Journal of Plant Biotechnology

  • 제29권4호
  • /
  • Pages.235-240
  • /
  • 2002
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  • 1229-2818(pISSN)
  • /
  • 2384-1397(eISSN)
한국식물생명공학회 (The Korean Society of Plant Biotechnology)
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감자 특이 Internal Control DNA 증폭용 Primer와 이를 이용한 유전자 변형 감자의 경쟁적 이중 PCR 검정법

Primer for the Potato Specific Internal Control DNA and Screening Method for the Genetically Modified Potatoes by Competitive Duplex-PCR

  • 서효원 (농촌진흥청 고령지농업시험장) ;
  • 이정윤 (농촌진흥청 고령지농업시험장) ;
  • 조현묵 (농촌진흥청 고령지농업시험장) ;
  • 김숭열 (농촌진흥청 고령지농업시험장)
  • Seo, Hyo-Won (Highland Crop Research Division, National Highland Agricultural Experiment Station, RDA) ;
  • Yi, Jung-Yoon (Highland Crop Research Division, National Highland Agricultural Experiment Station, RDA) ;
  • Cho, Hyun-Mook (Highland Crop Research Division, National Highland Agricultural Experiment Station, RDA) ;
  • Kim, Sung-Yeul (Highland Crop Research Division, National Highland Agricultural Experiment Station, RDA)
  • 발행 : 2002.12.01
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초록

현재 유전자 변형 작물의 검정에는 CaMV 35S프로모터 혹은 NOS 터미네이터 등과 같이 형질전환에 널리 이용하는 유전인자를 검출하기 위한 PCR 기술이 널리 이용되고 있다. 본 연구에서는 유전자변형 감자를 검정하기 위한 새로운 기술로서 감자특이 internal control과 CaMV 35S 프로모터 혹은 NOS 터미네이터에 특이적인 프라이머를 이용한 경쟁적 이중PCR 방법을 개발하였다. 감자 유전자의 RAPD 결과 이용한 모든 품종에서 약 530 bp인 homozygous DNA 밴드를 증폭하는 특이 primer (rAGU4A)를 찾아 감자특이 internal control 증폭용으로 이용하였다. 이 프라이머에 의해 증폭되는 DNA는 TC 염기의 반복 빈도가 높은 repetitive 혹은 microsatellite DNA (AF541972)로 판단되었다. 이와 같은 단일 프라이머 internal control 증폭용으로 이용한 경쟁적 이중 PCR은 비특이적 PCR 산물을 줄일 수 있고, 기존 방법들에 비해 경제적이다. 현재 상품화되어 있는 유전자 변형 감자품종인 'New Leaf'의 경우도 형질전환에 이용한 유전인자로 CaMV 35S 프로모터와 NOS 터미네이터가 모두 이용되었으므로 이 기술을 이용할 경우 현재까지 상품화된 유전자 변형감자들의 효율적인 검정 기술로 이용될 수 있을 것으로 기대된다.

We report the new method for the screening of genetically modified potato by competitive duplex-PCR using the potato specific single oligomer primer for the internal control and CaMV 35S promoter or NOS terminator specific primers. The single oligomer primer (rAGU4A) amplify the potato specific internal control band from the homozygous potato genomic DNA in the RAPD profiles of all analyzed potato varieties. The 530 bp internal control DNA was amplified independently to CaMV 35S promoter or NOS terminator DNA and identified as repetitive or microsatellite DNA of potato (AF541972). With this new technique, the transgenic potatoes which were transformed with vectors contained the different foreign genes are analyzed. In case of the commercialized transgenic potato varieties, 'Hew Leafs', those two genetic factors are used for promoter and terminator respectively So, this new PCR technique should be a promising method of cost effective and accurate screening for the commercialized GM potatoes on market.

