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TRANSFORMASI MENGGUNAKAN Agrobacterium tumefaciens PADA TUNAS DAUN Kalanchoe mortagei DAN Kalanchoe daigremontiana 1 DAN 2 Dewanto, Hamami Alfasani; Suhandono, Sony
Chimica et Natura Acta Vol 4, No 2 (2016)
Publisher : Departemen Kimia

Show Abstract | Download Original | Original Source | Check in Google Scholar | Full PDF (355.284 KB) | DOI: 10.24198/cna.v4.n2.10679

Abstract

Cocor bebek adalah tumbuhan sukulen yang mampu memproduksi tunas adventif (reproduksi vegetatif) pada tepian daunnya. Kemampuan reproduksi vegetatif ini menghasilkan tanaman yang sama dalam waktu yang singkat, sehingga memungkinkan untuk dijadikan sebagai bioreaktor protein rekombinan. Transformasi dilakukan menggunakan Agrobacterium tumefaciens pada tunas daun cocor bebek dari spesies Kalanchoe mortagei dan Kalanchoe daigremontiana. Optimasi dilakukan mencakup: galur A. tumefaciens, kerapatan optis dari kultur A. Tumefaciens, konsentrasi acetosyringone, teknik ko-kultivasi, pH medium dan komposisi medium ko-kultivasi. Hasil optimasi transformasi secara transien menunjukan bahwa perbedaan galur A. tumefaciens, kerapatan optis, konsentrasi acetosyringone menghasilkan ekspresi transien yang relatif sama secara kualitatif. Berdasarkan uji GUS teknik ko-kultivasi dengan infiltrasi vakum dan pH medium 5,5 menghasilkan ekspresi transien lebih baik dibandingkan dengan perendaman dan pH medium 7,0. Medium ko-kultivasi M9 menghasilkan ekspresi transien yang lebih baik dibandingkan dengan medium ½MS0. Tunas daun K. daigremontiana 2 menunjukan ekspresi transien yang lebih baik dibandingkan K. mortagei dan K. daigremontiana 1.
Construction of Binary Vector With Wound Inducible Promoter for Hbsag Expression: Development of Plant-Based Edible Hepatitis B Vaccine from Indonesian Isolate Suhandono, Sony; Rachman, Ernawati A.Giri; Zainuddin, Ima M.; Utari, Putri Dwi; Supraba, Apsari; Estiati, Amy
ANNALES BOGORIENSES Vol 11, No 1 (2007): Annales Bogorienses
Publisher : Research Center for Biotechnology - Indonesian Institute of Sciences (LIPI)

Show Abstract | Download Original | Original Source | Check in Google Scholar | DOI: 10.1234/22

Abstract

Hepatitis B is a serious infectious disease In the third world countries including Indonesia . Vaccination is  the most effective way to prevent the spread of the disease;  therefore the demand for HBV vaccine is high.  In order to produce more vaccine at lower cost, transgenic plant can be chosen to express the vaccine with the above criteria. Several researches were successfully producing transgenic plants expressing HBsAg that formed virus-like particles and  induced  immune response  in  human.  However. HBsAg expression  in  transgenic plant needs to be  improved especially on gene  expression control system. Here, we describe the construction of HBsAg .  structural gene under the control of wound  inducible promoter, MeEFl promoter from  manihot esculenta Crantz. The HBsAg gene was amplified using PCR from HBV genome isolated from an  Indonesian patient. The gene was subsequently fused with VSPaS signal peptide, which targeted the reticulum endoplasm of plant cell . The construct was cloned into binary expression vector for Agrobacterium plant Transformation  in  near future.Keywords: HBsAg. VSPaS signal peptide and MeEFI promoter
Analysis of Microsatellite Allele That potential as Resistance Marker of Giant Gouramy (Osphronemus gouramy Lac.) to Aeromonas hydrophila Kusumawardhani, Meirina Kartika; Kusumawaty, Diah; Suhandono, Sony
Aquacultura Indonesiana Vol 15, No 1 (2014): Volume 15 Issue 1 Year 2014
Publisher : Indonesian Aquaculture Society (MAI)

