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Journal : Jurnal AgroBiogen

Construction of Begomovirus AV1 Gene Candidate into pBI121 and Its Introduction into Tobacco by using Agrobacterium tumefaciens Vector Santoso, Tri J.; Herman, Muhammad; Hidayat, Sri H.; Aswidinnoor, Hajrial; ., Sudarsono
Jurnal AgroBiogen Vol 7, No 1 (2011): Jurnal AgroBiogen
Publisher : Jurnal AgroBiogen

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Abstract

Infection of Begomovirus has caused leaf curl disease in tomato. This infection has significantly impact on yield losses of tomato production. Recently, in Indonesia there was no effectively way to control this disease.  The use of resistant tomato variety is one of strategies to control this virus. Genetic engineering technology gives an opportunity to develop the transgenic tomato resistant to Begomovirus through pathogen derived resistance (PDR) approach. The objectives of this study were to construct the  Begomovirus AV1 candidate gene in the pBI121 and to introduce the construct into tobacco plant genome through Agrobacterium tumefaciens vector. A series activites in gene construct have been conducted include PCR amplification of AV1 gene using a pair of specific primer, cloning the gene into pGEM-T easy, transformation of the clone into Escherichia coli DH5α competent cell, construct the gene into pBI121, and transform the construct into A. tumefaciens. Leaf segments of in vitro tobacco plant were transformed by co-cultivation with A. tumefaciens containing ToLCV-AV1 construct. In the research activitiy, Indonesian  Begomovirus  AV1 gene  was  successfully amplified and inserted in expression vector plasmid pBI121. Tobacco transformants carrying kanamycin-resistant gene (nptII gene) were regenerated and established in the glasshouse. Those transformant plants are expected containing the AV1 gene.
Delivering of Over-Expression Construct OsWRKY76 Candidate Gene in Rice cv. Nipponbare through Agrobacterium tumefaciens Apriana, Aniversari; Sisharmini, Atmitri; Enggarini, Wening; Sudarsono, Sudarsono; Khumaida, Nurul; Trijatmiko, Kurniawan R.
Jurnal AgroBiogen Vol 7, No 1 (2011): Jurnal AgroBiogen
Publisher : Jurnal AgroBiogen

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Abstract

Plant genetic improvement can be done through classical breeding or genetic engineering. WRKY is a transcription factor involved in regulating plant defense responses. OsWRKY76 gene is located in a narrow segment of chromosome 9 which is identified previously to be related to wide spectrum resistance in rice. A sequence of OsWRKY76 (+1.200 bp) has available in the gene bank and it makes possible to isolate, clone, and construct the gene into over-expression vector. The aim of this research was to assemble an over-expression construct of OsWRKY76 candidate gene and introduce it into rice through Agrobacterium-mediated transformation. A construct of pCAMBIA-1301::35S::OsWRKY76 has been successfully assembled and transformed into embryogenic calli of rice cv. Nipponbare using A. tumefaciens strain Agl-1 and EHA 105. A number of 126 independent lines has been produced, in which Agl-1 showed 3.8 times more efficient than EHA 105. PCR analysis of randomly selected 25 independent lines showed that all of them positively contained hptII gene, a selectable marker used in the over-expression construct of the OsWRKY76 candidate gene. Based on the result, it could be concluded that the over-expression construct of OsWRKY76 candidate gene have been successfully introduced into the tissue of Nipponbare.
Molecular Analysis and Effectiveness Assay of AV1 Gene in Transgenic Tobacco for Resistance to Begomovirus Tri Joko Santoso; Muhammad Herman; Sri H. Hidayat; Hajrial Aswidinnoor; Sudarsono Sudarsono
Jurnal AgroBiogen Vol 8, No 2 (2012): Agustus
Publisher : Balai Besar Penelitian dan Pengembangan Bioteknologi dan Sumber Daya Genetik Pertanian

Show Abstract | Download Original | Original Source | Check in Google Scholar | DOI: 10.21082/jbio.v8n2.2012.p45-53

