In planta Transformation of Wheat Apical Meristem:A Preliminary Study

doi:10.3321/j.issn:1000-7091.2005.01.004

Abstract

Abstract: Realizing the importance of in planta transformation for wheat to obviate the inconvenience of callus based transformation through Agrobacterium and particle bombardment an attempt was made for direct transformation of apical meristem. The study provides a positive clue for success of in planta transformation of wheat apical meristem. Coleoptiles of growing seedlings were cut and exposed meristems were wounded with fiberglass brush and inoculated with Agrobacterium.Apical meristems of 1一11 days old seedlings were inocufated with Agrobacterium tumefaciens LBA4404 containing hph-GUS genes in its plasmid and transient GUS expression was determined in regenerated shoots one week after co-cultivation. When apical meristems from 1一2 days old seedlings were inoculated then 2 to 3 florets from inflorescences were tested for stable GUS expression. One-day-old seedlings were inoculated with Agrobacterium tumefaciens AGL1 harboring nptII and BADH genes in its plasmid and regenerated seedlings were selected on 100 mg/L kanamycin in vermiculite. Results indicate that transient GUS frequency decreased with increasing age of the seedlings with maximum(10.7%)in two days old seedlings. Numbers of GUS positive inflorescences were more when apical meristems from one-day-old seedlings were inoculated compared with 2 days old seedlings. Presence of glucose did not enhance the frequency of GUS expression in inflorescences. PCR test did not confirm the presence of GUS transgene in the progeny of plants with GUS positive inflorescences. Sixty-three kanamycin resistant green plants(13.6%)were obtained but only three plants were found PCR positive. Only one plant produced seeds.

Key words: Wheat, In planta, Apical meristem, Transformation, GUS, nptII-BADH

CLC Number:

S513

RAZZAQ Abdul, ZHANG Yan-min, YANG Fan, YOU Yan-jie, ZHAO He, MA Zhi-ying, WANG Hai-bo. In planta Transformation of Wheat Apical Meristem:A Preliminary Study[J]. ACTA AGRICULTURAE BOREALI-SINICA, 2005, 20(1): 17-22. doi: 10.3321/j.issn:1000-7091.2005.01.004.

