbTC overexpressing transgenic sweetpotato plants showed an enhanced tolerance to various abiotic stresses. (5) Taken together, results of this study suggest that the manipulation of lipophilic antioxidants represents useful resource for developing crops with increased antioxidant levels and abiotic stress resistance. And it would help mitigate the global food, nutrition and energy security issues in the face of global climate change, thus facilitating the establishment of a sustainable society. 산업혁명 이후 급속한 산업화와 세계 인구 증가는 심각한 글로벌 기후변화뿐만 아니라 식량, 에너지, 보건문제를 초래하고 있다. UN 식량농업기구 (FAO)는 2050년 세계인구는 97억 명, 2100년에는 110억 명에 도달하게 될 것이며 현재 추세대로 식량과 에너지를 사용한다면 2050년에는 식량과 에너지가 각각 지금의 1.7배와 3.5배 이상 필요할 것이라 전망했다. 기후위기시대, 고령화 시대에 대응하기 위해서는 환경스트레스에 잘 견디고 영양성분이 강화된 식량작물 개발이 필요하다. 고구마[Ipomoea batatas (L.) Lam.]는 세계 6대 식량작물로 전분 뿐만아니라 비타민C, 베타카로틴 등 각종 항산화물질, 식이섬유, 칼륨 등이 풍부하여 건강식품으로 재평가되고 있다. 저분자 항산화물질은 환경스트레스로부터 식물체를 보호하고 인간의 노화방지, 각종 질병 예방 등에 중요한 역할을 한다. 저분자 항산화물질은 카로티노이드, 토코페롤 (비타민E)과 같은 지용성 항산화물질과 아스코르브산 (비타민C), 글루타티온, 폴리페놀과 같은 수용성 항산화물질로 크게 분류할 수 있다. 선행연구에서 분리한 고구마 Orange (IbOr) 단백질은 강한 샤페론활성을 가지면서 카로티노이드 축적뿐만 아니라 고온, 건조, 염 등 다양한 환경스트레스에 저항성을 유도하였다. IbOr 단백질은 고온 등 스트레스 조건에서 카로티노이드 생합성에 중요한 phytoene synthase (IbPSY) 단백질과 광합성 광계II 단백질의 하나인 oxygen-evolving enhancer protein 2–1 (IbPsbP) 단백질과 결합하여 식물을 보호하는 것이 확인되었다. 또한 고구마에서 토코페롤 생합성에 중요한 유전자들이 분리되어 환경스트레스 관점에서 유전자 발현 등이 조사된 바 있다. 본 연구에서는 고구마의 지용성 항산화물질인 카로티노이드와 토코페롤을 형질전환기술을 이용한 대사조절로 지용성 항산화물질을 다량 함유하면서 복합 환경스트레스에 잘 견디는 복합기능성 고구마 개발을 위한 기반기술을 확보하고자 연구를 수행하여 다음과 같은 주요 결과들을 도출하였다. 1) 고구마 IbOr 단백질 염기서열에서 96번째 아미노산을 아르기닌 (R)에서 히스티딘 (H)로 치환시킨 IbOr-R96H 변이체를 제작하여 저장뿌리 속이 흰색인 고구마 품종 (Xushu 29)에 과발현 한 형질전환 캘러스와 식물체를 개발하였고 변이체의 기능을 IbOr (IbOr-WT)와 비교하였다. IbOr-R96H 배양세포는 IbOr-WT 배양세포보다 전체 카로티노이드와 베타카로틴 함량이 각 약 13.3배와 약 39.3배 증가하였다. 또한 IbOr-R96H 배양세포는 고염 (150 mM NaCl), 고온 (47°C) 스트레스에서 내성을 나타내었다.IbOr-WT 과발현 고구마와 IbOr-R96H 과발현 고구마의 저장뿌리에서 IbOr-WT은 노란색, IbOr-R96H는 옅은 주황색의 표현형을 띄었다. IbOr-R96H 과발현 저장뿌리의 전체 카로티노이드와 베타카로틴 함량은 IbOr-WT 식물체에 비해 각각 최대 약 6.1배와 99.3배 증가하였다. 특히 IbOr-R96H 식물체 저장뿌리의 베타카로틴 함량은 NT식물체에 비해 최대 약 186.2배 증가되었다. 또한 잎 절편 (leaf disc)를 이용한 고온 (47°C) 스트레스 내성 실험에서 IbOr-R96H 과발현 형질전환 고구마 식물체가 IbOr-WT식물체보다 47°C 고온에 더 강한 저항성을 가지는 것을 확인하였다. 2) IbOr 단백질의 기능을 자세히 규명하기 위해 yeast two hybrid (Y2H) screening을 통해 IbOr 단백질과 상호작용하는 새로운 여러 후보 단백질들을 탐색하여, 그 중 하나인 chlorophyll a,b-binding (CAB) 단백질의 기능을 규명하고자 하였다. 고구마에서 IbCAB 유전자를 분리하여 co-immunoprecipitation (Co-IP), bimolecular fluorescence complementation (BiFC) assay 실험으로 IbCAB 단백질이 IbOr-WT 단백질, IbOr-R96H 단백질과 상호작용한다는 것을 확인하였다. 현재 IbOr 단백질과 IbCAB 단백질의 상호작용 조절 메커니즘을 규명하기 위해 빛 등 다양한 환경스트레스 조건에서 IbCAB의 발현 패턴분석 등을 분석 중이다. 3) 고구마에 존재하는 토코페롤 생합성에 관여하는 유전자 가운데 4-hydroxyphenylpyruvate dioxygenase (IbHPPD)와 tocopherol cyclase (IbTC) 유전자의 기능을 이해하고자 이들 유전자를 과발현 하는 형질전환 고구마 (Xushu 29 품종 이용)를 제작하여 분석하였다. IbHPPD 식물체는 NT 식물체에 비해 HPPD 저해 제초제인 sulcotrione (1 mM), 염 (200 mM NaCl), 건조, 산화스트레스 (5 μM MV) 내성을 보였으며 건조스트레스 후에 잎에서 α-tocopherol 함량이 NT식물체에 비해 최대 27%까지 증가하였다. IbTC 식물체는 스트레스가 없는 정상 조건일 때 잎에서 α-tocopherol 함량이 NT 식물체에 비해 최대 3.3배 증가했고, 고염 (200 mM NaCl), 건조, 산화 (3 μM MV) 스트레스에서 내성이 증가한 것을 확인 할 수 있었다. 본 연구에서 얻어진 결과를 토대로 종합적인 고찰과 전망은 다음과 같다. (1) 카로티노이드 축적 관련 IbOr-R96H 유전자가 IbOr-WT와 함께 카로티노이드 함량을 대폭 증가시키면서 고온 등 환경스트레스에 강한 품종을 개발할 수 있음이 강하게 제시되었다. (2) IbOr-R96H 연구결과는 고구마 유전자 편집기술(CRISPR-Cas9-mediated base-editing techniques)에 직접 활용될 수 있을 뿐만 아니라 다양한 식물에도 적용할 수 있을 것으로 기대된다. (3) IbOr 단백질과 상호작용하는 IbCAB 단백질을 처음으로 분리하여 IbOr이 광합성에 매우 중요하게 관여함이 시사되어 후속연구가 기대된다. (4) 토코페롤 생합성 유전자 (IbHPPD와 IbTC) 과발현 형질전환연구에서 토코페롤 대사조절로 복합 환경스트레스 내성식물체 개발에 활용될 수 있음이 시사되었다. 결론적으로 본 연구에서 얻어진 결과를 발전시킨다면 기후위기와 고령화 시대에 대응할 수 있는 환경스트레스에도 강하고 지용성 항산화물질을 고생산하는 복합기능 고구마 개발에 기여할 것으로 기대된다.