Abstract

Genetic transformationof tobacco plant using AV1 gene was conducted atthe previously experiment and generated transgenic tobaccoplants positively carrying the selectable marker nptII gene.The objectives of this experiment were to (1) analyze thepresence of Begomovirus AV1 gene in T0 generation putativetransgenic tobacco plants using PCR technique with specificprimers and its correlation with resistance phenotype, (2)analyze the integration and copy number of the transgene inT0 generation putative transgenic tobacco plants and itscorrelation with resistance response, (3) screen the T0generation putative transgenic tobacco plants with the targetvirus infection and to detect the presence of the virus in thetransgenic plant tissue using universal primers. PCRdetection of AV1 gene in tobacco transgenic was conductedby using specific primer for Begomovirus AV1 gene.Meanwhile, Southern Blot analysis was conducted by usingthe AV1 gene probe. The effectiveness of AV1 gene intobacco transgenic was tested by inoculation of target virususing whiteflies vector. Result of the experiments showedthat there was a positive correlation between the presenceof the AV1 transgene in T0 generation putative transgenictobacco plants and the resistant phenotype. Transgenicplants with a single copy integration of the transgeneexhibited more resistant than the multiple copy one. andnon transgenic plant. The resistance as a result of AV1 geneexpression was indicated with no symptom in T0 generationtransgenic tobacco plants and the accumulation of the virusin the transgenic plants tissue. Northern and Westernhybridization analysis need to be perfomed for investigatingthe presence of mRNA or protein accumulation so that theresistance mechanism of the AV1 gene could be explainedmore detail.
Regenerasi Jeruk Siam melalui Embriogenesis Somatik Ali Husni; Agus Purwito; Ika Mariska; Sudarsono Sudarsono
Jurnal AgroBiogen Vol 6, No 2 (2010): Oktober
Publisher : Balai Besar Penelitian dan Pengembangan Bioteknologi dan Sumber Daya Genetik Pertanian

Show Abstract | Download Original | Original Source | Check in Google Scholar | DOI: 10.21082/jbio.v6n2.2010.p75-83