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참고문헌

  1. Anonymous (2002) http://www.bats.ch/about/projekte.htm
  2. Bonierbale MW, Plaisted RL, Tanksley SD (1991) Application of restriction fragment length polymorphisms and genetic mapping to potato breeding. In : International Potato Center, Molecular Methods for Potato Improvement. Report of the Planning Conference on 'Application of Molecular Techniques to Potato Germplasm Enhancement'. CIP, Peru, PP. 149-157
  3. Brunnert HJ, Spener F, Borchers T (2001) PCR-ELISA for the CaMV-35S promoter as a screening method for genetically modified Roundup Ready soybeans. Eur Food Res Technol 213: 366-371 https://doi.org/10.1007/s002170100371
  4. Chiueh LC, Chen YL, Daniel TCS (2002) Study on the detection method of six varieties of genetically modified maize and processed foods. J Food Drug Anal 10:25-33
  5. Elliott AR, Campbe JA, Dugdale B, Brettel RSI, Grof CPL (1999) Green-fluorescent protein facilitates rapid in vivo detection of genetically transformed plant cells. PIant Cell Rep 18:707-714 https://doi.org/10.1007/s002990050647
  6. Hall TC (2002) Current status of GMO: development and perspective. In : International symposium on agrobiotechnology for 21st century, RDA, Suwon Korea, PP.79-102
  7. $H\ddot{u}bner$ P, Studer E, $L\ddot{u}thy$ J (1999) Quantitative competitive PCR for the detection of genetically modified organism in food. Food Control 10:353-358 https://doi.org/10.1016/S0956-7135(99)00074-2
  8. Grunst T, Kephart D (2001) Wizard Magnetic DNA purification system for food: Part II. Semi-automated DNA isolation and analysis of GMO foods. Pomega Note 76:19-22
  9. James C (2002) Global hectarage of GM crops in 2001. Crop Biotech Brief Vol.II No. 1
  10. Kang HW, Park DS, Go SJ, Eun MY (2002) Fingerprinting of diverse genomes using PCR with Universal Rice Primers generated from prpetitive sequence of Korean Weed Rice. Mol Cells 13:281-287
  11. Lipp M, Brodmann P, Puetsch K, Pauwels J, Anklam E (1999). IUPAC collaborative trial study of a method to detect genetically modified soybeans and maize in dried powder. J AOAC Int 82: 923-928
  12. $Mannerl\ddot{o}ef$ M, Tenning P (1996) Screening of transgenic plant by multiplex PCR. Plant Mol Biol Rep 15:38-45 https://doi.org/10.1007/BF02772111
  13. Meyer R (1999) Development and application of DNA analytical methods for the detection of GMOs in food. Food Control 10: 391-399 https://doi.org/10.1016/S0956-7135(99)00081-X
  14. Seo HW (2001) Genetic engineering of virus resistant potatoes and development of nucleic acid diagnostic methods using coat protein gene of Potato leafroll virus. PhD thesis, Konkuk University, Seoul. pp. 51-81
  15. Seo HW, Yi JY, Cho HM, Cho JH, Park YE (2001) Development of maturity controllable transgenic potato with GA 3beta-hydroxylase. In : Annual Research Report of National Highland Agiicultural Experiment Station, RDA, Korea. pp. 342-345
  16. Spoth B, Strauss E (1999) Screening for genetically modified organisms in food using Promegas Wizard Resin. Promega Notes 73:23-25
  17. Tao Z, Cai XF, Yang SL, Gong Y (2001) Detection of exogenous genes in genetically modified with multiplex polymerase chain reaction. Plant Mol Biol Rep 19:289-298 https://doi.org/10.1007/BF02772827
  18. Teresa MF, Julapark C, Tanksley SD (1995) Microprep protocol for extraction of DNA from tomato and other herbaceous plants. Plant Mol Biol Rep 13: 207-209 https://doi.org/10.1007/BF02670897
  19. Tozzini AC, Martinez MC, Lucca, MF, Rovere CV, Distefano AJ, del Vas M, Hopp HE (2000) Semi-quantitative detection of genetically modified grains based on CaMV 35S promoter amplification. Electric Journal of Biotechnology 3:149-153
  20. Vickers JE, Graham GC, Henry RJ (1996) A protocol for the screening of putatively transformed plant for bar, the selectable marker gene, using the polymerase chain reaction. Plant Mol Biol Rep 14:363-368 https://doi.org/10.1007/BF02673368
  21. Wurz, A, Bluth A, Zeltz P, Pfeifer C, Willmund R (1999) Quantitative analysis of genetically modified organism (GMO) on processed food by PCR-based methods. Food Control 10:385-389 https://doi.org/10.1016/S0956-7135(99)00080-8
  22. Yi JY, Lee SW, Park KW (1998) Introduction of Shiva gene into tobacco and potato using tissue-specific tomato PAL promoter. Korean J. PIant Tissue Culture 25:109-113