Morphological characterization of ornamental sweetpotato cultivars and molecular analysis of a new virus

Kim, Do Yeon 서울시립대학교 일반대학원 2024 국내박사

본 연구는 식용 고구마와 관상용 고구마의 형태적 특성을 비교하고 메꽃과를 감염시키는 새로운 바이러스의 분자생물학적 특성을 규명하기 위해 수행되었다. 1장에서는 고구마의 형태적 특성을 비교하였는데, 형태적 특성은 품종에 따라 상당한 차이를 보였다. 식용 고구마(신율미, 베니하루카)는 잎 색깔이나 모양에서 큰 차이를 보이지 않았다. 그러나 관상용 고구마(스위트캐롤라인퍼플, 마르가리타, 골드핑거, 블랙하트, 모닝퍼플, 모닝화이트)는 품종 간 잎 색깔과 모양이 매우 다양한 것으로 나타났다. 그러나 국내육성품종인 모닝퍼플과 모닝화이트는 잎자루가 약해 쉽게 부러지는 것으로 나타났다. 관상용 고구마 품종 중 3 개 품종(스위트캐롤라인퍼플, 모닝퍼플, 모닝화이트)은 식재 후 약 60일이 지난 후부터 지속적으로 개화가 이루어져 관상용 식물로서 뛰어난 가치를 지니고 있는 것으로 나타났다. 이러한 결과는 화분이나 관상용 식물로 재배할 때 괴근을 형성하지 않는 관상용 품종으로 육종 시 참고할 수 있으며, 잠재적으로 고구마 시장이 원예분야로 확장될 수 될 수 있을 것으로 판단된다. 2장에서는 메꽃에서 신종바이러스를 발견하였다. 메꽃과 식물에는 고구마, 메꽃, 나팔꽃이 포함되며 고구마는 메꽃과 같은 영양번식 작물이다. 메꽃은 자연적으로 바이러스에 감염되지는 않지만 바이러스의 숙주가 되어 다른 작물에 영향을 미치는 것으로 알려져 있다. 새로운 바이러스는 어린잎에서는 바이러스 증상이 뚜렷하지만 성장하면서 증상은 없지만 바이러스는 존재하는 것으로 나타났고, Bindweed mottle virus(BWMV) 로 명명하였다. 같은 과에 속하는 다른 바이러스와 계통발생수를 비교했을 때, 지놈의 뉴클레오타이드 서열은 Mulcrilevirus와 밀접한 관련이 있는 것으로 나타났다. 그러나 외피단백질(Capsid Protein, CP)와 복제관련단백질(Replication- associated Protein, ReP)의 아미노산 서열은 Capulavirus와 높은 상동성을 나타냈다. 그러나 ORF 게놈 구조는 Topilevirus 와 유사하였다. 종합해보면, BWMV는 제미니바이러스과에 속하지만 기존의 속에서의 분류는 명확하지 않다. 3장에서는 고구마를 감염시키는 것으로 알려진 Sweetpotato pakakuy virus(SPPV)를 분자생물학적 특성을 분석하였다. 이 바이러스는 원형의 DNA 지놈을 가지고 있으며 배드나바이러스에 속한다. ‘신율미’에서 차세대 염기서열 분석으로 SPPV 를 검출하였지만 서열을 완성하지 못하였다. 이는 이 바이러스가 독립적인 바이러스가 아니라 숙주 유전체의 일부일 수 있다는 것을 시사한다. 바이러스인지 숙주지놈의 일부인지 확인하기 위해 고구마의 DNA를 PCR로 분석하였다. 역 PCR에서 나온 480bp 길이의 서열을 분석한 결과, SPPV의 서열이 야생고구마인 Ipomoea trifid의 지놈 서열과 연결되어 있는 것으로 밝혀졌다. 또한 서열은 15개 염색체 중 13개 염색체에 흩어져 있었다. 이 결과는 ‘신율미’에서 확인된 바이러스가 독립된 바이러스가 아님을 시사한다. 영양번식 작물은 감염된 바이러스가 후대에 계속 전파될 수 있기 때문에 각별한 주의가 필요하다. 이 바이러스는 고구마에 새로운 유형의 지속성 바이러스로 추정되며 고구마 생산에 미치는 영향에 대해서는 추가 연구가 필요하다고 판단된다. 본 연구에서는 고구마의 품종별 형태학적 특성을 구명하고 관상용으로서의 고구마의 잠재적인 가치를 발견하였다. 또한, 메꽃과 식물을 감염시킬 수 있는 신종바이러스를 메꽃에서 확인하였으며, 고구마 파카쿠이 바이러스가 독립적인 바이러스가 아닌 숙주게놈의 일부임을 확인하였고, 추후 이 바이러스가 실제 고구마 재배에 미치는 잠재적 영향에 대한 추가연구의 필요성을 제시하였다. This study was conducted to morphological characteristics of edible and ornamental sweetpotato and to characterize the molecular biology of a new virus infecting the family Convolvulaceae. In Chapter Ⅰ, the study compared the morphological characteristics of sweetpotatoes, which exhibited considerable variation among the cultivars. Edible sweetpotato (‘Shinyulmi’ and ‘Beniharuka’) did not show significant differences in leaf color or shape. However, ornamental sweetpotato (‘Sweet Caroline Purple’, ‘Margarita’, ‘Goldfinger’, ‘Blackheart’, ‘Morningpurple’, ‘Morningwhite’) demonstrated considerable variation in leaf color and shape among cultivars. The Korean varieties, ‘Morningpurple’ and ‘Morningwhite’, have weak petioles that are susceptible to breakage. Three of the ornamental sweetpotato cultivars ('Sweet Caroline Purple', 'Morning Purple', and 'Morning White') began flowering about 60 days after planting and