Abstract

The Regenerated of Siam Tangerin through SomaticEmbryogenesis. Ali Husni, Agus Purwito, Ika Mariska,and Sudarsono. Somatic embryogenesis occurs in mostplants that are cultured on a suitable medium in vitro.Somatic embryo may arise from single cells and the embryogeniccells are widely applicable in plant propagation,genetic manipulation and transgenic technologies. Thepresent study was carried out to develop an effectivesomatic embryogenesis technique to regenerate Siamtangerine plants. Materials used in this study were nucellartissues of young fruits (30-90 days post anthesis). Inductionof embryogenic calli was done by culturing the tissues onthree different basal media (MS, MW and MT). Embryomaturation was done on the MW medium + ABA (0; 0.1; 0.3;dan 0.5 mg/l), while germination to plantlet developmentwas done on WS medium + GA3 (0, 0.1, 0.3, and 0.5 mg/l).The results showed that among the three media, MW wasthe best medium for callus induction from nucellar tissueswith a success rates 98% for Simadu and 100% forPontianak. The best maturation of embryo somatic wasdone on MW medium + ABA 0.5 mg/l with success rates99%, while the best medium for germination and developmentinto planlets was MW medium + 0.5 mg/l GA3 with asuccess rate 58%.
Co-Authors , Agustiansyah , Rustikawati , Sukendah , Triadiati , Witjaksono ABDUL RACHMAD BUDIONO Ade Wachjar Adisyahputra Adisyahputra, Adisyahputra Agung Endro Nugroho Agus Purwito Agus Rachmat AHMAD RIDUAN Ahmad Surkati Ali Husni Amarillis, Shandra Amarullah, Muhamad Roffi Andrie Al Kausar Anita Yuliyanti Aniversari Apriana Anneke Pesik Anwar, Aswaldi Aprizal Zainal Arfan Nazhri Simamora Arief Hidayat Aris Munandar Asep Setiawan ASMONO, DWI Atmitri Sisharmini Bambang Heliyanto Bambang S. Purwoko BAMBANG SUGIRI Barahima Abbas C J Soegihardjo Cartealy, Imam Civi Cartealy, Imam Civi Catur Herison Cecep Kusmana Christoph Leuschner Clarry Sada Colleen M. Higgins, Colleen M. Darmawan Saptadi Deka Rotama Dewi Sukma Didiek Hadjar Goenadi Diky Anggoro DINARTY, DINY Dini Dinarty, Dini DINI NURDIANI Diny Dinarti Djoko Santoso DWI ANDREAS SANTOSA Dwi Asmono DWI HAPSORO Dwi Hapsoro Edi Guhardja Efendi, Darda Elina, Juanita ELSJE T. TENDA, ELSJE T. Endang Pudjihartati Endrizal Eneng Fakhrunnisa Enni Suwarsi Rahayu Erick Raynalta, Erick Fajar Laksono Suroso Fakhrunnisa, Eneng Fatsan, Ahmad FETRINA OKTAVIA Germini Alam Giyanto . Gusti Ayu Kade Sutariati H. M.H. Bintoro Djoefrie HAJRIAL ASWIDINNOOR Harsini ,, Harsini Hartati, Raden Roro Sri Haryadi Permana Hasriadi Mat Akin Hengky Novarianto Hengky Novarianto Hidayat Pawitan Hiroshi Ehara Hugo Volkaert, Hugo I MADE ARTIKA IBNUL QAYIM IDA HANARIDA SOMANTRI Iim Fatimah Ika Mariska Ikhsanudin Ikhsanudin Irman Firmansyah Ismail Maskromo Ismail Maskromo Iwa Sutardjo, Iwa Iwan Setiawan JAJAH KOSWARA Juang Gema Kartika Jumanto Jumanto Jumanto Jumanto, Jumanto Khoerul Anwar Khursatul Munibah Kukuh Murtilaksono Kukuh Setiawan Kurniasari, Praharisti Kurniawan R. Trijatmiko Kusumo Nugroho Kuswanhadi Kuswanhadi, Kuswanhadi Laksmita Prima Santi Laksono Trisnantoro Lila Yuwana Lollie Agustina P. Putri M. Herman Marbelisa Briliani Mardikaningsih, Agusti Marenda Ishak S MASKROMO, ISMAIL MEITY A. TULALO, MEITY A. Memen Surahman Meynarti Sari Dewi Ibrahim Muchammad Ali Safaat Muhammad Hasyim Bintoro MUHAMMAD HERMAN Muhammad Herman Muhammad Machmud Muhammad Machmud Mutia Dewi Yuniati Neviana Wahyuni Nihayati, Hanik Endang Niniet Indah Arvitrida Noor Cahaya NOVARIANTO, HENGKY Nur Khayati Nurhajati A. Mattjik Nursalam Nursalam Nurul Khumaida Paulus B.K. Santoso Purwaningsih Purwaningsih Purwoko, Devit purwoko, devit R B Gobel Ralf G. Dietzgen, Ralf G. Reni Indrayanti ROEDHY POERWANTO Rr Sri Hartati Rr Sri Hartati, Rr Sri Rr. Sri Hartati Rubiyo Rubiyo RULLY DYAH PURWATI RUSMILAH SUSENO S. Arsyad S. Ilyas Said Harran Saiful Akhyar Lubis SATRIYAS ILYAS Satriyas Ilyas Satya Nugroho Shinta Hadiyantina Sholeh Avivi Siswanto Siswanto Siti Mufidah Siti Sunarintyas SOEKISMAN TJITROSEMITO Sri H. Hidayat Sri H. Hidayat Sri H. Hidayat Sri Indarto Sri S. Harjadi Sri Soenarsih DAS Stephanie Wilamarta Sudari, Sudari Sudibyo Martono Sudirman Yahya SUDJINDRO SUDJINDRO Sugiyanta Sukendah S, Sukendah SUKENDAH SUKENDAH, SUKENDAH Sumali Wiryowidagdo Suryo Wiyono Susilo Indrawati Swastika, Maria Syafaruddin Syafaruddin TENDA, ELSJE T. Teuku Tajuddin TINCHE, nFn Tri J. Santoso Tri J. Santoso Tri Joko Santoso Tri Joko Santoso U.S WIRADISASTRA UJANG KHAIRUL Urai Salam Visia Assyafira Suwarto vyta andri setyo utami Wening Enggarini Widiatmaka . Widiatmaka Widiatmaka Widodo Widodo Wiwin Ambarwulan Yudiwanti Wahyu E. K. Yulis Setiya Dewi Yundari, Yundari Yuni Wibowo Yusnita Yusnita