Journal of Plant Biotechnology

The Korean Society of Plant Biotechnology (한국식물생명공학회)
권호 표지
DOI QR코드

DOI QR Code

Molecular Characterization and Event-Specific Marker Development of Insect Resistant Chinese Cabbage for Environmental Risk Assessment

환경위해성 평가를 위한 해충저항성 배추의 분자생물학적 특성 검정 및 계통 특이 마커 캐발

  • Lim, Sun-Hyung (Biosafety Division, National Institute of Agricultural Biotechnology, RDA) ;
  • Kim, Na-Young (Biosafety Division, National Institute of Agricultural Biotechnology, RDA) ;
  • Lee, Si-Myung (Biosafety Division, National Institute of Agricultural Biotechnology, RDA) ;
  • Woo, Hee-Jong (Biosafety Division, National Institute of Agricultural Biotechnology, RDA) ;
  • Shin, Kong-Sik (Biosafety Division, National Institute of Agricultural Biotechnology, RDA) ;
  • Jin, Yong-Moon (Biosafety Division, National Institute of Agricultural Biotechnology, RDA) ;
  • Cho, Hyun-Suk (Biosafety Division, National Institute of Agricultural Biotechnology, RDA)
  • 임선형 (농업생명공학연구원 생물안전성과) ;
  • 김나영 (농업생명공학연구원 생물안전성과) ;
  • 이시명 (농업생명공학연구원 생물안전성과) ;
  • 우희종 (농업생명공학연구원 생물안전성과) ;
  • 신공식 (농업생명공학연구원 생물안전성과) ;
  • 진용문 (농업생명공학연구원 생물안전성과) ;
  • 조현석 (농업생명공학연구원 생물안전성과)
  • Published : 2007.12.31
Download PDF
⟨ Previous Next ⟩

Abstract

Commercialization of genetically modified (GM) plants will be required the assessment of risks associated with the release of GM plants that should include a detailed risk assessment of their impacts in human health and the environment. Prior to GM plant release, applicants should provide the information on GM crops for approval. We carried out this study to provide the molecular data for risk assessment of the GM Chinese cabbage plants with insect-resistance gene, modified CryIAc, which we obtained by Agrobacterium-transformation. From the molecular analysis with GM Chinese cabbage, we confirmed the transgene copy number and stability, the expression of the transgene, and integration region sequences between the transgene and the Chinese cabbage genome. Based on the unique integration DNA sequences, we designed specific primer set to detect GM Chinese cabbage and set up the GM cabbage detection method by qualitative PCR analysis. Qualitative analysis with GM Chinese cabbage progenies analysis was revealed the same as the result of herbicide treatment. Our results provided the molecular data for risk assessment analysis of GM Chinese cabbage and demonstrated that the primer set proposed could be useful to detect GM Chinese cabbage.

유전자변형 작물의 상업화를 위해서는 유전자변형 작물이 식품으로서의 안전성과 환경에 미치는 영향에 관한 평가가 이루어져야한다. 이를 위해 개발자는 유전자변형 작물의 방출이전에 유전자변형 작물 개발에 관한 정보를 제출해야만 한다. 본 연구는 유전자변형 작물의 환경 위해성 평가를 위한 분자생물학적 자료를 제공하고자 수행하였다. 아그로 박테리움 형질전환법을 통하여 해충저항성 CryIAc 유전자가 도입된 배추 형질전환체를 획득한 후, 분자생물학적인 분석을 통하여 유전자의 copy수, 안정성, 식물체내에서의 발현을 확인하였고, T-DNA의 배추게놈내의 인접서열을 분석하였다. T-DNA의 게놈내 삽입 인접서열을 바탕으로 유전자변형 배추를 동정할 수 있는 프라이머를 제작하였고, 이를 이용한 검정방법을 수립하였다. 계통 특이 프라이머를 이용한 해충저항성 배추 후대의 PCR 분석결과와 제초체 처리결과가 서로 일치하였다. 본 연구 결과로 환경위해성 평가를 위한 해충저항성 배추의 분자생물학적인 자료를 획득하였으며, 개발된 프라이머는 해충저항성 배추의 검출을 위하여 유용하게 사용할 수 있음을 확인하였다.

