Agrobacterium Tumefaciens Based Transformation of Pelargonium x Hortrum cv. ‘Samba’ with Anti-1-aminocyclopropane-1-carboxylate Synthase cDNA to Regulate Ethylene Biosynthesis

DOI: 10.4236/eng.2012.410B002   PDF   HTML     2,847 Downloads   4,181 Views   Citations

Abstract

Phytohormone, ethylene plays an important role in plant growth and development including fruit ripening and flower senescence. The synthesis of 1-aminocylo-propane-1- carboxylate (ACC), the immediate precursor of ethylene, from S-adenosyl-methionine is catalyzed by ACC synthase; and which is also a rate limiting step in the ethylene biosynthetic pathway. Therefore, it plays a key role in ethylene biosynthesis and the genes that code for ACC synthase are of special interest. Moreover, in zonal geraniums, ethylene bursts released from cuttings can have profound impact on the viability of explants for plant propagation. Biotechnological approach involving genetic modification that may reduce ethylene levels has potential for increasing the shelf-life of cuttings for plant propagation. These considerations have led us to clone several cDNA of ACC synthase genes from Pelargonium x hortorum cv.  ‘Sincerity’. To transform geranium cells with Agrobacterium tumefaciens an in vitro regeneration  system was developed using very young petiole explants. An Antisense construct of ACC synthase cDNA (PHSacc41) ligated into binary vector pAM696 was introduced into A. tumefaciens EHA 105 cells. Petiole explants were incubated with the Agrobacterium for 15 min and then co-cultivated for several days on MS medium containing 5 mM BAP and  1 mM IAA in  the dark without the antibiotics. Selection for transformants was carried out in the presence of kanamycin and timentin. Transgenic plantlets generated were examined for inserted gene cassette by PCR and Southern blotting. Recovery of positive transformants that survived selection suggested that it is possible to transform and introduce genes via transformation in hybrid geraniums for genetic modification.

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R. Ranu, J. Fan, S. Krishnan, P. Agarwal. and A. Mitra, "Agrobacterium Tumefaciens Based Transformation of Pelargonium x Hortrum cv. ‘Samba’ with Anti-1-aminocyclopropane-1-carboxylate Synthase cDNA to Regulate Ethylene Biosynthesis," Engineering, Vol. 4 No. 10B, 2012, pp. 4-7. doi: 10.4236/eng.2012.410B002.

Conflicts of Interest

The authors declare no conflicts of interest.