continued to flower consistently. This information may be used to develop ornamental sweetpotatoes that do not form tuberous root when grown in pots or as ornamental plants, potentially expanding the sweetpotato market into horticulture. In Chapter Ⅱ, a new virus was detected in bindweed (Calystegia sepium). The Convolvulacea family includes sweetpotato, bindweed, and morning glory. Sweetpotato is a bindweed-like vegetative propagation crop. Bindweed is not naturally infected with viruses but has been known to become a host for viruses, affecting other crops. The new virus has been named Bindweed mottle virus (BMWV), as the symptoms of the virus are evident in the young leaves, but the virus is present without symptom in the older leaves. When comparing to viruses of other genera in the family, Geminiviridae, the phylogenic tree, showed that it is closely related to the Mulcrilevirus in terms of the identity of nucleotide sequences of genome. However, both the sequence of amino acids of coat protein (CP) and replication protein (Rep) were closely related to the Capulavirus. Otherwise, the genome organization was similar to Topilevirus. Taken together, BMWV belongs to family Geminiviridae without question, but the classification in the level of genus is not clear. In Chapter Ⅲ, the study analyzed the molecular biological characteristics of previously reported sweetpotato virus, Sweetpotato pakakuy virus (SPPV), known to infect sweetpotato. It has circular DNA genome and belongs to the family Badnavirus. When SPPV was detected by next generation sequencing of ‘Shinyulmi’, the attempt to complete the genome of SPPV had been failed. This may be suggested that the virus was not independent virus, rather it was a part of host genome. To confirm whether it was a virus or a part of the host genome, the DNA of sweetpotato was subjected to inverse PCR. The nucleotide sequences of the 480bp long product from inverse PCR revealed that the sequence of SPPV was connected to the genome sequence of wild sweetpotato, Ipomoea trifida. Furthermore, the sequences were scattered to thirteen out of fifteen chromosomes. These results indicated that the virus identified from ‘Shinyulmi’ was not independent virus. The vegetatively-propagated crops require special care because viruses are transmitted by seeds. These viruses likely mean a new type of persistent virus in sweetpotato, and the threat they present to sweetpotato production needs to be further investigated. In conclusion, this study reveled the ornamental potential of sweetpotato cultivars through morphological analysis, identified a new virus, Bindweed mottle virus (BMWV), in bindweed that can infect Convolvulaceae plants, and repoted that Sweetpotato pakakuy virus (SPPV) is an integrated part of the host genome rather than an independent virus, suggesting the need for further research on its potential impact on sweetpotato cultivation.

