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Organogenic Regeneration of Transgenic Plant by Agrobacteriummediated DNA transformation of Citrus Wirawan, I Gede Putu; Subandiyah, Siti; Ngurah Suprapta, Dewa; Arya, Nyoman; Supartana, Putu; Sudana, Made
Journal of Bioscience and Biotechnology Vol. 1, No. 1 Januari 2007
Publisher : Journal of Bioscience and Biotechnology

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Abstract

Internodal stem segment from citrus seedling were cultured and inoculatedwith Agrobacterium tumefaciens harboring binary Ti plasmid vector that containedthe genes for detecable marker ß-glucuronidase (GUS) and the selectable markerNptII. The result of this study show that shoots can be regenerated in media with 100?g/ml kanamycin and about 10 % of them were GUS+ shoots. Some of the GUS+shoots were confirmed by polymerase chain reaction (PCR) analysis for GUS genedetection in transformed shoots.
Molecular and Biochemical Detection of Fusarium Oxysporum f.sp. cubense as the Pathogen of Fusarium Wilt Disease on Banana (Musa spp.) ., masanto; ., parwito; wibowo, arif; subandiyah, siti
Jurnal Hortikultura Indonesia Vol 1, No 1 (2010): Jurnal Hortikultura Indonesia
Publisher : Perhimpunan Hortikultura Indonesia

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Abstract

Molecular and biochemical characteristics of Fusarium oxysporum f.sp. cubense (Foc) were detected. Six of Indonesian Foc isolates were artificially inoculated on “Ambon Kuning” banana. DNA of one-week culture isolates was extracted by three methods prior to PCR assay using Foc TR4 (tropical race 4) specific primer. Activity of extracellular enzyme was determined with reduction sugar, agar diffusion and SDS-PAGE assays. Statistical analysis revealed that all isolates insignificantly caused Fusarium wilt symptoms on tested banana with disease severity index ranging from 3 to 3.6. Maximum DNA concentration was obtained by CTAB method (766.25 µg mL-1), followed by SDS and alkaline lysis methods, i.e. 553.75 and 211.25 µg mL-1, respectively. PCR analysis showed that Bnt2 and Kjg1 isolates positively reacted to TR4 of Foc primer (DNA size of 1400 bp approximately). Reduction sugar and agar diffusion assays demonstrated that Kjg1 isolate significantly produced more extracellular enzyme, with 6.53 × 10-2 mg mL-1 in concentration and 20 mm in halo diameter. Meanwhile, SDS-PAGE assay viewed diverse bands of tested fungi (20.6 to 80 kDa), representing four extracellular enzymes. Positive PCR results highlighted the presence of Foc TR4 infecting banana in Indonesia. Various activities of extracellular enzymes did not influence the pathogenicity of Foc.   Key words: pathogenicity, DNA concentration, extracellular enzyme
Evaluation of Some Specific Primer Sets Development for Detecting Fusarium oxysporum f. sp. cubense Tropic Race 4 (Foc TR4) Originating from Indonesia Pratama, Yudha; Wibowo, Arif; Widiastuti, Ani; Subandiyah, Siti; Widinugraheni, Sri; Rep, Martijn
Jurnal Perlindungan Tanaman Indonesia Vol 22, No 1 (2018): (In Press)
Publisher : Universitas Gadjah Mada

Show Abstract | Download Original | Original Source | Check in Google Scholar | DOI: 10.22146/jpti.25037

