본문 바로가기
KERD
논문KCI 등재

고추 역병 저항성 후보 유전자 삽입을 통한 담배(Nicotiana benthamiana) 형질전환체 개발 최적화

Development of Agrobacterium-mediated Transformation of Nicotiana benthamiana through Insertion of Candidate Gene for Phytophthora capsici Resistance in Pepper

강원희|권지수|염지은
농업생명과학연구원 (Institute of Agriculture & Life Science, Gyeongsang National University)|2023|농업생명과학연구 (Journal of Agriculture & Life Science)|제57권 제1호|pp. 17-24

1

조회

0

다운로드

1

인용

0

참고문헌

키워드

Agrobacterium tumefaciens고추 역병균담배 형질전환저항성 유전자Candidate geneNicotiana benthamiana transgenicPhytophthora capsici

초록

고추역병균(Phytophthora capsici)은 고추 생육 전반에 걸쳐 병을 발생시켜 농가 소득에 큰 손실을 일으키고 있다. 고추 역병균 저항성은 양적형질 유전자좌(Quantitative Trait Loci, QTL)에 의해 조절되며 주동 유전자는 고추의 5번 염색체에 존재한다고 보고 되었지만, 후보 유전자의선발 및 저항성 유전자 규명 연구는 아직 초기 단계이다. 특히, 고추는 형질전환이 어려운 작물로써 병원균과의 상호작용 연구를 통한 저항성유전자 동정에 제한이 많다. 반면 고추와 같은 가지과 작물인 담배(Nicotiana benthamiana)는 병원균 상호작용 모델로 알려져 형질전환을 통해저항성 유전자 규명에 활용된다. 본 연구에서는 고추 역병 저항성 기작 규명을 위한 기초 연구로써, 식물 저항성 유사 유전자(Resistance GeneAnalog, RGA)를 선발하고, 이들 유전자들에 대한 담배 형질전환 기법 최적화 연구를 수행하였다. 고추 5번 염색체에 존재하는 고추 역병 저항성분자표지들을 분석하여 RGA 후보 유전자인 CaNBARC105, CaNBARC112 유전자를 동정하였다. 이들 유전자들에 대해 Agrobacteriumtumefaciens를 매개체로 하여 고추 RGA가 삽입된 담배 형질전환체를 개발하였다. 형질전환 여부는 유전자 특이적인 서열을 이용한 genomicPCR과 RT-PCR 검증을 통해 이들 형질전환 된 담배들의 생육 및 발달에 영향이 없다는 것을 확인하였다. 본 연구는 향후 고추 병 저항성 후보유전자들이 삽입된 담배 형질전환체는 고추 역병 저항성 유전자 규명 및 기작 연구에 기반이 될 것이다

Abstract

Phytophthora capsici (P. capsici) is a soil-borne fungal that causing Phytophthora blight with the significant problem to chili pepperfarming in wide-spread globally. To date, the resistance of P. capsici regulated by several minor quantitative trait loci (QTLs) a consistentmajor QTL on chromosome 5. The study is more needed such as the selection of candidate genes and identification of their function. Especially, the Capsicum annuum is difficult to transformation, and there are many limitations in identifying resistance genes throughinteraction studies with pathogens. Hence, Nicotiana benthamiana (N. benthamiana), a Solanaceae crop such as chili pepper, is knownas a pathogen interaction model source and is useful for identifying the function of resistance genes through transformation. In thisstudy, an investigation of the resistance mechanism to P. capsici, a plant resistance gene analog (RGA) was selected, and optimizationof N. benthamiana transformation technique was performed on these genes. The CaNBARC105 and CaNBARC112 were identified asRGA candidates through analysis of the known P. capsici resistance molecular markers on chromosome 5 in pepper. We developed a N. benthamiana transformation into which chili pepper RGA was inserted based on the Agrobacterium tumefaciens for these two genesand we confirmed through genomic PCR and RT-PCR using gene-specific sequence. Also, it was verified phenotypes that this transformedN. benthamiana was no effect on its growth and development. This transformed N. bentahmiana will be helpful as a basis for theidentification of the Phytophthora blight resistance gene and mechanistic research in chili pepper.