Keywords

References

  1. 농림부 (2002) 유전자변형농산물의 환경위해성 평가지침. 농림부고시 제2002-02호
  2. Berdal KG, Holst-Jensen A (2001) Roundup ready soybean event specific real-time quantitative PCR assay and estimation of the practical detection and quantification limits in GMO analysis. Eur Food Res Technol 213: 432-438 https://doi.org/10.1007/s002170100403
  3. Cankar K, Ravnikar M, Zel J, Gruden K, Toplak N (2005) Real-time polymerase chain reaction detection of cauliflower mosaic virus to complement the 35S screening assay for genetically modified organisms. J AOAC lnt 88: 814-822
  4. Collonnier C, Schattner A, Berthier G, Boyer F, Coue-Philippe G, Diolez A, Duplan MN, Fernandez S, Kebdani N, Kobilinsky A, Romaniuk M, De Beuckeleer M, De Loose M, Windels P, Bertheau Y (2005) Characterization and event specific-detection by quantitative real-time PCR of T25 maize insert. J AOAC Int 88: 536-546
  5. Hardegger M, Brodmann P, Herrmann A (1999) Quantitative detection of the 35S promoter and the NOS terminator using quantitative competitive PCR. Eur Food Res Technol. 209: 83-87 https://doi.org/10.1007/s002170050462
  6. Hernandez M, Pia M, Esteve T, Prat S, Puigdomenech P, Ferrando A (2003) A specific real-time quantitative PCR detection system for event MON810 in maize $YieldGard^{\circledR}$ based on the 3'-transgene integration sequence. Transgenic Res 12: 179-189 https://doi.org/10.1023/A:1022979624333
  7. Hupfer C, Hotzel H, Sachse K, Engel KH (1998) Detection of the genetically modification in heat products of Bt maize by polymerase chain reaction. Z Lebensm Unters Forsch 206: 203-207 https://doi.org/10.1007/s002170050243
  8. James C (2006) Global status of commercialized biotech/GM crops in 2006. ISAAA Briefs No.35-2006
  9. James D, Schmidt AM, Wall E, Green M, Marshi S (2003) Reliable detection and identification of genetically modified maize, soybean, and canola by multiplex PCR analysis. J Agri Food Chem 51: 5829-5834 https://doi.org/10.1021/jf0341159
  10. James D, Schmidt AM, Wall E, Green M, Masri S (2003) Reliable detection and identification of genetically modified maize, soybean, and canola by multiplex PCR analysis. J Agric Food Chem 51: 5829-5834 https://doi.org/10.1021/jf0341159
  11. Kim JS (2002) Composite of a linkage map of Brassica rapa (ssp. pekinenssiS) using EST clones and comparative genome study to Arabidopsis thaliana. PhD thesis, Kyunghee University, Suwon, Republic of Korea
  12. Kim YM, Sohn SH, Jeong SI, Yoon MS, Kim TS, Park YH (2002) Detection methods for genetically modified soybeans. J Korean Soc Agri Chem Biotechnol 45: 185-189
  13. Lee SW (2003) Current research status on the development of genetically modified plants in Korea. Kor J Plant Biotech 30: 1-6 https://doi.org/10.5010/JPB.2003.30.1.001
  14. Malhotra K, Foltz L, Mahoney WC, Schueler PA (1998) Interaction and effect of annealing temperature on primers used in differential display RT-PCR. Nucleic Acids Research 26: 854-856 https://doi.org/10.1093/nar/26.3.854
  15. Matsuoka T, Kuribara H, Akiyama H, Miura H, Goda Y, Kusakabe Y, Isshiki K, Toyoda M, Hino A (2001) A multiplex PCR method of detecting recombinant DNAs from five lines of genetically modified maize. J Food Hyg Soc Japan 42: 24-32 https://doi.org/10.3358/shokueishi.42.24
  16. Matsuoka T, Kuribara H, Takubo K, Akiyama H, Miura H, Goda Y, Kusakabe Y, Isshiki K, Toyoda M, Hino A (2002) Detection of recombinant DNA segments introduced to genetically modified maize (Zea mays). J Agric Food Chem 50: 2100-2109 https://doi.org/10.1021/jf011157t
