생명과학회지 (Journal of Life Science)

  • 제16권3호
  • /
  • Pages.540-545
  • /
  • 2006
  • /
  • 1225-9918(pISSN)
  • /
  • 2287-3406(eISSN)
한국생명과학회 (Korean Society of Life Science)
권호 표지
DOI QR코드

DOI QR Code

애기장대에 있어서 shoot 발달 연구를 위한 프로모터 trap 라인들의 제조 및 선별

Generation and Selection of Promoter Trap Lines for the Investigation of Shoot Development in Arabidopsis

  • 이화목 (부산대학교 분자생물학과) ;
  • 박희연 (부산대학교 분자생물학과) ;
  • ;
  • 이춘환 (부산대학교 분자생물학과) ;
  • 문용환 (부산대학교 분자생물학과)
  • Lee Hwa-Mok (Department of Molecular Biology, Pusan National University) ;
  • Park Hee-Yeon (Department of Molecular Biology, Pusan National University) ;
  • Zulfugarov Ismayil S. (Department of Molecular Biology, Pusan National University) ;
  • Lee Choon-Hwan (Department of Molecular Biology, Pusan National University) ;
  • Moon Yong-Hwan (Department of Molecular Biology, Pusan National University)
  • 발행 : 2006.06.01
PDF 다운로드
⟨ 이전 논문 다음 논문 ⟩

초록

T-DNA 매개 형질전환은 삽입 돌연변이를 가지는 형질전환 식물체를 만들기 위한 유용한 방법이다. 애기장대의 shoot 발달 과정에서 중요한 역할을 하는 유전자를 확인하기 위해, 프로모터 trap 식물체 라인을 제작하고 분석하였다. 이를 위해 효율적인 프로모터 trap 백터인 pFGL561을 제작하였다. pFGL561은 basta 저항성 유전자, multiple splicing donor acceptor 서열들, 그리고 프로모터가 없는 GFP 리포터 유전자를 포함하고 있다. Agrobacterium 균주인 GV3101에 pFGL561을 도입하고, 이를 매개로 하여 300개의 $T_1$ 프로모터 trap 식물체를 제작하였고, 이들 식물체에서 GFP 발현을 조사하였다. $T_1$ 식물체에서 GFP 발현 비율은 26.7%로 매우 높았고, 이러한 발현은 $T_2$ 식물체에서도 재확인되었다. 한편, 식물의 shoot 발달에 관여하는 주요 유전자를 동정하기 위해서, shoot에서 GFP발현을 보인 19개 $T_1$ 식물체에서 유래한 $T_2$ 식물체를 대상으로 표현형을 조사한 결과, 비정상적인 shoot발달을 보이는 6개 $T_1$ 라인을 확인하였다. 이들 식물체는 shoot발달의 조절 기작 분석에 매우 유용하게 사용될 수 있을 것이다.

T-DNA-mediated transformation is a common method for generating transgenic plants with insertional mutagenesis. In order to identify important genes involved in shoot development, a system of promoter trap insertional mutagenesis was employed in Arabidopsis thaliana. For this system, an efficient promoter trap vector, pFGL561 was developed. The pFGL561 includes a basta-resistant gene, an intron with multiple splicing donor and acceptor sites, and a promoter-less GFP reporter gene. Using floral-dipping method, we made total 300 $T_1$ promoter-trap lines which were screened for GFP expression. GFP signals in the $T_1$ plants were detected with high frequency, 26.7%, and the signals were reconfirmed in $T_2$ plants. To isolate the genes that are involved in shoot development, phenotypes were analyzed in $T_2$ plants of the 19 $T_1$ lines that had GFP signals in shoot apex, and 6 $T_1$ lines were selected that had abnormal shoot development. These lines will be very useful for the investigation of shoot development.