Functional characterization of SRD1 and SRD2 genes in storage root development in sweetpotato

노설아 School of Life Sciences and Biotechnology, Korea U 2011 국내박사

To understand how storage root of sweetpotato develops, MADS-box gene (SRD1) and its promoter were identified and functionally characterized. Also, three expansin genes, IbEXP1, IbEXP2, and IbEXPL1 were identified and especially, IbEXP1 (SRD2) gene was functionally characterized by analysis of transgenic plants. The transcript level of SRD1 was only detected at roots and the highest level of SRD1 transcript was present at storage roots of early SR (storage root) stage. In situ hybridization reveals that SRD1 mRNA was mainly found in the actively dividing cells, including the vascular and cambium cells of the young storage root. The transcript level of SRD1 in the fibrous roots was increased by exogenous IAA treatment. And SRD1 transcript level was increased when endogenous IAA content increased during the storage root development at early stage. SRD1-overexpressing sweetpotato plants cultured in vitro produced thicker and shorter fibrous roots than wild-type plants. The metaxylem and cambium cells of the fibrous roots of SRD1- overexpressing plants showed markedly enhanced proliferation, resulting in the fibrous roots of these plants showing an earlier thickening growth than those of wild-type plants. Taken together, these results demonstrate that SRD1 plays a role in the formation of storage roots by activating the proliferation of cambium and metaxylem cells to induce the initial thickening growth of storage roots in an auxin-dependent manner. The 3.0 kb promoter of SRD1 from sweetpotato (Ipomoea batatas cv. ‘White Star’) was isolated and its activity in transgenic Arabidopsis, carrot, and potato using a reporter gene (uidA) was characterized. The SRD1 promoter conferred root-specific expression in transgenic Arabidopsis, with SRD1 promoter activity increasing in response to exogenous IAA. A serial 5’ deletion analysis of the SRD1 promoter identified regions related to IAA-inducible expression as well as regions containing positive and negative elements, respectively, controlling the expression level. In transgenic carrot, the SRD1 promoter mediated strong taproot-specific expression, as evidenced by GUS staining being strong in almost the entire taproot, including secondary phloem, secondary xylem and vascular cambium. The activity of the SRD1 promoter gradually increased with increasing diameter of the taproot in the transgenic carrot and was higher than that of the CaMV35S promoter. The SRD1 promoter also directed strong tuber-specific expression in transgenic potato. Taken together, these results demonstrate that the SRD1 promoter directs strong expression restricted to the underground storage organs, such as fleshy taproots and tubers, as well as fibrous root tissues. Additionally, three expansin genes, IbEXP1, IbEXP2 and IbEXPL1, were identified from EST library. These genes were investigated in basal function of expansin gene by chilling stress. The transcriptional regulation of three sweetpotato expansin genes (IbEXP1, IbEXP2 and IbEXPL1) in response to various chilling temperatures (12, 16, 22, and 28℃) was examined. IbEXP1 mRNA levels in the 12℃-treated petiole showed a fluctuation (transient decrease?Crecovery?Cstable decrease) pattern for 48 h. In the leaf and petiole, IbEXP1 and IbEXPL1 exhibited a similar response to chilling in that their mRNA levels decreased at 22℃, increased at 16℃, and decreased dramatically at 12℃. In contrast, mRNA levels of IbEXP2 in the leaf fell gradually as the temperature fell from 28 to 12℃, while they remained unaltered in the petiole. In the root, mRNA levels of IbEXPL1 and IbEXP1 reached maximum levels at 16℃, and decreased significantly at 12℃. Among them, SRD2 (IbEXP1) was functionally characterized in antisense transgenic sweetpotato plants. The transcript level of SRD2 was detected at actively growing tissues such as petioles and fibrous roots at fibrous root stage. SRD2-antisense sweetpotato plants grown in vitro have shorter roots than wild-type plants. Five months after planting, the number and weight of both storage roots and shoots were increased in SRD2-antisense transgenic plants. These results suggest that SRD2 plays a role as negative regulator in storage root development of sweetpotato. These studies could provide an understanding for mechanism of storage root development and these results could contribute to the agriculture for improving crops with advantages.

Molecular and Physiological Studies on Tuberous Roots of Sweetpotato under Low Temperature Storage