  17. Murashige T, Skoog F (1962) A revised medium for rapid growth and bioassays with tobacco tissue cultures. Physiol Plant 15: 473-497 https://doi.org/10.1111/j.1399-3054.1962.tb08052.x
  18. Nielsen CR, Berdal KG, Holst-Jensen A (2004) Characterisation of the 5' -integration site and development of an eventspecific real-time PCR assay for NK603 maize from a low starting copy number. Eur Food Res Technol 219: 421-427
  19. Rho JK, Lee T, Jung SI, Kim TS, Park YH, Kim YM (2004) Qualitative and quantitative PCR methods for detection of three lines of genetically modified potatoes. J Agric Food Chem 52: 3269-3274 https://doi.org/10.1021/jf0499020
  20. Ronning SB, Vaitilingom M, Berdal KG, Holst-Jensen A (2003) Event specific real-time quantitative PCR for genetically modified Btll maize (Zea mays). Eur Food Res Technol 216: 347-354 https://doi.org/10.1007/s00217-002-0653-4
  21. Southern EM (1975) Detection of specific sequences among DNA fragments. J Mol Biol 98: 503-517 https://doi.org/10.1016/S0022-2836(75)80083-0
  22. Taverniers I, Windels P, Vaitilingom M, Milcamps A, Van Bockstaele E, Van Den Eede G, De Loose M (2005) Event-specific plasmid standards and real-time PCR methods for transgenic Bt11, Bt176, and GA21 Maize and transgenic GT73 canola. J Agric Food Chem 53: 3041-3052 https://doi.org/10.1021/jf0483467
  23. Vaitilingom M, Pijnenburg H, Gendre F, Brignon P (1999) Real-time quantitative PCR detection of genetically modified maximizer maize and RoundupReady soybean in some representative foods. J Agric Food Chem 47: 5261-5266 https://doi.org/10.1021/jf981208v
  24. Windels P, Bertrand S, Depicker A, Moens W, Van Bockstaele E, De Loose M. (2003). Qualitative and event-specific PCR real-time detection methods for StarUnk maize. Eur Food Res Technol 216: 259-263 https://doi.org/10.1007/s00217-002-0652-5
  25. Woo HJ, Lim SH, Lee KJ, Won SY, Kim TS, Cho HS, Jin YM. (2006) Current development status on the genetically modified crops in Korea. Korean J Intl Agri 18: 221-229
  26. Yang L, Pan A, Zhang K, Yin C, Qian B, Chen J, Huang C, Zhang D (2005) Qualitative and quantitative PCR methods for event specific detection of genetically modified cotton MON1445 and MON531. Transgenic Res 14: 817-831 https://doi.org/10.1007/s11248-005-0010-z
  27. Yang R, Xu W, Luo Y, Guo F, Lu Y, Huang K (2007) Event-specific qualitative and quantitative PCR detection of roundup ready event GT73 based on the 3' -integration junction. Plant Cell Rep 43: 250-257
  28. Zimmermann A, Liniger M, Luthy J, Pauli U (1998) A sensitive detection method for genetically modified $MaisGard^{TM}$ com using a nested PCR-system. Lebensm-Wissu Technol 31: 664-667 https://doi.org/10.1006/fstl.1998.0422

Cited by

  1. Qualitative and quantitative PCR detection of insect-resistant genetically modified rice Agb0101 developed in korea vol.40, pp.1, 2013, https://doi.org/10.5010/JPB.2013.40.1.018
  2. Event-specific qualitative and quantitative polymerase chain reaction methods for detection of insect-resistant genetically modified Chinese cabbage based on the 3′-junction of the insertion site vol.55, pp.3, 2012, https://doi.org/10.1007/s13765-012-2028-2
  3. Qualitative PCR Detection of Stack Gene GM Rice (LS28 X Cry1Ac) Developed in Korea vol.52, pp.1, 2009, https://doi.org/10.3839/jabc.2009.001