Show Abstract | Download Original | Original Source | Check in Google Scholar | Full PDF (157.124 KB) | DOI: 10.21534/ai.v15i1.30

Abstract

Giant gouramy is one of economical important freshwater fish. However, the production of the fish was declined because of a Motile Aeromonas Septicemia (MAS) disease. Giant gouramy with MAS disease caused an ulcer on their skin, and the worst case infection may cause death. The symptoms is vary widely depends on fish resistance to the disease. The aim of this study is to analyze microsatellite alleles that potential as a resistance marker against Aeromonas hydrophila. In previous studies, eleven microsatellite loci were isolated from giant gouramy genome. These loci were tested on DNA from resistant and susceptible giant gouramy that had been treated with Aeromonas hydrophila. Resistant giant gouramy was the gouramy that survived at least 50 days post-infection and ssusceptible giant gouramy was the gouramy that died before 50 days post- infection. Three of eleven microsatellite loci were found with unique alleles that appeared only in resistant giant gouramy and potential as resistance marker, which is the 342 bp allele at GE 1.9 locus with (GCA)10 (ACA) 6 motif, the 262 bp allele at GE 2.4 locus with (GCA) 6 (GGA)10 motif, and the 244 bp allele at GE 1.4 locus with (GCT) 9 (TA) 6 motif. All three loci were used to scan 48 giant gouramy broodstock from Tasikmalaya, Singaparna, and Sukabumi. Amplification of the microsatellite loci was performed by PCR with M13 tail sequence in forward primer and fluorescence dye. Successfully amplified alleles were analyzed with GeneAlEx 6.5 software. As a result, fragment 262 bp and 244 bp that contained in 43.75% of 48 gouramy broodstock predicted have a potency as resistance marker to Aeromonas hydrophila. For further study, microsatellite motifs have to be screened in gouramy progeny, because microsatellites are inhereted in Mendelian traits.
THE CONCEPTUAL CHANGE ASSESSMENT BASED ON ESSAY QUESTIONS IN CASE STUDY OF DNA/RNA AND INTRON TOPICS Kristianti, Tati; Widodo, Ari; Suhandono, Sony
Jurnal Penelitian Pendidikan IPA Vol 4, No 1 (2019): Juni 2019
Publisher : Perkumpulan Pendidik IPA Indonesia (PPII)

Show Abstract | Download Original | Original Source | Check in Google Scholar | DOI: 10.26740/jppipa.v4n1.p31-37

Abstract

Most of the study on conceptual change was analysed using multiple-choice, true/false and true/false-reason type of questions. However, these questions are unable to reveal the variation in understanding of the university students. In this study, we developed a new conceptual change assessment based on essay questions. Here, student responses on DNA and intron term are presented as case studies in order to show the application of this assessment. The assessment is able to classify the various degree of understanding in university students such as construction, revision, complementation, static and disorientation. Our findings in university students studying DNA term showed that of 70 university students, 34% (construction), 36% (revision), 21% (complementation), 4% (static) and 5% (disorientation). On intron term finds construction (4%), revision (14%), static (44%) and disorientation (38%). Overall analysis using various categories of understanding reveals that DNA term is easier to understand by the university students than the intron term. This assessment is useful to evaluate the conceptual change level of university students in the class. Therefore, the assessment might also be useful to evaluate various teaching strategies and other topics.Keywords: conceptual change, essay, assessment, DNA, Intron
Sekuens Gen Protein Kapsid Mayor L1 Human Papilomavirus 16 dari Isolat Klinik Asal Bandung Pradita, Anandayu; Sahiratmadja, Edhyana; Suhandono, Sony; Susanto, Herman
Majalah Kedokteran Bandung Vol 46, No 3 (2014)
Publisher : Faculty of Medicine, Universitas Padjadjaran

Show Abstract | Download Original | Original Source | Check in Google Scholar | Full PDF (976.647 KB)