Abstract

Fusarium oxysporum f. sp. cubense tropic race 4 (Foc TR4) strain which belong to Vegetative Compatibility Group (VCG) 01213 is the most devastating disease in global banana production. Validation of specific primer sets using the positive control (Foc TR4). In total, 50 isolates of Foc are collected from several banana production regions in Indonesia represent the group of VCG, races, genotype, cultivars, which are confirmed as Foc based on the tested using FocEf3 primer set, except Cjr-2 and Lmp-4 isolates. Foc-1/Foc-2 could amplify 34 Foc isolates included in Foc race 4. Three specific primer sets i.e. TR4-F/TR4-R, Six-1c, and TR4-F2/TR4-R1 are used to classify Foc isolates into Foc tropic race 4. TR4-F/TR4-R is known have the highest specificity as it could amplify 35 Foc isolates including positive controls (Foc TR4) compared to the other primer sets (Six-1c and TR4-F2/TR4-R1). This research indicates that there are a large number of diversity strains found in Foc isolates to be studied for further research. Race 4 of Foc (STR4 or TR4) is known to be widespread in several regions in Indonesia. Therefore, specific primer set development needs to be done to detect Foc TR4 and the most damaging strains on Foc TR4 based on molecular data.IntisariFusarium oxysporum f. sp. cubense ras 4 tropika (Foc TR4) yang termasuk ke dalam kelompok VCG 01213 merupakan patogen yang paling merusak dalam produksi tanaman pisang secara global. Validasi primer spesifik berbasis PCR menggunakan kontrol positif (Foc TR4). Total, 50 isolat Foc dikoleksi dari  beberapa daerah produksi pisang di Indonesia mewakili VCG, ras, genotipe dan kultivar yang dikonfirmasi sebagai isolat Foc berdasarkan pengujian menggunakan primer FocEf3, kecuali isolat Cjr-2 dan Lmp-4. Foc-1/Foc-2 dapat mengamplifikasi 34 isolat Foc yang termasuk ke dalam Foc ras 4. Selanjutnya tiga pasang primer spesifik yaitu TR4-F/TR4-R, Six-1c, dan TR4-F2/TR4-R1 digunakan untuk mengelompokkan isolat-isolat tersebut ke dalam isolat Foc ras 4 tropika. TR4-F/TR4-R diketahui memiliki spesifitas tertinggi karena dapat mengamplifikasi sebanyak 35 isolat Foc termasuk kontrol positif (Foc TR4) dibandingkan dengan primer lainnya (Six-1c dan TR4-F2/TR4-R1). Penelitian ini menunjukkan bahwa terdapat sejumlah besar keragaman strain yang terlihat pada isolat-isolat Foc tersebut untuk dapat dipelajari lebih lanjut. Ras 4 dari Foc (STR4 atau TR4) diketahui tersebar luas pada beberapa daerah di Indonesia. Oleh karena itu, perlu dilakukan pengembangan primer spesifik untuk mendeteksi Foc TR4 dan strain yang paling merusak pada Foc TR4 berdasarkan data molekuler.
Peranan Faktor Cuaca terhadap Infeksi dan Perkembangan Penyakit Bercak Ungu pada Bawang Merah Hadisutrisno, Bambang; Sudarmadi, Sudarmadi; Subandiyah, Siti; Priyatmojo, Achmadi
Jurnal Perlindungan Tanaman Indonesia Vol 1, No 1 (1995)
Publisher : Universitas Gadjah Mada

Show Abstract | Download Original | Original Source | Check in Google Scholar | DOI: 10.22146/jpti.9351

Abstract

Purple blotch disease caused by Alternaria porri (Ell.) Cif. is one of the most important diseases on shallots. The farmers have been using fungicides as a reliable control method, however, the chemical control method has to be considered for the negative effect on the environment and the danger to the other species and causes the resistance to a certain pesticide. Therefore, it is important to develop other method that is safe, effective, and efficient, like the use of climatic data to decide the control strategy.The result of the study on ecology of purple blotch disease carried out at the experimental station in Kramat-Tegal, Kledung-Temanggung, and KP4 Kalitirto in dry and rainy season (1991/1992) suggested that climatic factors played on important role on the daily conidium dissemination of Alternaria porri and on the intensity of purple blotch disease. Daily conidium dissemination which was one of the important component in the epidemy of purple blotch disease was influenced by the temperature, relative humidity, and the wind velocity. Conidium dissemination was occurred both during the day and night and at 10.00 am-14.00 pm when the air temperature and wind velocity were high with low relative humidity, conidium dissemination was maximum. On the other hand, at 22.00 pm – 02.00 am there was minimum conidium dissemination.The result on the effects of leaf wetness period on disease intensity suggested that high humidity was not the only factor caused disease infection but it has to be supported by the presence of thin film water on the leaf surface at least for 4 hours since the attachment of the conidia on the leaves.Key words: purple blotch, climatic factor, Alternaria porri, shallot
Prevalensi Nematoda Parasit pada Pertanaman Pisang di Daerah Istimewa Yogyakarta Indarti, Siwi; Rahayu TP, Bambang; Subandiyah, Siti; Indarti, Lilis
Jurnal Perlindungan Tanaman Indonesia Vol 17, No 1 (2011)
Publisher : Universitas Gadjah Mada

Show Abstract | Download Original | Original Source | Check in Google Scholar | DOI: 10.22146/jpti.9397

