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Optimization of micropropagation and Agrobacterium-mediated gene transformation to spinach (Spinacia oleracea L.)

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DOI: 10.4236/abb.2012.37109    3,636 Downloads   6,274 Views   Citations

ABSTRACT

Spinach is one of the dioecious plant which is considered as a model plant in genetic and molecular studies of sex determination because of its special characteristics such as low chromosome number and short life cycle. An efficient protocol for Spinacia oleracea Agrobacterium-mediated gene transformation was developed. The leaf disks, roots, hypocotyls and cotyledons of this plant were inoculated with LBA4404. LBA4404 carrying pCAMBIA3301 binary vector with 35SCaMV gusint and 35SCaMV bar cassettes. Effects of two preparation condition (induction of vir genes and noninduction) were considered. Also effects of different number days of co-cultivation and pre-culture of explants were examined. After co-cultivation, the explants were transferred to regeneration medium containing 250 mg·L-1 Carbeniciline. Transient expression efficiency was calculated based on the number of blue spots per explants one week after inoculation. Based on the results of transient expression, stable transformation was carried out. After formation of callus the histochemical GUS assay was carried out on some parts of them and other parts were leaved for being regenerated.

Conflicts of Interest

The authors declare no conflicts of interest.

Cite this paper

Naderi, D. , Zohrabi, Z. , Shakib, A. , Mahmoudi, E. , Khasmakhi-Sabet, S. and Olfati, J. (2012) Optimization of micropropagation and Agrobacterium-mediated gene transformation to spinach (Spinacia oleracea L.). Advances in Bioscience and Biotechnology, 3, 876-880. doi: 10.4236/abb.2012.37109.