References

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[21] Taylor, B.H., Manhart, J.R. & Amasino, R.R. (1993) Isolation and characterization of plant DNA. In: Glick, B.R. & Thompson, C.R. (eds) Methods in plant molecular biology and biotechnology. CRC Press, Boca Raton, pp 37-38.
[22] Agarwal, P. Ranu, RS.(2000) Regenration of plantlets from leaf and petiole explants of Pelargonium x hortorum. In viro CellDev Biol-Plant 36,392-396
[23] Boase, M.R., Deroles, S.C., Winefield, C.S., Butcher, S.M., Borst, N.K. & Butler, R.C. (1996) Genetic transformation of regal pelargoinium (Pelargonium x domesticum Dubonnet) by Agrobacterium tumefaciens. Plant Sci. 121:47-61. Krishnaraj, S., Bi, Y.-M. & Saxena, P.K. (1997) Somatic embryogenesis and Agrobacterium-mediated transformation system from scented geranium (Pelargonium sp. Frensham). Planta. 201:434-440.
[24] Pellegrineschi, A., Damon, J.P., Valtorta, N., Paillard, N. & Tepfer, D. (1994) Improvement of ornamental characters and fragrance production in lemon scented geranium through genetic transformation by Agrobacterium rhizogenes. Biotech. 12:64-68.
[25] White, J. (1993) Geranium, IV. Geneva, IL: Ball Publishing.
[26] Wood, H.J. (1966) Pelargoniums: a complete guide to their cultivation. UK: Wheaton & Co.
[27] Moore, H.E. Plant classification 3: Taxonomy in cultivation .In: Mastalerz, J.M., ed. Geraniums, a Penn State manual.University Park, Pennsylvania: Pennsylvania Flower Growers. 1971:14-15.
[28] Horn, W. (1994) Interspecific crossability and inheritance in Pelargonium. Plant Breed. 113:3-17.
[29] Mattoo AK, Suttle JC (1991) Plant Hormone Ethylene,CRCPress, Boca Raton, FL, 352 pp
[30] Yang, S.F. & Hoffman, N.E.(1984) Ethylene biosynthesis andits regulation in higher plants. Annu. Rev. Plant Physiol. 35:155-189.
[31] White, J. (1993) Geranium, IV. Geneva, IL: Ball Publishing.
[32] Wood, H.J. (1966) Pelargoniums: a complete guide to their cultivation. UK: Wheaton & Co.
[33] Moore, H.E. Plant classification 3: Taxonomy in cultivation .In: Mastalerz, J.M., ed. Geraniums, a Penn State manual.University Park, Pennsylvania: Pennsylvania Flower Growers. 1971:14-15.
[34] Horn, W. (1994) Interspecific crossability and inheritance in Pelargonium. Plant Breed. 113:3-17.
[35] Mattoo AK, Suttle JC (1991) Plant Hormone Ethy-lene,CRCPress, Boca Raton, FL, 352 pp
[36] Yang, S.F. & Hoffman, N.E.(1984) Ethylene biosynthesis andits regulation in higher plants. Annu. Rev. Plant Physiol. 35:155-189.
[37] Ranu, RS., Wang D. Fan J. Role of 1-aminocyclopropane-1-caboxylate (ACC) synthase genes and genes involved in ethylene signal transduction and rose flower senescence. Floriculture and Ornamenral Biotechnology Special issue on roses. 3, 104-110, 2009.
[38] Ranu RS. Chakrabarti, S. Lysa-Anne M. Volpe L-A. M., Brown B., Wang D., Fan, F. 1-Aminocyclopropane-1-carboxylate (ACC) Synthases of Rosa hybrida: Analysis of Genomic Gene Structure and the Cis-Acting Regulatory Elements in their Promoters. Genes, Genomes and Genetics. 2, 68-76, 2008
[39] Ma N, Cai L, Lu WJ, Tan H, Gao JP (2005) Exogenous ethylene influences flower opening of cut roses (Rosa hybrida) by regulating the genes encoding ethylene biosynthesis enzymes. Science in China, Series C 48, 434-444
[40] Ma N, Tan H, Liu X, Li Y, Gao J (2006) Transcriptional regulation of ethylene receptor and CTR genes involved in ethylene-induced flower opening in cut rose (Rosa hybrida) cv. Samantha. Journal of Experimental Botany 57, 2763-2773
[41] An, G., Ebert, P.R., Mitra, A. & Ha, S.B. (1988) Binary vectors in Plant Molecular Biology Manual, Ed, Klawcr Academic Publishers, Dordrect, Belgium A3, pp. 1-19.
[42] Mitra, A. & Higgins, D.W. (1994) The Chlorella virus adenine methyltransferase gene promoter is a strong promoter in plants. Plant Mol. Biol. 26:85-93
[43] Fan, J., Wang, D. & Ranu, R.S. (1996) Cloning and characterization of cDNA encoding 1-aminocyclopropane-1-carboxylate synthases from Pelargonium x hortorum cv. Sincerity. FASEB Journal,10:A1410
[44] Fan J, Wang D, Ranu RS (2007) Characterization and expression of 1-aminocyclopropane-1-carboxylate(ACC) synthase from geranium (Pelargonium x hortorum cv Sincerity). Journal of Plant Biochemistry and Biotechnology 16, 87-96
[45] Murashige, T. & Shook, F. (1962) A revised medium for rapid growth and bioassays with tobacco tissue cultures. Physiol. Plant. 15:473-497.
[46] Taylor, B.H., Manhart, J.R. & Amasino, R.R. (1993) Isolation and characterization of plant DNA. In: Glick, B.R. & Thompson, C.R. (eds) Methods in plant molecular biology and biotechnology. CRC Press, Boca Raton, pp 37-38.
[47] Agarwal, P. Ranu, RS.(2000) Regenration of plantlets from leaf and petiole explants of Pelargonium x hortorum. In viro CellDev Biol-Plant 36,392-396
[48] Boase, M.R., Deroles, S.C., Winefield, C.S., Butcher, S.M., Borst, N.K. & Butler, R.C. (1996) Genetic transformation of regal pelargoinium (Pelargonium x domesticum Dubonnet) by Agrobacterium tumefaciens. Plant Sci. 121:47-61.
[49] Krishnaraj, S., Bi, Y.-M. & Saxena, P.K. (1997) Somatic embryogenesis and Agrobacterium-mediated transformation system from scented geranium (Pelargonium sp. Frensham). Planta. 201:434-440.
[50] Pellegrineschi, A., Damon, J.P., Valtorta, N., Paillard, N. & Tepfer, D. (1994) Improvement of ornamental characters and fragrance production in lemon scented geranium through genetic transformation by Agrobacterium rhizogenes. Biotech. 12:64-68.

  
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