참고문헌 (37)

  1. Champouret N, "Functional genomics of Phytophthora infestans effectors and Solanum resistance genes", pp.2010
  2. Consortium TPGS, "Genome sequence and analysis of the tuber crop potato", Nature, 475, pp.189~195 (2011)
  3. Consortium TTG, "The tomato genome sequence provides insights into fleshy fruit evolution", Nature, 485, pp.635~641 (2012)
  4. "Producer prices" (2020)[URL]
  5. Goodin MM, "Nicotiana benthamiana : Its history and future as a model for plant–pathogen interactions", Mol. Plant Microbe Interact, 21, pp.1015~1026 (2008)
  6. Hue S., "Nonhost resistance of the plants induced by the phytopathogenic bacteria", Hortic. Sci. Technol, 19, pp.617~622 (2001)
  7. Ji P, "Effect of oxathiapiprolin on asexual life stages of Phytophthora capsici and disease development on vegetables", Ann. Appl. Biol, 166, pp.229~235 (2015)
  8. Kaila L., "Inheritance and stability of late blight resistance in potato population B3 of the international potato center", Master, pp.2015
  9. 강원희, "Genome-wide Identification, Classification, and Expression Analysis of the Receptor-Like Protein Family in Tomato", The Plant Pathology Journal, 34(5), pp.435~444 (2018), 한국식물병리학회[DOI]
  10. Kang WH, "Genome-wide analysis of Dof transcription factors reveals functional characteristics during development and response to biotic stresses in pepper", Sci. Rep, 6, pp.1~12 (2016)
  11. Won-Hee Kang, "Universal gene co-expression network reveals receptor-like protein genes involved in broad-spectrum resistance in pepper (<i>Capsicum annuum<", i>L.), 2022, pp.9, ~[DOI]
  12. 강원희, "Differential Responses of Capsicum Accessions to Foliar Blight and Root Rot by Phytophthora capsici", 원예과학기술지, 40(1), pp.85~93 (2022), 한국원예학회[DOI]
  13. Kang WH, "Transcriptome profiling of abiotic responses to heat, cold, salt, and osmotic stress of Capsicum annuum L", Sci. Data, 7, pp.1~7 (2020)
  14. 김동영, "Analysis of Genes Expressed during Pepper-Phytophthora capsici Interaction using EST Technology", 생명과학회지, 24(11), pp.1187~1192 (2014), 한국생명과학회
  15. 김동환, "Isolation of putative pepper defense-related genes against the pathogen Phytophthora capsici using suppression subtractive hybridization", macroarray and RNA-sequencing analyses, 2019, pp.60(5), 685~699[DOI]
  16. Hyoun-Joung Kim, "Marker Production by PCR Amplification with Primer Pairs from Conserved Sequences of WRKY Genes in Chili Pepper", Molecules and Cells, 25(2), pp.196~204 (2008), 한국분자세포생물학회
  17. Kim HJ, "BAC-derived markers converted from RFLP linked to Phytophthora capsici resistance in pepper (Capsicum annuum L.)", Theor. Appl. Genet, 118, pp.15~27 (2008)
  18. Kim MS, "Global gene expression profiling for fruit organs and pathogen infections in the pepper, Capsicum annuum L", Sci. Data, 5, pp.1~6 (2018)
  19. Kim N, "Development of clustered resistance gene analogs-based markers of resistance to Phytophthora capsici in chili pepper", Biomed Res. Int, 2019 (2019)[DOI]
  20. Kim S, "Genome sequence of the hot pepper provides insights into the evolution of pungency in Capsicum species", Nat. Genet, 46, pp.270~278 (2014)
  21. Kim SB, "Divergent evolution of multiple virus‐resistance genes from a progenitor in Capsicum spp", New Phytol, 213, pp.886~899 (2017)
  22. Kwon JS, "Leaf-to-whole plant spread bioassay for pepper and Ralstonia solanacearum interaction determines inheritance of resistance to bacterial wilt for further breeding", Int. J. Mol. Sci, 22, pp.2279~ (2021)[DOI]
  23. Lamour KH, "The oomycete broad-host-range pathogen Phytophthora capsici", Mol. Plant Pathol, 13, pp.329~337 (2012)
  24. Lee HA, "Plant NB-LRR proteins: Tightly regulated sensors in a complex manner", Brief Funct Genomics, 14, pp.233~242 (2015)
  25. 이흥렬, "QTL Analysis of Fruit Length Using rRAMP, WRKY, and AFLP Markers in Chili Pepper", Horticulture, Environment, and Biotechnology, 52(6), pp.602~613 (2011), 한국원예학회
  26. Lee JB, "Antifungal activity of Bacillus sp. AM-651 against Phytophthora capsici", Microbiol. Biotechnol. Lett, 36, pp.227~232 (2008)[DOI]
  27. Liu WY, "Combined use of bulked segregant analysis and microarrays reveals SNP markers pinpointing a major QTL for resistance to Phytophthora capsici in pepper", Theor. Appl. Genet, 127, pp.2503~2513 (2014)
  28. Ogundiwin EA, "Construction of 2 intraspecific linkage maps and identification of resistance QTLs for Phytophthora capsici root-rot and foliar-blight diseases of pepper (Capsicum annuum L.)", Genome, 48, pp.698~711 (2005)
  29. Renfiyeni Y, "Calli induction of some chili pepper (Capsicum annuum l.) genotypes as material for genetic transformation", Int. J. Agric. Sci, 1, pp.75~80 (2017)
  30. Seo E, "Genome-wide comparative analyses reveal the dynamic evolution of nucleotide-binding leucine-rich repeat gene family among Solanaceae plants", Front. Plant Sci, 7, pp.1205~ (2016)[DOI]
  31. 서은영, "Ectopic Expression of Capsicum-Specific Cell Wall Protein Capsicum annuum Senescence-Delaying 1 (CaSD1) Delays Senescence and Induces Trichome Formation in Nicotiana benthamiana", Molecules and Cells, 33(4), pp.415~422 (2012), 한국분자세포생물학회[DOI]
  32. Van Der Vossen E, "An ancient R gene from the wild potato species Solanum bulbocastanum confers broad-spectrum resistance to Phytophthora infestans in cultivated potato and tomato", Plant J, 36, pp.867~882 (2003)
  33. Van Ooijen G, "Structure–function analysis of the NB-ARC domain of plant disease resistance proteins", J. Exp. Bot, 59, pp.1383~1397 (2008)
  34. Wang P, "Molecular mapping of a gene conferring resistance to Phytophthora capsici Leonian race 2 in pepper line PI201234(Capsicum annuum L.)", MOl. Breed, 36, pp.1~11 (2016)
  35. Yeom SI, "Use of a secretion trap screen in pepper following Phytophthora capsici infection reveals novel functions of secreted plant proteins in modulating cell death", Mol. Plant Microbe Interact, 24, pp.671~684 (2011)
  36. Yeom SI, "A common plant cell‐wall protein HyPRP1 has dual roles as a positive regulator of cell death and a negative regulator of basal defense against pathogens", Plant J, 69, pp.755~768 (2012)
  37. You-Xiong Q, "Isolation and characterization of NBS-LRR resistance gene analogs from sugarcane", Acta Agron. Sin, 35, pp.631~639 (2009)