Abstract

A study to determine the prevalence and distribution of plant parasitic nematodes associated with banana was undertaken in banana growing areas at four districts (Bantul, Gunung Kidul, Kulon Progo, and Sleman) of Yogyakarta Special Province. Seven genera of plant parasitic nematodes were found on these area: Criconemoides, Helicotylenchus, Hoplolaimus, Meloidogyne, Pratylenchus, Radopholus, and Rotylenchulus. Genera Hoplolaimus and Meloidogyne were distributed at all districts and occurring in soil and root samples of banana cultivars, Ambon, Kepok, Koja, Klutuk, Raja, Tanduk, and Uter, respectively. Four genera, Hoplolaimus, Meloidogyne, Pratylenchus, and Radopholus were dominant and were found with high level of population than the others on each district. Pratylenchus mostly was found on banana cv Kepok with average population 348,2–2057,3 nematodes on total samples of 5g banana root and 100g soil. Penelitian untuk mengetahui keberadaan dan sebaran nematoda parasit tumbuhan yang menyerang tanaman pisang dilakukan dengan survei di empat daerah kabupaten (Bantul, Gunung Kidul, Kulon Progo, dan Sleman) yang berada di wilayah Daerah Istimewa Yogyakarta. Ditemukan tujuh genera nematoda parasit yang menyerang pisang yaitu: Criconemoides, Helicotylenchus, Hoplolaimus, Meloidogyne, Pratylenchus, Radopholus, dan Rotylenchulus. Genera Hoplolaimus dan Meloidogyne tersebar merata di semua daerah dan didapatkan pada masing-masing sampel akar dan tanah pada hampir semua kultivar pisang: Ambon, Kepok, Koja, Klutuk, Raja, Tanduk, dan Uter. Empat genera yang dominan dengan tingkat populasi tinggi di setiap daerah adalah Hoplolaimus, Meloidogyne, Pratylenchus, dan Radopholus. Nematoda Pratylenchus banyak ditemukan pada pisang kultivar Kepok dengan rata-rata populasi 348,2– 2057,3 nematoda pada setiap total sampel 5 g akar dan 100 g tanah.  
Pemurnian dan Deteksi Serologi Patchouli Mottle Virus pada Tanaman Nilam Hartono, Sedyo; Subandiyah, Siti; Munawarti, Aminatun; Mastuti, Retno; Indriani, Serafinah
Jurnal Perlindungan Tanaman Indonesia Vol 12, No 2 (2006)
Publisher : Universitas Gadjah Mada

Show Abstract | Download Original | Original Source | Check in Google Scholar | DOI: 10.22146/jpti.12044

Abstract

Patchouli mottle virus (PatMoV) is the most severe disease pathogen and causes substantial losses in many patchouli-producing regions in Indonesia. Serological detection tool  for the disease was developed in this  research. Virus isolation was conducted on Chenopodium amaraticolor resulted on  the homogenous local lesions 6 days after  inoculation. Virus  purification was obtained from 200g inoculated leaves resulted on 2 ml virus solution with the concentration of 1 mg/ml. Polyclonal antibodies were produced on rabbits. Harvested antiserum was used for further virus detection by Indirect-Enzyme linked Immunosorbent assay (I-ELISA) and dot-immunobanding assay (DIBA) techniques. The antibodies were  positively  reacted with purified  viruses, infected field collection of patchouli, and  inoculated C. amaranticolor. On the other hand un-inoculated C. amaranticolor samples and healthy patchouli generated from tissue cultures gave negative reaction with the antibodies. This is the first report of cheap practical antibody production for PatMoV detection in Indonesia.
Ekstraksi DNA Phytophthora palmivora Isolat Kelapa Priyatmojo, Achmadi; Subandiyah, Siti
Jurnal Perlindungan Tanaman Indonesia Vol 2, No 1 (1996)
Publisher : Universitas Gadjah Mada

Show Abstract | Download Original | Original Source | Check in Google Scholar | DOI: 10.22146/jpti.9358

Abstract

Coconut isolate of Phytophthora palmivora DNA was extracted from sporangium and mycelium using SDS and Proteinase-K method. Sporangium and mycelium were harvested from liquid and solid culture of Potato Dextrose medium.The result of the study showed that the DNA extracted from sporangium and mycelium of 8 days solid culture was seen as distinct band. On the other hand, the dNA from sporangium and mycelium of 24 days old liquid culture was fragmented with smear band appearance.Key words: DNA, Phytophthora palmivora
Karakterisasi dan Deteksi Cepat Bakteri Penyebab Penyakit Darah pada Pisang Edy, Nur; Subandiyah, Siti; Sumardiyono, Christanti; Widada, Jaka
Jurnal Perlindungan Tanaman Indonesia Vol 17, No 1 (2011)
Publisher : Universitas Gadjah Mada

Show Abstract | Download Original | Original Source | Check in Google Scholar | DOI: 10.22146/jpti.9390