Abstract

Kanker serviks disebabkan oleh infeksi kronik human papillomavirus (HPV) dengan genotipe HPV-16 sebagai HPV tersering yang menginfeksi epitel serviks. Protein penyelubung virus yang disebut kapsid mayor (L1) mempunyai peranan penting dalam menginfeksi epitel serviks. Tujuan penelitian untuk mengisolasi dan menganalisis sekuens gen L1 HPV-16. Pengetahuan mengenai sekuens gen L1 dapat memberikan informasi yang berguna, salah satunya yaitu untuk pengembangan vaksin. Pada studi ini, deoxyribonucleic acid (DNA) virus diekstraksi dari sediaan biopsi pasien kanker serviks yang diambil pada bulan Juni sampai Oktober 2010 di Kebidanan dan Kandungan RS Dr. Hasan Sadikin Bandung. Gen diamplifikasi dengan polymerase chain reaction menggunakan primer spesifik. Infeksi HPV-16 pada jaringan kanker dikonfirmasi dengan menggunakan kit komersial untuk tes genotipe HPV. Fragmen L1 kemudian diklon dan diinsersikan ke dalam pJET1.2/L1-16, kemudian dipotong dengan enzim BamHI dan BgIII untuk kemudian divalidasi dan disekuensing. Hasil sekuensing menunjukkan amplikon gen L1 HPV-16 sebesar 1.595 pasang basa. Analisis dari dua amplikon gen L1 HPV-16 menggunakan software BIOEDIT dan Basic Local Alignment Search Tool menunjukkan kesamaan ho mologi 99% dan 97% dengan sekuens L1 HPV-16 asal Thailand yang terregistrasi pada GenBank. Simpulan, telah dilakukan kloning sekuens gen L1 HPV-16 dari dua isolat klinik Bandung. Hasil kloning HPV-16 pada penelitian ini memberikan informasi tentang variasi sekuens yang perlu dipertimbangkan bagi pengembangan vaksin terutama bagi daerah spesifik seperti penduduk asal Indonesia.Kata kunci: Human papillomavirus, kanker serviks, gen L1 HPV-16 Sequence of Human Papilomavirus 16 Major Capsid L1 Gene from Clinical Isolates in BandungCervical cancer is strongly associated with chronic human papillomavirus (HPV) infection. HPV-16 is the most prevalent genotype infecting cervical epithelium. The major coat protein of viral particle (L1) plays a key role in the infection process. Our study aimed to isolate the HPV-16 L1 gene and analyze its sequence. Samples used were samples collected from the Department of Obstetrics and Gynaecology, Dr. Hasan Sadikin General Hospital, Bandung during the period of June to October 2010. In this study, the HPV-16 L1 sequence was analyzed from the viral deoxyribonucleic acid (DNA) extracted from biopsy sample of cervical cancer patient biopsy samples.The HPV-16 L1 amplification was performed using the polymerase chain reaction with specific primer. The HPV infection in the cervical tissue was confirmed by commercial HPV genotyping test. The L1 fragment was cloned into plasmid and the insert of the recombinant clone pJET1.2/L1-16 was digested using BamHI and BgIII. The amplicon result showed HPV-16 L1 gene with a length of 1.595 base pairs. The sequence analysis of two samples using software BIOEDIT dan Basic Local Alignment Search Tool revealed a high level of sequence similarity to L1 HPV-16 from Thailand (99% and 97%) as registered in GenBank. In conclusion, the L1 HPV-16 gene from Bandung isolates revealed variations from published sequence. Knowledge on L1 gene sequence may give additional information to the development of vaccine. Further study on vaccine development is currently ongoing using this HPV-16 clone that may be specific to Indonesian population. Key words: Cervical cancer, human papillomavirus, L1 HPV-16 DOI: 10.15395/mkb.v46n3.317
Segmentasi Citra Digital Objek Hasil Pengamatan In Situ Localization Gen gfp pada Tanaman Transforman Atqiya, Firas; Ihsani, Nisa; Sholahuddin, Muhammad Rizqi; Dwivany, Fenny Martha; Suhandono, Sony
Jurnal Pendidikan Multimedia (Edsence) Volume 1 No 2 (Desember 2019)
Publisher : Universitas Pendidikan Indonesia (UPI)

Show Abstract | Download Original | Original Source | Check in Google Scholar | DOI: 10.17509/edsence.v1i2.21575

