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Production of transgenic pineapple (Ananas cosmos (L.) Merr.) plants via adventitious bud regeneration

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Abstract

A new protocol for the production of transgenic pineapple plants was developed. Adventitious buds were induced directly from Agrobacterium-infected leaf bases and stem discs of in vitro plants, bypassing the establishment of callus cultures. Non-chimeric transgenic plants were obtained by multiple subculturing of primary transformants under increasing levels of selection. A total of 42 independent transgenic lines were produced from two cultivars with two different constructs: one containing a modified rice cystatin gene (Oc-IΔD86) and the other with the anti-sense gene to pineapple aminocyclopropane synthase (ACS). GUS histochemical staining provided the first evidence of the non-chimeric nature of the transformed plants. Their non-chimeric nature was further demonstrated by PCR analyses of the DNA extracted from individual leaves of a primary transformed plant and also from multiple plants propagated from a single transformation event. Southern hybridization confirmed random integration patterns of transgenes in the independent lines. For the Oc-IΔD86 gene, the expression at the mRNA level was detected via RT-PCR and its translation was detected by protein blot. Agronomic evaluation and bioassays of the transgenic plants will further validate the utility of this new tool for pineapple improvement.

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References

  • Agarwal, S.; Kanwar, K. Comparison of genetic transformation in Morus alba L. via different regeneration systems. Plant Cell Rep. 26: 177–185; 2007. doi:10.1007/s00299-006-0217-3.

    Article  PubMed  CAS  Google Scholar 

  • Agarwal, S.; Kanwar, K.; Saini, N.; Jain, R. K. Agrobacterium tumefaciens mediated genetic transformation and regeneration of Morus alba L. Sci. Hortic. 100: 183–191; 2004. doi:10.1016/j.scienta.2003.06.002.

    Article  CAS  Google Scholar 

  • Almeida, W. A. B.; Mourao, Filho, F. A. A.; Pino, L. E.; Boscariol, R. L.; Rodriguez, A. P. M.; Mendes, B. M. J. Genetic transformation and plant recovery from mature tissues of Citrus sinensis L. Osbeck. Plant Sci. 164: 203–211; 2003. doi:10.1016/S0168-9452(02)00401-6.

    Article  CAS  Google Scholar 

  • Bugos, R. C.; Chiang, V. L.; Zhang, X. H.; Campbell, E. R.; Podila, G. K.; Campbell, W. H. RNA isolation from plant tissues recalcitrant to extraction in guanidine. Biotechniques. 19: 734–737; 1995.

    PubMed  CAS  Google Scholar 

  • Chan, Y. K.; Coppens, G.; d’Eeckenbrugge, G. C.; Sanewski, G. M. Breeding and variety improvement. In: Bartholomew D. P.; Paull R. E.; Rohrbach K. G. (eds) The pineapple: botany, production and uses. CABI, Honolulu, pp 33–55; 2002.

    Google Scholar 

  • Cui, M. L.; Ezura, H.; Nishimura, S.; Kamada, H.; Handa, T. A rapid Agrobacterium-mediated transformation of Antirrhinum majus L. by using direct shoot regeneration from hypocotyl explant. Plant Sci 166: 873–879; 2004. doi:10.1016/j.plantsci.2003.11.021.

    Article  CAS  Google Scholar 

  • Espinosa, P.; Lorenzo, J.; Iglesias, A.; Yabor, L.; Menéndez, E.; Borroto, J.; Hernández, L.; Arencibia, A. Production of pineapple transgenic plants assisted by temporary immersion bioreactors. Plant Cell Rep. 21: 136–140; 2002. doi:10.1007/s00299-002-0481-9.

    Article  CAS  Google Scholar 

  • FAOSTAT, (2008) http://faostat.fao.org/site/567/DesktopDefault.aspx?PageID = 567#ancor. In:

  • Firoozabady, E.; Heckert, M.; Gutterson, N. Transformation and regeneration of pineapple. Plant Cell, Tissue Organ Cult. 84: 1–16; 2006. doi:10.1007/s11240-005-1371-y.

    Article  Google Scholar 

  • Firoozabady, E.; Moy, Y. Regeneration of pineapple plants via somatic embryogenesis and organogenesis. In Vitro Cell Dev. Biol. 40: 67–74; 2004. doi:10.1290/1543-706X(2004)040<0067:UAPTOA>2.0.CO;2.

