* Corresponding Author
Stable Transformant of Phalaenopsis amabilis Somatic Embryo Carrying 35S::AtRKD4 Develops Into Normal Phenotype of Transgenic Plant
https://doi.org/10.22146/jtbb.59210 Naufal Ghozi Aditya Perdana(1), Windi Mose(2), Muhammad Dylan Lawrie(3), Jose Gutierrez-Marcos(4), Endang Semiarti(5*)
(1) Faculty of Biology, Universitas Gadjah Mada
(2) Faculty of Mathematics and Natural Sciences, Pattimura University
(3) Faculty of Biology, Universitas Gadjah Mada
(4) School of Life Sciences, University of Warwick
(5) Faculty of Biology, Universitas Gadjah Mada
(*) Corresponding Author
Abstract
Phalaenopsis amabilis (L.) Blume orchid is an Indonesian national flower. The number of these orchids in their natural habitat is very limited, therefore plant propagation efforts are needed. One of the promising methods is plant propagation by inserting embryo gene AtRKD4 from a model plant Arabidopsis thaliana into the orchid genome to produce many somatic embryos. From previous research, we have obtained 28 plant P. amabilis transformants carrying the AtRKD4 gene, however, it was unknown whether these plants have normal phenotypes and growth similar to their parents. Therefore, descriptions on growth and morphology are needed. This research aimed to evaluate the phenotype of P. amabilis carrying 35S::AtRKD4 the transformants grown in greenhouse. To achieve it, AtRKD4 gene integration stability on transformants genome was analyzed. Morphology and cross-section anatomy structure on transformant and non-transformant plantswere described. The stability of AtRKD4 gene integration in the plant genome was confirmed by amplification of the AtRKD4 gene from genomic DNA with Polymerase Chain Reaction (PCR) using a specific primer for AtRKD4 and ACTIN genes as the internal control. The quantitative data from morphology and anatomy measurements were analyzed statistically using ANOVA. The results showed that AtRKD4 was stably integrated into the genome of P. amabilis transformants and all transformant plants showed similar morphology and anatomy characteristics as non-transformant plants. The AtRKD4 embryo gene was stably integrated into the orchid genome and the transformant plants grow normally without significant changes in phenotype.
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Feng J.-H., Chen J.-T., 2014. A novel in vitro protocol for inducing direct somatic embryogenesis in Phalaenopsis aphrodite without taking explants. Scientific World Journal, pp.1-7.
Hsing H.-X., Lin Y.-J., Tong C.-G., Li M.-J., Chen Y.-J., Ko S.-S., 2016. Efficient and heritable transformation of Phalaenopsis orchid. Botanical Studies, 57(30).
Jeong, S., Palmer, T. & Lukowitz, W., 2011. The RWP-RK factor GROUNDED promotes embryonic polarity by facilitating YODA MAP kinase signaling. Current Biology, 21(15), pp.1268-1276.
Kőszegi, D., Johnston, A.J., Rutten, T., Czihal, A., Altschmied, L., Kumlehn, J., Wüst, S.E.J., Kirioukhova, O., Gheyselinck, J., Grossniklaus, U. & Bäumlein, H., 2011. ‘Members of the RKD transcription factor family induce an egg cell-like gene expression program’. Plant Journal, 67(2), pp.280–291.
Mirani, A.A., Abul-Soad, A.A. & Markhand, G.S., 2017. Effect of different substrates on survival and growth of transplanted orchids (Dendrobium nobile cv.) into net house, International Journal of Horticulture and Floriculture, 5(4), pp.310-317.
Mose, W., Indrianto, A., Purwantoro, A. & Semiarti, E., 2017. The influence of thidiazuron on direct somatic embryo formation form various types of explant in Phalaenopsis amabilis (L.) Blume. Hayati Journal of Biosciences, 24, p.202.
Mose, W., Daryono, B.S., Indrianto, A., Purwantoro, A. & Semiarti, E., 2020. Direct somatic embryogenesis and regeneration of an Indonesian orchid Phalaenopsis amabilis (L.) Blume under a variety of plant growth regulators, light regime, and organic substances. Jordan Journal of Biological Sciences, 13(4), pp.509-518.
Mursyanti, E., Purwantoro, A., Moeljopawiro, S. & Semiarti, E., 2015. Induction of somatic embryogenesis through overexpression of AtRKD4 genes in Phalaenopsis “Sogo Vivien”. Indonesian Journal of Biotechnology, 20(1), p.43.