Abstract

Penelitian berbasis biomolekuler membutuhkan beragam penggunaan perangkat lunak pengolah data. Salah satunya yaitu kebutuhan perangkat lunak yang mampu mengolah data citra digital pada proses segmentasi warna. Dalam penelitian biomolekuler, segmentasi warna dapat digunakan untuk menganalisis pendaran warna hijau sebagai hasil ekspresi gen gfp. Gen pelapor ini banyak digunakan dalam proses rekayasa genetik tumbuhan maupun hewan yaitu: memonitor ekspresi gen, in situ localization, biosensor, physiological indicators, dan studi interaksi protein. Sinar UV pada panjang gelombang eksitasi 450-490 nm dapat diserap dan diemisikan oleh molekul protein GFP sebagai warna hijau. Adanya pendaran hijau tersebut diharapkan hanya muncul sebagai penanda terekspresinya gen gfp. Namun demikian, pada sampel tumbuhan terkandung senyawa metabolit sekunder yang dapat menyerap dan mengemisikan sinar UV sebagai warna hijau. Adanya warna hijau selain hasil ekspresi gen gfp ini tentunya dapat menyebabkan hasil analisis in situ localization menjadi bias. Oleh karena itu, diperlukan teknik pengolahan citra digital yang mampu memilah warna hijau hasil ekspresi gen gfp dan warna hijau dari emisi senyawa metabolit tumbuhan. Tujuan  penelitian  ini  adalah untuk memisahkan objek hasil ekspresi gen gfp pada citra digital jagung transforman dengan warna hijau yang diemisikan oleh senyawa metabolit sekunder jagung menggunakan pengolahan citra digital. Proses yang digunakan adalah color filtering, thresholding, dan Canny edge detection. Hasil penelitian yang diperoleh berupa citra yang mengandung citra objek hasil ekspresi gen gfp yang telah tersegmentasi pada sayatan melintang akar jagung transforman.
THE CONCEPTUAL CHANGE ASSESSMENT BASED ON ESSAY QUESTIONS IN CASE STUDY OF DNA/RNA AND INTRON TOPICS Kristianti, Tati; Widodo, Ari; Suhandono, Sony
Jurnal Penelitian Pendidikan IPA Vol 4, No 1 (2019): Juni 2019
Publisher : Perkumpulan Pendidik IPA Indonesia (PPII)

Show Abstract | Download Original | Original Source | Check in Google Scholar | DOI: 10.26740/jppipa.v4n1.p31-37

Abstract

Most of the study on conceptual change was analysed using multiple-choice, true/false and true/false-reason type of questions. However, these questions are unable to reveal the variation in understanding of the university students. In this study, we developed a new conceptual change assessment based on essay questions. Here, student responses on DNA and intron term are presented as case studies in order to show the application of this assessment. The assessment is able to classify the various degree of understanding in university students such as construction, revision, complementation, static and disorientation. Our findings in university students studying DNA term showed that of 70 university students, 34% (construction), 36% (revision), 21% (complementation), 4% (static) and 5% (disorientation). On intron term finds construction (4%), revision (14%), static (44%) and disorientation (38%). Overall analysis using various categories of understanding reveals that DNA term is easier to understand by the university students than the intron term. This assessment is useful to evaluate the conceptual change level of university students in the class. Therefore, the assessment might also be useful to evaluate various teaching strategies and other topics.Keywords: conceptual change, essay, assessment, DNA, Intron
Cloning and Characterization of P5CS1 and P5CS2 Genes from Saccharum officinarum L. under Drought Stress Iskandar, Hayati Minarsih; Widyaningrum, Dwiyantari; Suhandono, Sony
Journal of Tropical Crop Science Vol 1 No 1 (2014): Journal of Tropical Crop Science
Publisher : Department of Agronomy and Horticulture, IPB University

Show Abstract | Download Original | Original Source | Check in Google Scholar | Full PDF (512.987 KB) | DOI: 10.29244/jtcs.1.1.23-30