Abstract

Blood disease of banana is one of the most serious banana disease in Indonesia. Although the disease has became the subject of quarantine it eventually spread and found in most provinces in Indonesia. The aim of this research were to identify the blood disease bacterium (BDB) using morphological observation, biochemical assay, pathogenicity testing of hosts range using infectivity titration and rapid detection by Polymerase Chain Reaction (PCR). The results showed that the blood disease bacterium could be differentiated from Ralstonia solanacearum race 2, the causal agent of Moko disease and R. solanacearum tobacco isolates. BDB isolates were not able to hydrolyze gelatin, Tween 80, starch, and were not able to produce nitrite from nitrate. They were only able to produce acid from galactose and glycerol. The pathogenicity test indicated that the BDB was only able to infect the banana/plantain and was not able to infect tomato, eggplant, and chili. Rapid detection using PCR method showed that the 121F/R primers was able to amplify the BDB genome and was not able to amplify the genome of R. solanacearum tobacco isolates.Penyakit darah pada pisang masih merupakan kendala utama dalam budidaya pisang di Indonesia. Walaupun patogen penyakit darah sudah merupakan OPT karantina, namun saat ini penyakit sudah tersebar di seluruh provinsi di Indonesia. Penelitian ini bertujuan untuk mengidentifikasi bakteri penyebab penyakit darah dengan karakterisasi morfologi, biokimia, kisaran inang, dengan infectivity titration dan deteksi cepat menggunakan PCR. Hasil penelitian menunjukkan bahwa bakteri penyakit darah (BDB) dapat dibedakan dengan Ralstonia solanacearum ras 2, penyebab penyakit Moko dan R. solanacearum isolat tembakau. Isolat BDB tidak dapat menghidrolisis gelatin, Tween 80, pati dan tidak dapat menghasilkan nitrit dari nitrat. Bakteri ini hanya menghasilkan asam dari galaktosa dan gliserol. Hasil uji patogenisitas menunjukkan bahwa bakteri penyakit darah (BDB) hanya dapat menginfeksi pisang dan plantain dan tidak dapat menginfeksi tomat, terung dan cabai. Deteksi cepat dengan PCR menunjukkan bahwa primer 121F/R dapat mengamplifikasi genom bakteri penyakit darah (BDB) dan tidak dapat mengamplifikasi R. solanacearum isolat tembakau. 
Pengaruh Jenis Mangsa dan Suhu pada Perkembangan Menochilus sexmaculatus Fabricius (Coleoptera: Coccinellidae) dan Peranannya dalam Pengendalian Diaphorina citri Kuwayama (Hemiptera: Psyllidae) Ramadhan, Tris Haris; Trisyono, Y. Andi; Mahrub, Eddy; Wijonarko, Arman; Subandiyah, Siti; Beattie, George Andrew Charles
Jurnal Perlindungan Tanaman Indonesia Vol 14, No 1 (2008)
Publisher : Universitas Gadjah Mada

Show Abstract | Download Original | Original Source | Check in Google Scholar | DOI: 10.22146/jpti.11874

Abstract

Diaphorina citri Kuwayama (Hemiptera: Psyllidae) is the vector of citrus greening (Huanglongbing) bacterium and the most serious impediment to citrus culture. Classical biological control of this psyllid vector should contribute to suppress their population. This research was conducted to determine the performance of Menochilus sexmaculatus Fabricius (Coleoptera: Coccinellidae) when they were fed with D. citri. The larval performance index of M. sexmaculatus on D. citri compared with Aphis craccivora Koch (Hemiptera: Aphididae) diet was 1.3.M. sexmaculatus fed with D. citri had lower fitness than those fed with A. craccivora as shown by longer larval stadium, lower adult dry weight, less number of egg produced and lower percentage of egg hatched. M. sexmaculatus grew best at the temperature of 27oC. Employing the exclusion procedure under field condition,M. sexmaculatus could reduce the population of D. citri up to 90%. These findings showed that theM. sexmaculatus could be a potential predator in reducing D. citri, particularly when the more preferred prey A. craccivora was not present. Diaphorina citri Kuwayama (Hemiptera: Psyllidae) merupakan kelompok Psyllid yang menularkan penyebab penyakit Huanglongbing yang sangat berbahaya pada tanaman jeruk. Pengendalian hayati klasik telah banyak memberikan kontribusi dalam pengendalian di lapangan. Penelitian ini dilakukan untuk melihat penampilan Menochilus sexmaculatus Fabricius (Coleoptera: Coccinellidae) dengan pakan D. citri. Indeks penampilan larva M. sexmaculatus dengan pakan D. citri dibandingkan dengan Aphis craccivora Koch (Hemiptera: Aphididae) adalah 1,3. M. sexmaculatus yang diberi pakan D. citri menunjukkan penurunan kebugaran dibandingkan jika diberi pakan A. craccivora seperti yang ditunjukkan dengan stadium larva lebih lama, penurunan berat kering serangga dewasa, telur yang dihasilkan lebih sedikit, dan penurunan jumlah telur yang menetas. Menggunakan metode eksklusi pada kondisi lapangan, M. sexmaculatus mampu menurunkan populasi D. citri sampai 90%. M. sexmaculatus berkembang baik pada suhu 27oC. Hasil penelitian ini menunjukkan bahwa M. sexmaculatus berpotensi untuk menurunkan populasi D. citri, terutama jika inang utamanya A. craccivora tidak ditemukan.
Peningkatan Ketahanan Tanaman Pisang Kepok Kuning terhadap Penyakit Darah melalui Variasi Somaklonal dan Simbiosis Endofitik Wibowo, Arif; Joko, Tri; Subandiyah, Siti; Mariska, Ika; Supriyati, Yati; Suryadi, Yadi; Roostika, Ika
Jurnal Perlindungan Tanaman Indonesia Vol 16, No 1 (2010)
Publisher : Universitas Gadjah Mada