    Article  Google Scholar 

  • Graham, M.; Smith, M. K.; Hardy, V. G.; Ko, H. L. A method of plant transformation. International Patent WO 01/33943 A33941; 2001.

  • He, X.; Miyasaka, S. C.; Fitch, M. M.; Moore, P. H.; Zhu, Y. J. Agrobacterium tumefaciens-mediated transformation of taro (Colocasia esculenta (L.) Schott) with a rice chitinase gene for improved tolerance to a fungal pathogen Sclerotium rolfsii. Plant Cell Rep. 27: 903–909; 2008. doi:10.1007/s00299-008-0519-8.

    Article  PubMed  CAS  Google Scholar 

  • Jefferson, R. A. Assaying chimeric genes in plants: The GUS gene fusion system. Plant. Mol. Biol. Report. 5: 387–405; 1987. doi:10.1007/BF02667740.

    Article  CAS  Google Scholar 

  • Kato, C. Y.; Nagai, C.; Moore, P. H.; Zee, F.; Kim, M. S.; Steiger, D. L.; Ming, R. Intra-specific DNA polymorphism in pineapple (Ananas comosus (L.) Merr.) assessed by AFLP markers. Genet. Resour. Crop Evol. 51: 815–825; 2004. doi:10.1007/s10722-005-0005-x.

    Article  CAS  Google Scholar 

  • Ko, H. L.; Campbell, P. R.; Jobin-D’ecor, M. P.; Eccleston, K. L.; Graham, M. W.; Smith, M. K. The introduction of transgenes to control blackheart in pineapple (Ananas comosus L.) cv. Smooth Cayenne by microprojectile bombardment. Euphytica. 150: 387–395; 2006. doi:10.1007/s10681-006-9124-5.

    Article  CAS  Google Scholar 

  • Lakshmanan, P.; Geijskes, R. J.; Wang, L.; Elliott, A.; Grof, C. P.; Berding, N.; Smith, G. R. Developmental and hormonal regulation of direct shoot organogenesis and somatic embryogenesis in sugarcane (Saccharum spp. interspecific hybrids) leaf culture. Plant Cell Rep. 25: 1007–1015; 2006. doi:10.1007/s00299-006-0154-1.

    Article  PubMed  CAS  Google Scholar 

  • Lazo, G. R.; Stein, P. A.; Ludwig, R. A. A DNA transformation-competent Arabidopsis genomic library in Agrobacterium. Biotechnology (N Y). 9: 963–967; 1991. doi:10.1038/nbt1091-963.

    Article  CAS  Google Scholar 

  • Lin, R. C.; Ding, Z. S.; Li, L. B.; Kuang, T. Y. A rapid and efficient DNA minipreparation suitable for screening transgenic plants. Plant. Mol. Biol. Report. 19: 379a–379e; 2001.

    Article  CAS  Google Scholar 

  • Mathews, H.; Dewey, V.; Wagoner, W.; Bestwick, R. K. Molecular and cellular evidence of chimaeric tissues in primary transgenics and elimination of chimaerism through improved selection protocols. Transgenic. Res. 7: 123–129; 1998.

    Article  CAS  Google Scholar 

  • May, G. D.; Afza, R.; Mason, H. S.; Wiecko, A.; Novak, F. J.; Arntzen, C. J. Generation of transgenic banana (Musa acuminata) plants via Agrobacterium-mediated transformation. Bio/technology 13: 486–492; 1995. doi:10.1038/nbt0595-486.

    Article  CAS  Google Scholar 

  • Mezzetti, B.; Costantini, E. Strawberry (Fragaria x ananassa). Methods Mol. Biol. 344: 287–295; 2006.

    PubMed  CAS  Google Scholar 

  • Mezzetti, B.; Pandolfini, T.; Navacchi, O.; Landi, L. Genetic transformation of Vitis vinifera via organogenesis. BMC Biotechnol. 2: 18; 2002. doi:10.1186/1472-6750-2-18.

    Article  PubMed  Google Scholar 

  • Nagai, C.; Rosal, L. Pineapple micropropagation. Hawaii Sugar Planters’ Association Annual Report 33–34; 1995.

  • Nan, G. L.; Nagai, C.; Sipes, B. Field evaluation of tissue culture-derived pineapple plant at pre-flowering stage. Hawaii Agriculture Research Center Annual Report 24–25; 1997.