지창윤 과학기술연합대학원대학교 2018 국내박사

전 세계의 급격한 인구증가와 산업화는 심각한 식량과 에너지 불균형 문제를 일으키고 있다. UN 식량농업기구(FAO)는 2050년 세계인구가 약 91억명에 도달하며, 현재보다 약 1.7배의 식량과 3.5배 이상의 에너지가 필요할 것이라고 전망하고 있다. 이러한 글로벌 식량, 에너지 문제를 해결하기 위해서는 사막화 지역이나 저온지역 등 조건불리지역에도 잘 자랄 수 있는 작물 개발뿐만 아니라 에너지 절감을 위한 수확 후 저장관리기술이 필요하다. 최고의 건강식품 및 산업용 작물로 재평가 되고 있는 고구마 [Ipomoea batatas (L.) Lam]는 고온, 건조, 고염 등 조건불리지역에 재배가 용이하지만, 열대지역 기원 특성으로 인해 저온에 취약해 저온지역 재배가 어려울 뿐만 아니라 수확 후 저온저장에 따른 냉해로 인해 산업화의 장애가 되고 있다. 특히, 에너지 관점에서 겨울철 고구마의 저온저장성이 큰 문제가 되고 있다. 본 연구에서는 생명공학기술을 이용하여 고구마 덩이뿌리의 저온저장성을 향상시키기 위해, 저온저장 조건에서의 고구마의 생리, 생화학적 특성을 규명하고, 전사체 분석을 활용한 고구마 저온저장관련 유전자 네트워크 규명 및 유용유전자를 발굴하였다. 또한, 저온내성 및 저온저장성 관련 유용유전자를 도입한 형질전환 고구마를 제작하였다. 주요 결과와 중요한 의미는 다음과 같다. 1) 고구마 덩이뿌리 (cv. 율미)의 적정 (13℃) 및 저온 (4℃) 저장조건에서 0주, 6주간 저장한 시료로부터 생리, 생화학적 특성을 이해하고, 저온저장 전사체 분석을 통해 유전자 네트워크를 처음으로 규명하였다. DEG 분석법을 통해, 저온저장시 전반적으로 ‘불포화지방산 생합성’, ‘병원균 방어’, ‘페닐알라닌 대사’, ‘리그닌 합성’ 관련 유전자들의 발현이 증가하는 반면, ‘글리세롤인지질’, ‘항산화효소’, ‘탄수화물 및 에너지 대사’ 관련 유전자들의 발현이 감소함을 확인하였고, 이를 통해 저온저장관련 핵심유전자들을 확보하였다. 2) 저온저장기간에 따른 명확한 유전자 네트워크 기작을 규명하기 위해, 저온저장성 우수품종 (cv. Xushu 15-1) 및 취약품종 (cv. Xushu 15-4)의 고구마 덩이뿌리의 저온저장에 따른 생리, 생화학적 분석 결과를 근거로 0주, 저장초기(2주), 저장후기(6주) 시료를 선택하여 비교전사체분석에 이용하였다. DEG 분석을 통해, 우수품종이 취약품종에 비해 저장기간 동안 ‘막관련 스트레스 반응’, ‘번역’, ‘소포체 단백질 프로세싱’, ‘병원균 방어’, ‘탄수화물 대사’ 관련 유전자들의 발현이 전반적으로 높음을 확인하였다. 또한, AP2/ERF, Zinc finger, WRKY, NAC를 포함한 전사인자들의 유전자 발현의 증폭을 확인하였다. 더불어, SNP 탐지기술을 통해 106개의 후보 SNP를 탐색하였으며 분석 중에 있다. 추후 저온저장성 관련 핵심유전자를 추가로 확보하고, 저장성 관련 분자마커 개발에 활용하고자 한다. 3) 애기장대 유래 ribosomal P3 (AtP3B) 단백질은 식물 특이적 리보좀 P-단백질로서, RNA와 단백질을 보호하는 샤페론 기능으로 인해 저온과 고온에서 내성에 관여한다고 알려져 있다. AtP3B 유전자를 고구마 (cv. Xushu 29)에 과발현시킨 식물체 (이하 OP식물체)는 저온스트레스 (4℃)에서 WT식물체에 비해 높은 광합성 효율과 낮은 막투과성, 과산화수소 함량을 유지함으로써 저온스트레스에 증가된 내성을 보였다. 또한, 고온스트레스 (45℃)에서도 OP식물체는 WT식물체에 비해 높은 광합성효율과 낮은 전해질 누출을 나타냄으로써 높은 내성을 보였다. 더욱이, 수확한 OP덩이뿌리의 저온저장성을 평가한 결과, WT덩이뿌리에 비해 낮은 전해질 누출과 막투과성을 나타냄으로써 저온저장성이 향상됨을 확인 할 수 있었다. 4) 저온저장 전사체분석 결과로부터 분리한 저온저장성 관련 핵심유전자 가운데 lignin-forming anionic peroxidase (IbLfAP) 유전자와 fatty acid desaturase 8 (IbFAD8) 유전자의 기능을 분석하기 위해 이들을 과발현하는 형질전환 고구마 (cv. Xushu 29)를 제작하였고, 저온 및 저장성을 포함한 다양한 환경스트레스 조건에서 제반 특성을 분석할 예정이다. 본 연구를 통해 전사체 분석으로부터, 고구마 저온저장 중 발생하는 분자생물학적 기초 정보를 확보하였으며, SNP 분석을 통해 저온저장성 향상 품종 개발을 위한 육종선발마커에 이용할 수 있을 것으로 기대한다. 또한, 개발된 저온내성 및 저온저장성 향상 형질전환 고구마는, 저온 등 조건불리지역 재배 및 수확 후 저장 관점에서 21세기 인류가 당면한 식량 및 에너지 문제를 해결하는데 도움이 될 것이라고 기대한다. The dramatic increase in population and industrialization around the world has caused imbalances in the supply of food and energy. Food and Agriculture Organization (FAO) of UN estimate that the world population will be more than 9.1 billion by 2050 and require approximately 1.7 times the food and more than 3.5 times energy of today. In order to cope with these food and energy requirements, it is necessary to not only develop new crops with increased productivity that can be grown on marginal lands including desertification land and low temperature regions, but also, need to develop energy-saving postharvest storage technology. Sweetpotato [Ipomoea batatas (L.) Lam], which has been re-evaluated as the best health food and industrial crop to produce starch and useful materials such as low molecular weight antioxidants, is easy to cultivate in unfavorable conditions such as heat, drought and high salt. In light of these properties, sweetpotato is an industrially valuable starch crop. Nevertheless, sweetpotato originated tropical regions is vulnerable to low temperature, making it difficult to cultivate in low temperature regions. In addition, postharvest low temperature storage damage is a major constraint for sweetpotato industrialization due to problems such as chilling injury. In particular, low temperature storage of sweetpotato in winter season is a big problem from the view point of energy. In this study, to understand mechanism and improve storage ability of tuberous roots of sweetpotato under low temperature storage, I identified low temperature storage-related key genes and gene network using transcriptome analysis based on physiological and biochemical characteristics at low temperature storage conditions. In addition, transgenic sweetpotato plants expressing key genes involved in low temperature tolerance and storage ability were generated. The major results and significance in this study are as follows. 