/ / Recommend / Download Citations

URL: http://www.hbnxb.net/EN/10.3321/j.issn:1000-7091.2005.01.004

http://www.hbnxb.net/EN/Y2005/V20/I1/17

References

[1] Birch R G. Plant transformation:problems and strategies for practical application [J]. Ann Rev Plant Physiol Plant Mol Biol, 1997, 48:297- 326.
[2] Karp A. Origins causes and uses of variation in plant tissue cultures [A]. In:Vasil I K, Thorpe T A. Plant Cell and Tissue Culture [M]. Dordrecht, The Netherlands:Kluwer Academic Publishers, 1994.139- 151.
[3] Chee P P, Slighton J L. Transformation of soybean (Glycine max) via Agrobacterium tumefaciens and analysis of transformed plants [A]. In:KMA Gartland,Davey M Reds. Agrobacterium Protocols:Methods in Molecular Biology [M]. Totowa, NJ:Humana Press,1995. 101- 119.
[4] Christou P Ford, T L Kofron M. The development of a variety independent gene-transfer method for rice [J].Trends Biotechnol, 1992, 10:239- 247.
[5] Lorence A, Verpoorte R. Gene transfer and expression in plants [J]. Methods Mol Biol,2004, 267:329-50.
[6] Bent A F. Arabidopsis in planta transformation:Uses,mechanisms, and prospects for transformation of other species[J]. Plant Physiol,2000,124:1540- 1547.
[7] Graves A C F, Goldman S L. The transformation of Zea mays seedlings with Agrobacterium tumefaciens-detection of T-DNA specific enzyme activities[J]. Plant Mol Biol,1986, 7:43- 50.
[8] Gould J, Banister S, Fahima M, et al. Regeneration of Gossypium hirsutum. and G. biarbadense from shoot apex[J]. Plant Cell Rep,1991a,10:12- 16.
[9] Bechtold N, Ellis J, Pelletier G. In planta Agrobacterium mediated gene transfer by infiltration of adult Arabidopsis plants[M]. C R Acad Sci Paris, Life Sciences,1993, 316:1194- 1199.
[10] Katavic V, Haughn G W, Reed D, et al. In planta transformation of Arabidopsis thaliana [J]. Mol Gen Genet,1994, 245(3):363- 370.
[11] Feldmann K A, Marks M D. Agrobacterium-mediated transformation of germinating seeds of Arabidopsis thaliana:a non-tissue culture approach [J]. Mol Gen Genet, 1987,208:1- 9.
[12] Rosegrant M W, Sombilla M A, Gerpacio R V,et al.Global food markets and US exports in 21 st century [A]. Paper presented at the Illinois World Food and Sustainable Agriculture Program Conference. Meeting the Demand for Food in 21st century:Challenges and Opportunities, 28 th May[C], Univ Illionois, UrbanaCahmopaign, USA,1997.
[13] Vasil I K, Reway F A, Vasil I K. Regeneration of plants from embryogenic suspension culture protoplasts of wheat [J]. Bio Tech, 1990,8:429- 433.
[14] Vasil V, Castillo A M, Fromm M E, et al. Herbicide resistant fertile transgenic wheat plants obtained by microprojectile bombardment of regenerable embryogenic callus [J]. Bio Tech,1992, 10:667-674.
[15] Ahokas H. Transfection of germinating barley seed electrophoretically with exogenous DNA [J]. Theor Appl Genet, 1989, 77:469- 472.
[16] Oard J H, Paige D F, Simmonsand J A, et al. Transient gene expression in maize, rice and wheat cells using an air gun apparatus [J]. Plant Physiol, 1990, 92:334- 339.
[17] Chen D F, Dale P J. A comparison of methods for delivering DNA to wheat:the application of wheat dwarf virus DNA to seeds with exposed apical meristems [J].Transgenic Research, 1992,1:93- 100.
[18] Simmonds J, Stewart P, Simmonds D. Regeneration of Triticum aestivum apical explants after microinjection of germ line progenitor cells with DNA [J]. Phsiologia Plantarum, 1992, 85:197- 206.
[19] Iglesias V A, Gisel A, Bilang R, et al. Transient expression of visible marker genes in meristem cells of wheat embryos after biloistic micro-targeting [J]. Planta, 1994, 192:84-91.
[20] Potrykus I. Gene transfer to cereals:an assessment[J].Bio/Biotechnology, 1990, 8:535- 542.
[21] Jefferson R. Assaying chimeric genes in plants:The GUS gene fusion system [J]. Plant Molecular Biology Reporter, 1987, 5:387- 405.
[22] Doyle J J, Doyle J I. Isolation of plant DNA from fresh tissues [J].Focus, 1990, 12:13-15.
[23] Wrigt M S, Koechler S M, Hinchee M A,et al. Plant regeneration by organogenesis in Glycin max [J]. Plant Cell Rep, 1986, 5:150- 154.
[24] Stachel S E, Messens E, Van Montagu M, et al. Identification of the signal molecules produced by wounded plant cells that activate T-DNA transfer in Agrobacterium tumefaciens [J]. Nature, 1985, 318:624-629.
[25 ]Seo M S, Bae C H, Choi D O, et al. Investigation of transformation efficiency of rice using Agrobacterium tumefaciens and high transformation of GPAT (glycerol-3- phosphate aceyltransferase) gene relative to chilling tolerance [J]. Korean J Plant Biotechnology, 2002,29:85- 92.
[26] Shimoda N,Toyoda- Yamamoto A, Nagamine J, et al.Control of expression of Agrobacterium vir genes by synergistic actions of phenolic signals molecules and monosaccharides [C]. Proc Natl Acad Sci, 1990, 87:6684- 6688.
[27] Hiei Y, Ohta S, Komari T,et al. Efficient transformation of rice (Oryza sativa) mediated by Agrobacterium and sequence analysis of the boundaries of the T-DNA [J]. Plant J, 1994,72, 233- 244.
[28] Hauptmann R M, Vasil V, Ozia-Akin P, et al. Evaluation of selectable markers for obtaining stable transformants in Gramineae [J]. Plant Physiol, 1988, 85:602- 606.

Please choose a citation manager

Content to export