  • Nontaswatsri, C.; Fukai, S. Carnation (Dianthus caryophylus L.). Methods Mol. Biol. 344: 311–320; 2006.

    PubMed  CAS  Google Scholar 

  • Rangan, T. S. Pineapple. In: Ammirato P. V.; Evans D. A.; Sharp W. R.; Yamada Y. (eds) Handbook of plant cell culture. Macmillan, New York, pp 373–536; 1984.

    Google Scholar 

  • Rohrbach, K. G.; Leal, F.; d’Eeckenbrugge, G. C. History, distribution and world production. In: Bartholomew D. P.; Paull R. E.; Rohrbach K. G. (eds) The pineapple: botany, production and uses. CABI, Honolulu, pp 1–12; 2002.

    Google Scholar 

  • Sharma, K. K.; Anjaiah, V. V. An efficient method for the production of transgenic plants of peanut (Arachis hypogaea L.) through Agrobacterium tumefaciens-mediated genetic transformation. Plant Sci. 159: 7–19; 2000. doi:10.1016/S0168-9452(00)00294-6.

    Article  PubMed  CAS  Google Scholar 

  • Sharma, K. K.; Bhatnagar-Mathur, P. Peanut (Arachis hypogaea L.). Methods Mol. Biol. 343: 347–358; 2006.

    PubMed  Google Scholar 

  • Song, G. Q.; Sink, K. C. Optimizing shoot regeneration and transient expression factors for Agrobacterium tumefaciens transformation of sour cherry (Prunus cerasus L.) cultivar Montmorency. Sci. Hortic. 106: 60–69; 2005. doi:10.1016/j.scienta.2005.02.018.

    Article  CAS  Google Scholar 

  • Song, G. Q.; Sink, K. C. Transformation of Montmorency sour cherry (Prunus cerasus L.) and Gisela 6 (P. cerasus x P. canescens) cherry rootstock mediated by Agrobacterium tumefaciens. Plant Cell Rep. 25: 117–123; 2006. doi:10.1007/s00299-005-0038-9.

    Article  PubMed  CAS  Google Scholar 

  • Sripaoraya, S.; Marchant, R.; Power, J. B.; Davey, M. R. Herbicide-tolerant transgenic pineapple (Ananas comosus) produced by microprojectile bombardment. Ann. Bot. 88: 597–603; 2001. doi:10.1006/anbo.2001.1502.

    Article  CAS  Google Scholar 

  • Sun, H. J.; Uchii, S.; Watanabe, S.; Ezura, H. A highly efficient transformation protocol for Micro-Tom, a model cultivar for tomato functional genomics. Plant Cell Physiol. 47: 426–431; 2006. doi:10.1093/pcp/pci251.

    Article  PubMed  CAS  Google Scholar 

  • Syamala, D.; Devi, P. Efficient regeneration of sorghum, Sorghum bicolor (L.) Moench, from shoot-tip explant. Indian J. Exp. Biol. 41: 1482–1486; 2003.

    PubMed  CAS  Google Scholar 

  • Terakami, S.; Matsuta, N.; Yamamoto, T.; Sugaya, S.; Gemma, H.; Soejima, J. Agrobacterium-mediated transformation of the dwarf pomegranate (Punica granatum L. var. nana). Plant Cell Rep. 26: 1243–1251; 2007. doi:10.1007/s00299-007-0347-2.

    Article  PubMed  CAS  Google Scholar 

  • Tournier, V.; Grat, S.; Marque, C.; El Kayal, W.; Penchel, R.; de Andrade, G.; Boudet, A. M.; Teulieres, C. An efficient procedure to stably introduce genes into an economically important pulp tree (Eucalyptus grandis x Eucalyptus urophylla). Transgenic Res. 12: 403–411; 2003. doi:10.1023/A:1024217910354.

    Article  PubMed  CAS  Google Scholar 

  • Trusov, Y.; Botella, J. R. Silencing of the ACC synthase gene ACACS2 causes delayed flowering in pineapple [Ananas comosus (L.) Merr.]. J. Exp. Bot. 57: 3953–3960; 2006. doi:10.1093/jxb/erl167.