1) The first large-scale transcriptome analysis of sweetpotato tuberous roots stored at optimal (13℃) or low temperature (4℃) for 6 weeks was performed based on its physiological and biochemical analyses. In differentially expressed genes (DEG) analysis, genes involved in ‘biosynthesis of unsaturated fatty acids’, ‘pathogen defense’, ‘phenylalanine metabolism’, and ‘lignin biosynthesis’ were generally up-regulated at low temperature. By contrast, genes involved in ‘glycerophospholipid metabolism’, ‘antioxidant enzyme’, ‘carbohydrate metabolism’, and ‘energy metabolism’ were generally down-regulated under low temperature. These findings provided insight into key genes related to tuberous roots of sweetpotato under low temperature storage. 2) To investigate clear gene network mechanism under low temperature, comparative transcriptome analysis using tuberous roots of cultivars with high (cv. Xushu 15-1) and low (cv. Xushu 15-4) cold storage ability were carried out. Based on the physiological and biochemical analyses, samples were selected for 0 week, early storage (2 weeks), and late storage (6 weeks) and used for comparative transcriptome analysis under low temperature storage. The DEG analysis showed that the superior variety (high cold storage ability) generally showed a higher level of ‘membrane-associated stress response’, ‘translation’ , ‘endoplasmic reticulum protein processing’, ‘pathogen defense’, and ‘carbohydrate metabolism’ compared to weak variety (low cold storage ability). In addition, we confirmed the gene expression of transcription factors including AP2/ERF, Zinc finger, WRKY, and NAC. Furthermore, 106 candidate single nucleotide polymorphism (SNP) were analyzed through SNP molecular marker analysis. We will further secure key genes related to low temperature storage and use them for development of molecular markers related to storage ability. 3) P3 proteins are plant-specific ribosomal P-proteins that act as both RNA and protein chaperones to increase cold and heat stress tolerance in Arabidopsis. Here, I generated transgenic sweetpotato plants expressing the Arabidopsis ribosomal P3 (AtP3B) gene under the control of the CaMV 35S promoter (referred to as OP plants). Under low temperature stress (4℃), the OP plants exhibited enhanced low temperature tolerance with higher photosynthesis efficiency, less membrane permeability, and less hydrogen peroxide levels than wild type (WT) plants. The OP plants also displayed greater heat tolerance than WT plants, which was associated with higher photosynthesis efficiency and lower ion leakage after high temperature stress (45℃). Furthermore, the tuberous roots of OP transgenic sweetpotato showed improved storage ability with lower levels of ion leakage and membrane permeability compared to WT tuberous roots under low temperature storage condition at 4℃. 4) Transgenic sweetpotato plants were generated to analyze the function of lignin-forming anionic peroxidase (IbLfAP) and fatty acid desaturase 8 (IbFAD8), as representative genes related to low temperature storage, isolated from the transcriptome data. The characteristics of transgenic plants under various environmental stress conditions including low temperature and storability will be carried out. In conclusion, transcriptome and molecular data obtained from this study will be a valuable asset as a breeding selection marker and provide the basic information of molecular biology during low temperature storage of sweetpotato. In addition, transgenic sweetpotato with enhanced low temperature tolerance and storage ability may help to boost food security and energy problems for sustainable development in 21st century.