Mursyanti, E., Purwantoro, A., Moeljopawiro, S. & Semiarti, E., 2016. Micropropagation of mini orchid hybrid Phalaenopsis “Sogo Vivien”. Journal of Tropical Biodiversity and Biotechnology, 1(1), pp.45-53.
Palma, M.A., Chen, Y., Hall, C., Bessler, D. & Leatham, D., 2010. Consumer preferences for potted orchids in the Hawaiian market, HortTechnology, 20(1), p.239.
Pamarthi, R.K., Devadas, R., Kumar, R., Rai, D., Babu, P.K., Meitei, A.L., De L.C., Chakrabarthy, S., Barman, D. & Singh, D.R., 2019. PGR diversity and economic utilization of orchid. International Journal of Current Microbiology and Applied Sciences, 8(10), pp.1865-1887.
Saputro, T.B., Semiarti, E. & Purwantoro, A., 2018. Phenotypic and molecular characterization of multishoots development in transgenic Phalaenopsis Amabilis (L.) Blume harboring 35S::KNAT1 (KNOTTED-LIKE Arabidopsis thaliana 1). Biotropia, 25(1), pp.11-21.
Schuiteman, A., 2010. Proceeding International Seminar on Orchid Conservation and Agribusiness, Faculty of Biology UGM.
Semiarti, E., Indrianto, A., Purwantoro, A., Isminingsih, S., Suseno, N., Ishiwaka, T., Yoshioka, Y. Machida, Y. & Machida, C., 2007. Agrobacterium-mediated transformation of the wild orchid species Phalaenopsis amabilis. Plant Biotechnology, 24(3), pp.265-272.
Semiarti, E., Indrianto, A., Purwantoro, A., Machida, Y. & Machida, C., 2011. Agrobacterium-mediated transformation of Indonesian orchids for micropropagation, in M.A. Alavarez (ed.). Genetic Transformation, pp. 215-240.
Semiarti, E., Mursyanti, E., Suyoko, A., Perdana, F.S.W., Widyastuti, C.T. & Subchan, A.N., 2018. Stability of T-DNA integration in Phalaenopsis “Sogo Vivien” transgenic orchid carrying 35S::Gal4::AtRKD4::GR. Biology’. Medicine, & Natural Product Chemistry, 7(1), pp.5-13.
Setiari, N., Purwantoro, A., Moeljopawiro, S. & Semiarti, E., 2018. Micropropagation of Dendrobium phalaenopsis orchid through overexpression of embryo gene AtRKD4’, AGRIVITA Journal of Agricultural Science, 40(2), p.285.
Shekarriz, P., Kafi M., Deilamy S. D., Mirmasoumi M., 2014. Coconut water and pepton improve seed germination and protocorm-like body formation of hybrid Phalenopsis. Agriculture Science Development, 3, pp.317-322.
Sutikno, 2016. Bahan Ajar Mikroteknik Tumbuhan. [English: Plant Microtechnic Teaching Materials]. Faculty of Biology, Universitas Gadjah Mada. Yogyakarta. pp.28-32.
Waki, T. Hiki, T., Watanabe, R., Hashimoto, T. & Nakajima, K., 2011. The Arabidopsis RWP-RK protein RKD4 triggers gene expression and pattern formation in early embriogenesis. Current Biology, 21, pp.1277-1281.
Yang, Y., Wang N., Zhao S, 2020. Functional characterization of a WRKY family gene involved in somatic embryogenesis in Panax ginseng. Protoplasma, 257, pp.449-458.
Yuan, X-Y., Jiang, S-H., Wang, M-F., Ma, J., Zhang, X-Y. & Cui, B., 2014, Evaluation of internal control for gene expression in Phalaenopsis by quantitative real-time PCR. Applied Biochemistry and Biotechnology, 173, pp.1431-1445.
Zimmerman, J.L., 1993. Somatic embryogenesis: a model for early development in higher plants. Plant Cell, 5, pp.1411–1423.
Zulwanis, Nintya Setiari, Jose Gutierrez-Marcos, and Endang Semiarti , 2020. The expression of AtRKD4 transgene during induction of somatic embryogenesis in transgenic Dendrobium phalaenopsis orchid carrying 35S::GR::AtRKD4, AIP Conference Proceedings 2260, 060015.
DOI: https://doi.org/10.22146/jtbb.59210
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