    Article  PubMed  CAS  Google Scholar 

  • Urwin, P. E.; Atkinson, H. J.; Waller, D. A.; McPherson, M. J. Engineered oryzacystatin-I expressed in transgenic hairy roots confers resistance to Globodera pallida. Plant J. 8: 121–131; 1995. doi:10.1046/j.1365-313X.1995.08010121.x.

    Article  PubMed  CAS  Google Scholar 

  • Urwin, P. E.; Green, J.; Atkinson, H. J. Expression of a plant cystatin confers partial resistance to Globodera, full resistance is achieved by pyramiding a cystatin with natural resistance. Mol Breed. 12: 263–269; 2003. doi:10.1023/A:1026352620308.

    Article  CAS  Google Scholar 

  • Urwin, P. E.; Lilley, C. J.; McPherson, M. J.; Atkinson, H. J. Resistance to both cyst and root-knot nematodes conferred by transgenic Arabidopsis expressing a modified plant cystatin. Plant J. 12: 455–461; 1997. doi:10.1046/j.1365-313X.1997.12020455.x.

    Article  PubMed  CAS  Google Scholar 

  • Wakasa, K. Variation in the plants differentiated from the tissue culture of pineapple. Japan. J. Breed. 29: 13–22; 1979.

    Google Scholar 

  • Wakasa, K. Pineapples. In: Bajaja Y. P. S. (ed) Biotechnology in agriculture and forestry 5: trees II. Springer, Berlin, pp 13–22; 1989.

    Google Scholar 

  • Wang, G.; Xu, Y. Hypocotyl-based Agrobacterium-mediated transformation of soybean (Glycine max) and application for RNA interference. Plant Cell Rep. 27: 1177–1184; 2008. doi:10.1007/s00299-008-0535-8.

    Article  PubMed  Google Scholar 

  • Wei, H.; Wang, M. L.; Moore, P. H.; Albert, H. H. Comparative expression analysis of two sugarcane polyubiquitin promoters and flanking sequences in transgenic plants. J. Plant Physiol. 160: 1241–1251; 2003. doi:10.1078/0176-1617-01086.

    Article  PubMed  CAS  Google Scholar 

  • Yabor, L.; Espinosa, P.; Arencibia, A. D.; Lorenzo, J. C. Pineapple [Ananas comosus (L.) Merr.]. Methods Mol. Biol. 344: 219–226; 2006.

    PubMed  CAS  Google Scholar 

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Acknowledgements

We thank Mrs. Josephine Buenafe for technical support and Dr. Eden A. Perez for bringing the subculturing procedure to our attention. This project was funded by USDA ARS Specific Cooperative Agreements to the College of Tropical Agriculture and Human Resources entitled “Molecular Improvement of Pineapple” (59-5320-0-170) and “Hawaii Pineapple Improvement” (58-5320-5-785).

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Author notes

  1. G.-L. Nan

    Present address: Biological Sciences, Stanford University, 385 Serra Mall, Stanford, CA, 94305-5020, USA

Authors and Affiliations

  1. Hawaii Agriculture Research Center, P.O. Box 100, Kunia, HI, 96759, USA

    M.-L. Wang, L. Xiong, X. He, C. Nagai & G.-L. Nan

  2. Department of Tropical Plant and Soil Sciences, University of Hawaii at Manoa, 3190 Maile Way, Honolulu, HI, 96822, USA

    G. Uruu, K. T. Cheah & R. E. Paull

  3. Department of Plant and Environment Protection Sciences, University of Hawaii at Manoa, 3190 Maile Way, Honolulu, HI, 96822, USA

    J. S. Hu, B. S. Sipes & K. G. Rohrbach

  4. Centre for Plant Sciences, University of Leeds, Leeds, LS2 9JT, UK

    H. J. Atkinson

  5. USDA, ARS, PWA, Pacific Basin Agricultural Research Center, 64 Nowelo Street, Hilo, HI, 96720, USA

    P. H. Moore

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  1. M.-L. Wang
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Correspondence to R. E. Paull.

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Wang, ML., Uruu, G., Xiong, L. et al. Production of transgenic pineapple (Ananas cosmos (L.) Merr.) plants via adventitious bud regeneration. In Vitro Cell.Dev.Biol.-Plant 45, 112–121 (2009). https://doi.org/10.1007/s11627-009-9208-8

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  • DOI: https://doi.org/10.1007/s11627-009-9208-8

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