References

[1] Bao, J.H., Chin, D.P., Fukami, M., Ugaki, M., Nomura, M. and Mii, M. (2009) Agrobacterium-mediated transformation of spinach (Spinacia oleracea) with Bacillus thuringiensis cry1Ac gene for resistance against two common vegetable pests. Plant Biotechnology, 26, 249-254. doi:10.5511/plantbiotechnology.26.249
[2] Geekiyanage, S., Takase, T., Watanabe, S., Fukai, S. and Kiyosue, T. (2006) The combined effect of photoperiod, light intensity and GA3 on adventitious shoot regeneration from cotyledons of spinach (Spinacia oleracea L.) Plant Biotechnology, 23, 431-435. doi:10.5511/plantbiotechnology.23.431
[3] Neskovic, M. and Radojevic, L. (1973) The growth of and morphogenesis in tissue cultures of Spinacia oleracea L. Bulletin De Linstitute Et Du Jardin Botaniques De Luniversite De Beograd, 8, 35-37.
[4] Al-Khayri, J.M., Huang, F.H., Morelock, T.E., Busharar, T.A. and Gbur, E.E. (1991) Genotype dependent response of spinach cultivars to in vitro callus induction and plant regeneration. Plant Science, 78, 121-127. doi:10.1016/0168-9452(91)90168-8
[5] Al-Khayri, J.M., Huang, F.H., Morelock, T.E. and Busharar, T.A. (1992) Stimulation of shoot regeneration in spinach callus by gibberellic acid. Horticultural Science, 27, 1046.
[6] Komai, F., Okuse, I. and Harada, T. (1996) Somatic embryogenesis and plant regeneration in culture of root segments of spinach (Spinacia oleracea L.). Plant Science, 113, 203-208. doi:10.1016/0168-9452(95)04285-7
[7] Komai, F., Okuse, I. and Harada, T. (1996) Effective combinations of plant growth regulators for somatic embryogenesis from spinach root segments. Journal of the Japanese Society for Horticultural Science, 65, 559-564. doi:10.2503/jjshs.65.559
[8] Knoll, K.A., Short, K.C., Curtis, I.S., Power, J.B. and Davey, M.R. (1997) Shoot regeneration from cultured root explants of spinach (Spinacia oleracea L.): A system for Agrobacterium transformation. Plant Cell Reports, 17, 96-101. doi:10.1007/s002990050359
[9] Zhang, H.X. and Zeevaart, J.A.D. (1999) An efficient Agrobacterium tumefaciens-mediate transformation and regeneration system for cotyledons of spinach (Spinacia oleracea L.). Plant Cell Reports, 18, 640-645. doi:10.1007/s002990050635
[10] Ishizaki, T., Komai, F. and Masuda, K. (2001) Screening for strongly regenerative genotypes of spinach in tissue culture using subcultured root explants. Plant Cell, Tissue and Organ Culture, 67, 251-255. doi:10.1023/A:1012791611632
[11] Molvig, L. and Rose, R.J. (1994) A regeneration protocol for Spinacia oleracea using gibberellic acid. Australian Journal of Botany, 42, 763-769. doi:10.1071/BT9940763
[12] Goto, T., Miyazaki, M. and Oku, M. (1998) Varietal variations in plant regenerative potential from protoplasts in spinach (Spinacia oleracea L.). Journal of the Japanese Society for Horticultural Science, 67, 503-506.
[13] Leguillon, S., Charles, G. and Branchard, M. (2003) Plant regeneration from thin cell layers in Spinacia oleracea. Plant Cell, Tissue and Organ Culture, 74, 257-265. doi:10.1023/A:1024042522940
[14] Mii, M., Nakano, M., Okuda. K. and Iizuka, M. (1992) Shoot regeneration from spinach hypocotyls segments by short term treatment with 5,6-dichloro-indole-3-acetic acid. Plant Cell Reports, 11, 58-61. doi:10.1007/BF00235253
[15] Ishizaki, T., Megumi, C., Komai, F., Masuda, K. and Oosawa, K. (2002) Accumulation of a 31-kDa glycoprotein in association with the expression of embryogenic potential by spinach callus in culture. Plant Physiology, 114, 109-115. doi:10.1034/j.1399-3054.2002.1140115.x
[16] Davey, M.T., Rech, E.L. and Mulligan, B.J. (1989) Direct DNA transfer to plant cells. Plant Molecular Biology, 13, 273-285. doi:10.1007/BF00025315
[17] Cervera, M., Pina, J.A., Juarez, J., Navarro, L. and Pena, L. (1998) Agrobacterium mediated transformation of citrange: Factors affecting transformation and regeneration. Plant Cell Reports, 18, 271-278. doi:10.1007/s002990050570
[18] Cao, X., Liu, Q., Rawland, L.J. and Hammerschlag, F.R. (1998) GUS expression in Blueberry (Vaccinium spp.) factors influencing Agrobacterium mediated gene transfer efficiency. Plant Cell Reports, 18, 266-270. doi:10.1007/s002990050569
[19] Shuangxia, J., Xianlong, Z., Shaoguang, L., Yichun, N., Xiaoping, G. and Chao, H. (2005) Factors affecting transformation efficiency of embryogenic callus of upland cotton (Gossypium hirsutum) with Agrobacterium tumefaciens. Plant Cell, Tissue and Organ Culture, 81, 229-237. doi:10.1007/s11240-004-5209-9
[20] Moralejo, M., Rochange, F., Boudet, A.M. and Teulieres, C. (1998) Generation of transgenic Eucalyptus globulus plant lets through Agrobacterium tumefaciens-mediated transformation. Australian Journal of Plant Physiology, 25, 207-212. doi:10.1071/PP97041
[21] Geeta, N., Venkatachalam, P. and Lakshmi, S. (1999) Agrobacterium mediated genetic transformation of pigeon pea (Cajanus cajan L.) and development of transgenic plants via direct organogenesis. Plant Biotechnology, 16, 213- 218. doi:10.5511/plantbiotechnology.16.213
[22] Lawrence, P.K. and Koundal, K.R. (2001) Agrobacterium tumefaciens mediated transformation of pigeon pea (Cajanus cajan L. Millsp) and molecular analysis of regenerated plants. Current Science, 80, 1428-1432.
[23] Jefferson, R.A. (1987) Assaying chimeric genes in plant-The GUS gene fusion system. Plant Molecular Biology Reporter, 4, 387-405.
[24] Murashige, T. and Skoog, F. (1962) A revised medium for rapid growth and bioassay with tobacco tissue cultures. Plant Physiology, 15, 473-497. doi:10.1111/j.1399-3054.1962.tb08052.x

  
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