Detail High Efficiency and Regeneration Protocol of Tobacco Transformation mediated by Agrobacterium tumefaciens LBA4404 (presented at Workshop: Biofuel Research & Development: An Asian Perspective session 5 March 2011 in Keystone Symposia 40th Meeting Season Singapore, 1-6 March 2011) Bibliografi Author: Karmawan, Listya Utami ; Angliana Topik: Transformation; A. tumefaciens; tobacco; protocol; JABFUNG-FTB-LUK-2022 Bahasa: (EN )     Penerbit: Keystone Symposia on Molecular and Cellular Biology     Tempat Terbit: Singapore    Tahun Terbit: 2011     Jenis: Poster Fulltext: 2011_Seminar Int_Keystone_highres.pdf (9.94MB; 3 download) Abstract Nicotiana tabacum (tobacco) was known as a model in plant biotechnology. Nowadays, transgenic tobacco was used for biofuel production. New traits can be introduced to this plant via Agrobacterium mediated transformation procedure. Inefficient transformation efficiency in tobacco leaf disk protocol was caused by low regeneration rate and necrosis after the process of co-cultivation. In this study, we tried to combine techniques to increase the transformation and regeneration efficiency. Cell density of A. tumefaciens lower than 10^10 CFU/ml which was reached at optical density (lambda = 600nm) 0.30 was used for transformation. Concentration of 250 ppm cefotaxime was used to increase the inhibition of A. tumefaciens growth. This concentration did not inhibit the formation of tobacco shoot. Explant regeneration on the selection medium with increased hygromycin concentration from 5 ppm up to 25 ppm enhanced the success of transgenic shoot formation. The best transformation efficiency was 43.70±3.39% obtained from transformation with addition of 50 mikroM acetosyringone and 0.05% Tween20 in the inoculation stage compared to control, which was only 11.67±4.41%. Putative transgenic shoot was confirmed using 35S primers and generated different lengths of amplicons (122, 123 and 449 bp). This result was consistent with in silico test using FASTPCR 6.0.165. In conclusion, this combined protocol can be used as alternative method for tobacco transformation. Opini Anda Klik untuk menuliskan opini Anda tentang koleksi ini! Lihat Sejarah Pengadaan  Konversi Metadata   Kembali

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