Show Abstract | Download Original | Original Source | Check in Google Scholar | DOI: 10.22146/jpti.11738

Abstract

One of the obstacles that was encountered in the banana cultivation is blood disease. Blood diseases is caused by Ralstonia solanacearum that is subsequently revised to become blood disease bacteria (BDB). Until now the control of banana blood disease is very difficult. Control of banana blood disease with chemical injections and soil treatment is not effective. This study was aimed to obtain Kepok Kuning cultivar banana seedlings which was resistant towards blood disease obtained from in vitro selection by using BDB growing filtrate and induced resistance by inoculation of antagonistic endophytic bacteria.The observation of Kepok Kuning banana explants treated with various concentrations of BDB growing filtrate showed that the percentage of living explants decreased to 83.33% when the BDB growing filtrate concentration increased to 15%. Treatment of banana explants with BDB growing filtrate also affected the number of roots, shoots, and leaves. Treatment with a single antagonistic endophytic bacteria suppressed the intensity of banana blood disease to 0% in comparison with the mixture of antagonistic endophytic bacteria if Kepok Kuning banana explants were not treated with BDB growing filtrate. When Kepok Kuning banana explants were treated with BDB growing filtrate, the intensity of banana blood disease suppressed to 0% after the high concentration of BDB growing filtrate and the antagonistic endophytic bacteria mixture were applied. Salah satu kendala yang dihadapi dalam usaha budidaya pisang adalah adanya penyakit darah. Penyakit darah disebabkan oleh bakteri Ralstonia solanacearumyang selanjutnya direvisimenjadi bakteri darah pisang (Blood Disease Bacteria orBDB). Sampai saat ini pengendalian penyakit darah pisang sangatsukar dilakukan. Pengendalian penyakit darah dengan suntikan bahan kimia dan perlakuan tanah tidak efektif untuk diaplikasikan. Penelitian ini bertujuan untukmendapatkan bibit pisang kultivar Kepok Kuning yang tahan penyakit darah yang diperoleh dariseleksi in vitro dengan menggunakan filtrat pertumbuhan BDB dan induksi ketahanan melalui inokulasi jasad renik endofitik yang bersifat antagonis. Hasil pengamatan terhadap eksplan pisang Kepok Kuning yang diperlakukan dengan berbagai konsentrasi filtrat BDB menunjukkan bahwa pada eksplan yang diperlakukan dengan filtrat pertumbuhan BDB, persentase tumbuhnya akan berkurang hingga 83,33% jika konsentrasifiltrat pertumbuhanBDBmencapai 15%. Selain itu perlakuan planlet pisang dengan filtrat pertumbuhan BDB akan mempengaruhi jumlah akar, tunas, dan daun. Perlakuan dengan jasad renik endofitik antagonis secara tunggal mampu menekan intensitas penyakit darah hingga 0% jika dibandingkan dengan perlakuan campuran apabila sebelumnya eksplan pisang Kepok Kuning tidak diperlakukan dengan filtrat pertumbuhan BDB. Apabila sebelumnya planlet pisangKepokKuning diperlakukan dengan filtrat pertumbuhan BDB maka mampu menekan intensitas penyakit darah hingga 0% jika konsentrasi filtrat pertumbuhanBDBsemakin tinggi dan diperlakuan campuran jasad renik endofitik.