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Journal : Indonesian Journal of Biotechnology

Early detection of the orchid flowering gene PaFT1 in tobacco cells using a GFP reporter

Indonesian Journal of Biotechnology Vol 21, No 1 (2016)
Publisher : Universitas Gadjah Mada

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Abstract

Here we describe a novel method of using green fluorescence protein (GFP) as a reporter gene for early detection of an integrated T­DNA containing the orchid flowering gene, PaFT1 (Phalaenopsis aphrodite Flowering locus T1) in the tobacco genome. Functional assays that report the presence of exogenous DNA early in development are especially useful in plants where the desired phenotype is only apparent after long periods of vegetative growth. The objective of this study is to establish a method for detecting an inserted Phalaenopsis orchid flowering gene and examining its function in tobacco. The p35S::PaFT1­ 35S::GFP construct was introduced into Agrobacterium tumefaciens strain EHA101. Transformed tobacco leaves were cultured on MS medium with addition of 1 mgL-1 NAA+3 mgL-1 BAP+50 mgL-1 Kanamycin+300 mgL-1 timentin for selection. Results showed bright green GFP fluorescent signals in 11 out of 15 (73%) tobacco leaf cells at a 2­month time point after transformation. GFP and PaFT1 fragments were amplified in genomic PCR using GFP and PaFT1 specific primers. The accumulated PaFT1 transcripts were observed in 3 month­old transgenic tobacco plants containing p35S::PaFT1­35S::GFP. Green florescence was observed only in the transgenic plants at the 5 month­old stage but not in the wild type controls.

Co-Authors . Indarto ., Yusetyowati Abdullah, Teuku Aceng Komarudin Mutaqin Achmad subhi Gumilar Adyatma, Rengga Agus Supriyadi Agus Yodi Gunawan Ahmad Sofwan Ahmad Yusron, Ahmad Akriono, Muhammad Luqman Amri, Khabib Saiful Andries Lionardo, Andries Ani Rusilowati Anis Khiyaroh, Anis Arca Datam Sugiarto, Arca Datam Arif Budi Satrio Ash Shiddiq, Dimas Ghufron Ash-Shiddiq, Dimas Ghufron Askin, Askin Askin, Askin Astuti, Desi Widi Aulanni’am a Aziza, Siti Nur Budi Prayitno BUDI SETIADI DARYONO Cahyono Purbomartono DAHRI, IRSYAD Dani, Andrea Tri Rian Denny Irbianto Devi, Mike Indriana Dinda, Susanne Dini Siswani Mulia Djumadi Djumadi Edijatno Edijatno Eka Dewi Nurjayanti Eka Haryani, Eka Eko Prasetyo Eko Pudjadi, Eko Elida Novita Endah Subekti Endang Semiarti Erly Anna Rosita Ernawiati, Eti Ervi Rachma Dewi, Ervi Rachma F. Parasmawati Firman Fuad Wirakusumah Fitri, Susi Gofar Ismail Hamid Ahmad Hardiyanto Hardiyanto Hari Wijayanto Harsi D. Kusumaningrum Heri Nugroho Hery Maryanto, Hery Husaini . Husin, Farid I Wayan Suteja Ikhwanudin Ikhwanudin Indarto - Indarto Indarto Indriani . Ishak Abdulhak Istanto Istanto Istianah Istianah Jang, Soenghoe JB. Suparyatmo Suparyatmo, JB. Suparyatmo Jenny Ratna Suminar Joko Suryanto Joko Tri Sujiwo Khairunnissa, Aulia Khoiriyah, Naila Ulfa Komalasari, Gantina Lalu A. Zaenuri Latipah, Latipah Lawrie, Muhammad Dylan Lepong, Piter Lepong, Piter lina marlina, lina LUKMAN ILHAM, LUKMAN Lutfi Aris Sasongko M Nur Ihsan Malihatin Munawaroh, Malihatin Martha Lulus Lande, Martha Lulus Masyhudi, Lalu Mateus Sakundarno Adi, Mateus Sakundarno Moh. Djazari Muhamad Afandi, Muhamad Muhammad Ade Salim Muhardjo Pudjojono, Muhardjo Muslim, Khaerul Nadjadji Anwar Nafrida Ulfah, Nafrida Nanang Sugiarto Nanda Arista Rizki Nasir Saleh Nathan Hindarto Nieke Karnaningroem Nina Fitriyati Ningtias, Rahayu Ninik Hasanah, Ninik Nuada, I Wayan Nugraha E. Suyatma, Nugraha E. Nugroho Arifianto Nur Ducha Nur Hidayah, Afifah Nurjazuli Nurjazuli NURTITI SUNUSI Pradana, Hendra Andiananta Pramudya Annisa Firdausi Pratiwi Trisunuwati Puji Wahyumi, Puji Rahmadiyanti, Ridhani Ramdani, Aulia Ratna P, Kenah Renan Subantoro Rini Rita T. Marpaung Risma Sake, Risma Rita Irma, Rita Rossi Prabowo Rosyid Ridho Royaningsih, Nanik RUBEN DHARMAWAN Saefudin Saefudin Sahat, Afro Aransa Sasono Wibowo Satrijo Budiwibowo, Satrijo Saud Irsyad, Nasihin Savitri, Intan Maya Setiawan Setiawan Setyono, Bagus Dwi Hari Shinta Arum Wahyuningsih, Shinta Arum Shofia Nur Awami Sitti Raodhah, Sitti Soekarsono Hardjono, Soekarsono Sri Hartini Sri Heny Wahyuningsih, Sri Heny Sri Suparwitri, Sri St. A. Rahayuningsih Stiawan, Agus Sucipto Sucipto Sudaryati ., Sudaryati Suhardjo Widodo Dan, Suhardjo Widodo Suhartono Suhartono Suharyo Hadisaputro Sukadi, Desi Rahmawati Sukarti Moeljopawiro Suprapti Supardi Suprati Supardi Supriyanto Supriyanto Suroso . Susiarno, Hadi Sutawanir Darwis Sutiman B. Sumitro Sutrisno sutrisno Suyadi Suyadi Suyitno Suyitno Suyoto Suyoto Syahrul Basri Syaripuddin Syaripuddin Tino Bachtiar, Tino TITIK SUNDARI Tri Jalmo Trinil Susilawati Utami Dyah Syafitri Utami Kurniawati Wildana Latif Mahmudi, Wildana Latif Yanti Setianti Yudhatama, Tri Galih Yudi Widodo Yulianto Nugroho Yulianty Yulianty Yulinah S, Elin Yuwono Yuwono Zulkifli Zulkifli