Volatile Composition, Sensory Attributes and Quantitative Trait Loci (QTLs) Associated With Flavor Traits in Cooked Sweetpotato (Ipomoea batatas)

Abugu, Modesta Nnedinso North Carolina State University ProQuest Dissertat 2025 해외박사(DDOD)

Consumers have identified flavor as one of the major traits that drive the consumption and liking of cooked sweetpotato. Knowledge of the key flavor-related volatile organic compounds (VOCs) are also highly valuable for breeders as they seek to develop new varieties for various industry segments. However, the lack of phenotypic data and modern breeding tools limits the selection of consumer preferred varieties and development of new flavor types for different markets. To bridge this gap, we carried out three studies focused on: 1) understanding the nature of VOCs in cooked sweetpotato; 2) identifying the key compounds that drive unique flavor attributes; and 3) exploring the genetics of these important VOCs.In Chapter one, we review the sweetpotato VOCs identified in the literature and summarize how these VOCs influence sensory perception and consumer preferences. Over 400 VOCs have been identified in cooked sweetpotato with over 76 known to be aroma active. Suggested mechanisms of formation of these active aroma VOCs are described. However, specific compounds that drive flavor in cooked sweetpotato are yet to be fully characterized. To aid the characterization of these VOCs, Chapter two focused on identifying key predictors of sweetpotato flavor through modelling the compounds identified in a biparental sweetpotato genetic mapping population (n = 416) with the flavor attributes identified in a subset of the population (n = 42) using Partial Least Square regression (PLS). 1284 VOCs differentiated the 416 genotypes, among which 240 were annotated in mass spectra matching. Additionally, the 42 genotypes were differentiated by 12 flavor attributes and 297 VOCs, including 158 not previously reported in sweetpotato. "Sweetpotato", "caramel/sweet aromatics", "cooked carrot", "pumpkin/squash", and "floral" were dominant distinguishing flavors, positively associated with benzyl mandelate, 2-furanmethanol, D-limonene, (-)-cis-myrtanol, and δ -3-carene respectively. Significant but less distinguishing flavors were "baked potato", "roasted chestnut", "earthy", "cooked green", and "sharp/sour aromatics", which were positively associated with 2(5H)- furanone, (Z)-4-hexen-1-ol, 2 unidentified VOCs, nerol oxide, (-)-myrtenol, and phenylethyl alcohol. The knowledge obtained from the VOCs, and sensory and prediction analyses form a basis for targeted flavor improvement by breeding.Understanding the genetic basis and identification of major or minor quantitative trait loci (QTLs) underlying the flavor traits may enable marker-assisted selection strategies in sweetpotato breeding programs. In Chapter three, we explored the genetic basis of the important flavor traits identified in the previous study, using a QTL analysis of 178 VOCs. 88 major-effect QTLs associated with 65 VOCs were detected in one growing season. Among them, 31 QTLs were identified for sesquiterpenes, while 15 and 4 QTLs were identified for monoterpenes, and carotenoid derived VOCs respectively. No QTLs were identified for the Maillard Reaction (MR) compounds which are known to predict "sweetpotato" and "caramel" flavor, suggesting that their formation is not genetically controlled. However, significant QTLs were detected for monoterpenes and apocarotenoids that positively predict "cooked carrot" flavor. This research will provide valuable insights on the components of sweetpotato flavor to guide breeders in developing new varieties that will ultimately enhance consumer experience.Finally, in Chapter four, we present a narrative review paper on the impact of international aid investment in plant breeding research in Sub-Saharan Africa (SSA), using root and tuber crops (Cassava, Sweetpotato, Yams, Plantains/Bananas and Potato) as a case study. Our review shows that investment in plant breeding research has significantly improved technology transfer, and availability of new tools for development of new/improved varieties. However, translating these tools to the development of improved varieties and uptake by farmers has not been completely successful. This is due, in part, to our lack of knowledge and ability to breed for texture and flavor traits that consumers prefer. In summary, advancing plant breeding research for development in SSA requires time, autonomy, an appreciation of agronomic and regional flavor characteristics and a more decentralized funding structure for local needs to be met.

Genomic Tools for Sweetpotato Variety Development: Genomic Selection, Fusarium Wilt Resistance, and High-Throughput Phenotyping for Guava Root-Knot Nematode Resistance

Fraher, Simon Phillip North Carolina State University ProQuest Dissertat 2025 해외박사(DDOD)

Sweetpotato (Ipomoea batatas (L.) Lam.) (2n = 6x = 90) is among the most important vegetable crops. Breeding sweetpotato cultivars for increased yield and quality traits, as well as resistance to pests and diseases, has been the focus of breeding programs globally. Genomic tools to facilitate earlier breeding decisions, like marker-assisted and genomic selection, have lagged largely due to the complex genetics of sweetpotato. Resources now exist that have allowed breeders to implement these technologies, including the development of reference genomes and genomic tools specific to polyploid crops. Chapter 1 of this dissertation is a literature review describing these resources and tools in more detail, as well as the production and breeding of sweetpotato and some of the constraints to variety development.Chapter 2 describes strategies for high-throughput phenotyping resistance to Meloidogyne enterolobii, the guava root-knot nematode, a quarantined pest in the state of North Carolina. These approaches utilized machine learning to identify and count nematode eggs, information which can inform breeders as to the resistance level of a given sweetpotato genotype. Using convolutional neural networks and human-defined parameters, a hybrid machine learning model was able to detect eggs as well as human evaluators (M. enterolobii R2 = 0.985, M. incognita R2 = 0.992, M. javanica R2 = 0.983). Automated counting protocols have the potential to save hundreds of hours of labor while enhancing genetic gain for resistance for plant breeders.Chapter 3 details a QTL analysis focused on resistance to Fusarium oxysporum f.sp. batatas, commonly called Fusarium wilt disease. This was once the most important disease in US sweetpotato production, however, cultivar resistance has largely addressed this issue. In breeding for resistance to M. enterolobii, population-level resistance to Fusarium wilt disease may decline as some nematode-resistant lines lack Fusarium resistance. Over three trials, we bioassayed a 454-clone biparental mapping population, NCDM04-0001 x 'Covington', and found a single major locus on chromosome 10, herein named qIbFo-10.1, that explained 33.8% of variation for resistance to Fusarium wilt, suggesting resistance may be controlled by one or a few tightly-linked loci. This locus should be considered a priority target for marker-assisted breeding.Chapter 4 of this dissertation addresses quantitative traits, including yield, shape, and USDA grade by deploying genome-wide markers which can be used to predict the parental breeding value of a given individual for these traits. We utilized the DArTag genotyping platform and performed field trials on both a training and breeding population, each with 528 genotypes representing the NC State sweetpotato breeding program's diversity. We compared BLUPs generated using mixed models and compared pedigree, genomic, and hybrid relationship matrices for trait predictions. Predictive abilities for the genomic BLUP model were highest in the breeding population for storage root count (r = 0.195) and length to diameter ratio (r = 0.317), and highest in the training population for total yield (r = 0.363) and USDA No. 1 storage root packout (r = 0.408). This provides evidence that genomic selection can increase the rate of genetic gain for quantitative traits in sweetpotato for the first time.Chapter 5 summarizes my experiences in the breeding program and includes details about several projects that were outside the scope of this dissertation. This chapter also identifies future research targets, including the use of drones for high-throughput phenotyping and future applications of genomic selection.This research strives to increase the rate of genetic gain for necessary traits for the sweetpotato breeding community. With the availability of genomic and high-throughput phenotyping tools, sweetpotato breeders can improve traits at the population-level, ultimately increasing the likelihood of selecting the next major sweetpotato variety.