키워드

참고문헌

  1. Azpiroz-Leehan, R. and K. A. Feldmann. 1997. T-DNA insertion mutagenesis in Arabidopsis: Going back and forth. Trends Genet. 13, 152-156 https://doi.org/10.1016/S0168-9525(97)01094-9
  2. Bouchez, D., C. Camilleri and M. Caboche. 1993. A binary vector based on Basta resistance for in planta transformation of Arabidopsis thaliana. Methods Mol. Biol. 316, 1188-1193
  3. Campisi, L., Y. Yang, Y. Yi, E. Heilig, B. Herman, A. J. Cassista, D. W. Allen, H. Xiang and T. Jack. 1999. Generation of enhancer trap lines in Arabidopsis and characterization of expression patterns in the inflorescence. Plant J. 17, 699-670 https://doi.org/10.1046/j.1365-313X.1999.00409.x
  4. Casson, S. A., P. Chilley, J. E. Topping, I. M. Evans, M. Souter and K. Lindsey. 2002. The POLARIS gene of Arabidopsis encodes a predicted peptide required for correct root growth and leaf vascular patterning. Plant Cell 14, 1705-1721 https://doi.org/10.1105/tpc.002618
  5. Chen, H., R. S. Nelson and J. L. Sherwood. 1994. Enhanced recovery of transformants of Agrobacterium tumefaciens after freeze-thaw transformation and drug selection. Biotechniques 16, 664-668
  6. Chin, H. G., M. S. Choe, S. H. Lee, S. H. Park, J. C. Koo, N. Y. Kim, J. J. Lee, B. G. Oh, G. H. Yi, S. C. Kim, H. C. Choi, M. J. Cho and C. D. Han. 1999. Molecular analysis of rice plants harboring an Ac/Ds transposable element- mediated gene trapping system. Plant J. 19, 615-23 https://doi.org/10.1046/j.1365-313X.1999.00561.x
  7. Clough, S. J. and A. F. Bent. 1998. Floral dip: a simplified method for agrobacterium-mediated transformation of Arabidopsis thaliana. Plant J. 16, 735-43 https://doi.org/10.1046/j.1365-313x.1998.00343.x
  8. David, W., J. Meinke, C., Michael, D., Caroline, D., Stevenson, Rounsley and K., Maarten. 1998. Arabidopsis thaliana : A model plant for genome analysis. Science 282, 622-682
  9. Feldmann, K. A. and M. D. Marks. 1987. Agrobacteriummediated transformation of germinating seeds of Arabidopsis thaliana: A non-tissue culture approach. Mol. Gen. Genet. 208, 1-9 https://doi.org/10.1007/BF00330414
  10. Jeon, J. S., S. Lee, K. H. Jung, S. H. Jun, D. H. Jeong, J. Lee, C. Kim, C. S. Jang, K. Yang, J. Nam, K. An, M. J. Han, R. J. Sung, H. S. Choi, J. H. Yu, J. H. Choi, S. Y. Cho, S. S. Cha, S. I. Kim and G. An. 2000. T-DNA insertional mutagenesis for functional genomics in rice. Plant J. 22, 561-70 https://doi.org/10.1046/j.1365-313x.2000.00767.x
  11. Krysan, P. J., J. C. Young and M. R. Sussman. 1999. T-DNA as an insertional mutagen in Arabidopsis. Plant Cell 11, 2283-2290 https://doi.org/10.1105/tpc.11.12.2283
  12. Lindsey, K., W. Wei, M. C. Clarke, H. F. McArdle, L. M. Rooke and J. F. Topping. 1993. Tagging genomic sequences that direct transgene expression by activation of a promoter trap in plants. Transgenic Res. 2, 33-47 https://doi.org/10.1007/BF01977679
  13. Lindsey, K., J. F. Topping, P. R. Muskett, W. Wei and K. L. Home. 1998. Dissecting embryonic and seedling morphogenesis in Arabidopsis by promoter trap insertional mutagenesis. Symp. Soc. Exp. Biol. 51, 1-10
  14. Martha, C., Alvarado, M. Laura, Zsigmond, K. Izabella, C. Agnes, K. Csaba and M. S. Laszlo. 2004. Gene trapping with firefly luciferase in Arabidopsis. Tagging of stress-responsive genes. Plant Physiology 134, 18-27 https://doi.org/10.1104/pp.103.027151
  15. McKinney, E. C., N. Ali, A. Traut, K. A. Feldmann, D. A. Belostotsky, J. M. McDowell and R. B. Meagher. 1995. Sequencebased identification of T-DNA insertion mutations in Arabidopsis: Actin mutants act2-1 and act4-1. Plant J. 8, 613-622 https://doi.org/10.1046/j.1365-313X.1995.8040613.x
  16. Murashige, T and F. Skoog. 1962. A revised medium for rapid growth and bioassays with tobacco tissue culture. Physiol Plant 15, 473-497 https://doi.org/10.1111/j.1399-3054.1962.tb08052.x
  17. Springer, P. S. 2000. Gene traps: tools for plant development and genomics. Plant Cell 12, 1007-1020
  18. Sundaresan, V., P. Springer, T. Volpe, S. Haward, J. D. Jones, C. Dean, H. Ma and R. Martienssen. 1995. Patterns of gene action in plant development revealed by enhancer trap and gene trap transposable elements. Genes Dev. 9, 1797-1810 https://doi.org/10.1101/gad.9.14.1797
  19. Topping, J. F., W. Wei and K. Lindsey. 1991. Functional tagging of regulatory elements in the plant genome. Development 112, 1009-1019
  20. Topping, J. F., F. Agyeman, B. Henricot and K. Lindsey. 1994. Identification of molecular markers of embryogenesis in Arabidopsis thaliana by promoter trapping. Plant J. 5, 895-903 https://doi.org/10.1046/j.1365-313X.1994.5060895.x
  21. Wei, W., D. Twell and K. Lindsey. 1997. A novel nucleic acid helicase gene identified by promoter trapping in Arabidopsis. Plant J. 11, 1307-1314 https://doi.org/10.1046/j.1365-313X.1997.11061307.x
  22. Weigel, D. 2000. Activation tagging in Arabidopsis. Plant Physiol. 122, 1003-1014 https://doi.org/10.1104/pp.122.4.1003
  23. Winkler, R. G., M. R. Frank, D. W. Galbraith, R. Feyereisen and K. A. Feldmann. 1998. Systematic reverse genetics of transfer-DNA-tagged lines of Arabidopsis: Isolation of mutations in the cytochrome P450 gene superfamily. Plant Physiol. 118, 743-750 https://doi.org/10.1104/pp.118.3.743