관련 논문AI추천

고추의 역병 저항성 품종 개발을 위하여 Agrobacterium tumefaciens를 이용한 elicitin 유전자 도입권태룡, 김경민, 김창길, 신동현, 오중열, 이문중, 한증술한국식물생명공학회 · Journal of Plant Biotechnology2007 · 제34권 제1호고추 유전자변형체 후대 생육특성 검정권태룡, 김경민, 김창길, 신동현, 오중열, 이문중, 한증술한국자원식물학회 · 한국자원식물학회지2008 · 제21권 제4호Soybean mosaic virus의 외피단백질유전자 형질전환 담배의 Heterologous potyvirus에 대한 저항성김율호, 손수인, 이부영, 이장용, 이현숙, 허한순한국육종학회 · 한국육종학회지2004 · 제36권 제5호고추약한모틀바이러스 병원형 P1,2 및 P1,2,3 생물검정을 통한 저항성 고추유전자원 선발곽해련, 김빛샘, 김성훈, 노나영, 이수경, 최유미, 한범수, 허온숙, 황애진한국육종학회 · 한국육종학회지2022 · 제54권 제1호Fluidigm 시스템을 이용한 고추 유전자원의 주요병 유전형 평가고호철, 금보라, 노나영, 이수경, 최유미, 허온숙, 왕샤오한한국육종학회 · 한국육종학회지2021 · 제53권 제4호Agrobacterium을 이용한 고추의 Transient Expression 시스템성은수, 정영희, 최도일한국식물생명공학회 · Journal of Plant Biotechnology2004 · 제31권 제3호

이 간행물의 다른 논문 (14건)

국내외 칡 제거 방법에 대한 비교 고찰박지현, 이상태, 김기우, 서다혜, 박준형pp. 1-15음식물류폐기물 혼합 가축분 퇴비 연용 시 시설 토마토 생육 및수량에 미치는 영향조현종, 심재홍, 이승하, 나홍식, 정영재, 권오연pp. 25-34콩 재배시 식이유황 처리에 따른 생육 및 수량성 평가채세은, 심두보, 전승호, 조영손, 오승가pp. 35-43문관과 미성숙 종자로부터 부정아 유도를 통한 재분화안찬훈, 이현석pp. 45-50중부지방 대왕참나무의 수간곡선식 추정에 관한 연구임종수, 이영진, 강진택, 장용환pp. 51-59
원문 보기