Functional identification of IbAGL17 related to root formation and differentiation of sweetpotato (Ipomoea batatas (L.) Lam.)

Kwon, Na Yeong 서울시립대학교 일반대학원 2020 국내석사

Previous studies reported that IbAGL17 of sweetpotato is a MADS-box transcription factor and is highly expressed in tuberous roots. This study was conducted to analyze the structure and expression of IbAGL17 in sweetpotato and to investigate the relationship with IbAGL17 and the tuberous root formation. Amino acid sequence analysis of IbAGL17 confirmed that it belongs to the AGL17 subfamily. Genes in the subfamily are known to be involved in various plant developments such as flowering, germination, and root or nodule development. The qRT-PCR analysis showed that IbAGL17 was root-specifically expressed in sweetpotato and especially most significantly expressed in the tuberous root. It was also highly expressed in the pigmented root at 45 days, the early stage of development of the tuber, and most highly in the tuberous root during 60-90 days. In addition, Further experiments showed that IbAGL17 expression was regulated by various stress conditions. Based on those results, IbAGL17 may play a specific role in early signaling on the tuberous root formation of sweetpotato and may be involved in the stress response of plants. This study can be used as a basic research data on the formation of sweetpotato tuber and as a foundation on improving the productivity of sweetpotato. 이전의 연구로 고구마의 유전자 IbAGL17이 MADS-box 전사인자이고, 괴근에서 많이 발현한다는 것을 알 수 있었다. 이에 따라 본 연구는 고구마에서 분리한 유전자 IbAGL17의 구조와 발현을 분석하여 괴근 형성과의 관계를 알아보기위해 진행하였다. IbAGL17의 단백질 서열 분석 결과, AGL17 subfamily에 속하는 것을 확인할 수 있었다. 이 subfamily에 속하는 단백질들은 개화, 발아, 뿌리나 뿌리혹의 발달과 같은 식물의 다양한 발달에 관여하는 것으로 알려져 있다. IbAGL17의 발현 양상은 뿌리특이적인 발현을 보였으며, 다양한 뿌리 기관 중 괴근에서 가장 특이적으로 발현하였다. 또한, 괴근이 발달하는 초기 단계인 45일째의 pigmented root에서 많이 발현하였으며, 괴근이 두꺼워지는 60~90일동안 tuberous root에서 가장 많이 발현하였다. 그리고 추가적인 실험을 통해 다양한 스트레스 조건들에 의해 IbAGL17의 발현이 조절된 것을 확인할 수 있었다. 이러한 결과를 통해 보았을 때 IbAGL17은 고구마의 괴근 형성에 관한 초기 신호에 특이적인 역할을 하는 것으로 보이며, 이와 더불어 식물의 스트레스 반응에도 관여할 것으로 추정된다. 따라서 본 연구는 고구마의 괴근 형성에 대한 기초 연구자료로서의 활용 및 고구마의 생산성 증진에 관한 자료로 이용될 수 있을 것이다.

Managing Weeds that are Hosts for Root-Knot Nematodes in Sweetpotato

Ippolito, Stephen James ProQuest Dissertations & Theses North Carolina Sta 2025 해외박사(DDOD)

Weed interference with sweetpotato can reduce yield and leave it susceptible to pathogens and insects. Guava root-knot nematode (Meloidogyne enterolobii) is an emerging pest in North Carolina. Guava root-knot nematode can severely impact sweetpotato yield and quality. In addition, it is known for its aggressive nature, having a wide variety of both weed and crop hosts. Research was conducted to evaluate which weeds may act as a host for root-knot nematodes and identify herbicides which could be registered to control identified weed hosts in sweetpotato. In addition, yellow nutsedge (Cyperus rotundus L.) is a troublesome pest. However, there are few options for yellow nutsedge control in organic production systems. Research was conducted to develop a management program utilizing subsequent mowing and herbicide applications to improve yellow nutsedge control.A survey was conducted to identify which weeds may be acting as a host for root knot nematodes in sweetpotato producing counties in North Carolina. Summer weeds were excavated, then the roots were cleaned and examined for the presence or absence of root-knot nematode galling. The weed species with the highest prevalence across fields sampled and root-knot nematode incidence rate were: annual morningglory (Ipomoea spp.), prink purslane (Portulaca pilosa L.), carpetweed (Mollugo verticillata L.), Palmer amaranth (Amaranthus palmeri S. Watson), and Florida pusley (Richardia scabra L.).