Abstract

Increasing world sugar demand might be fulfilled with land extensification which include the use of dry area. Development of drought tolerance and high productivity sugarcane variety could be achieved  by plant genetic engineering. Under drought condition, proline will be accumulated and functioned as an osmoregulator in plant cells. ∆1-pyrroline-5-carboxylate synthase (P5CS) is one of the important enzymes in proline biosynthesis.  This enzyme is encoded by P5CS gene family. We cloned two homologous P5CS genes from sugarcane,  SoP5CS1 (Accession Number : KF178299) and SoP5CS2 (Accession Number : KF178300), which encode 729 and 716 amino acid polypeptides. The identity between these two genes was 74% based on nucleotide sequences. The SoP5CS1 gene had 98% identity with SbP5CS1 (Accession Number : GQ377719.1) and SoP5CS2 had 99% identity with SaP5CS (Accession Number : EF113257.1). In this experiment, sugarcane plantlets  were exposed to medium containing PEG 6000 (40%) for 12, 24, 48, and 72 hours. Proline concentration was measured after treatment and genes expression were analyzed by real time-qPCR. The results showed that the proline concentration was increased 12 folds (9.8 umol.g-1) after 48-hours stress treatment. The highest expression of SoP5CS1 occured at 24-hours treatment with approximately 16 times from plant without PEG (control plant) and decreased gradually at 48 and 72 hours treatment. The highest expression of SoP5CS2 occured at 24-hours drought stress with approximately 3.6 folds compared to control. In drought treatment, the expression level SoP5CS1 was higher than SoP5CS2 and has increased significantly at 12-hours treatment. It is suggested that the SoP5CS1 gene contributes more significantly to the production of proline during drought stress than SoP5CS2. Hence, SoP5CS1 could potentialy be used as a marker to screen sugarcane variety for drought tolerance and for the development of transgenic plant tolerant to drought.Keywords: cloning, drought, expression, P5CS, sugarcane
The transformation of dbr2 and p19 Genes into Artemisia annua L. by Agrobacterium tumefaciens Mediation: Metabolites Analysis Sy, Khairunnisa; Kristianti, Tati; Suhandono, Sony; Elfahmi, Elfahmi
Indonesian Journal of Pharmaceutical Science and Technology 2024: Suppl. 6, no. 3 (The 3rd Mandala Waluya International Conference on Pharmaceutical Science and
Publisher : Indonesian Journal of Pharmaceutical Science and Technology

Show Abstract | Download Original | Original Source | Check in Google Scholar | DOI: 10.24198/ijpst.v6i3.54535

Abstract

The low level of Artemisinin in Artemisia annua L. is associated with its biosynthetic pathway. Genetic engineering offers a potential solution to increase its metabolites. The research objectives were to transform the double bond reductase (dbr2), one of the critical genes in the biosynthesis of Artemisinin, and p19, an anti-silencing gene, into A. annua L. by the mediation of Agrobacterium tumefaciens and assess their impact on metabolites production. The dbr2 and p19 genes were verified by analyzing PCR products. Plasmids pCAMBIA1303-DBR2-P19 were transformed into A. tumefaciens and subsequently introduced into the leaves from tissue culture of A. annua L, using vacuum infiltration and assessed by GUS assay. UPLC-ESI-MS/MS measured artemisinic acid (AA), dihydroartemisinic acid (DHAA), and Artemisinin contents. The transformation in A. tumefaciens was successful using a freeze-thaw method. The tissue culture of A. annua L. has been infected by A. tumefaciens using vacuum infiltration. Based on the GUS histochemical assay, the gene has been successfully inserted in the leaves with an efficiency of 48.2%. The result from UPLC-ESI-MS/MS showed that the level of Artemisinin in the transformation sample with and without pCAMBIA-dbr2-p19 was detected but not quantifiable while in wild-type leaves were quantified at 0.008% in fresh weight. The AA and DHAA were not detectable but only in the wild type. The transformation was successful, but the quantification of DHAA and AA was unsuccessful because of the low quantity in the samples.
A robust in planta Agrobacterium‐mediated transformation in red chili (Capsicum annuum L.) Hamdani, Anti Damayanti; Nugroho, Syarul; Esyanti, Rizkita Rachmi; Faizal, Ahmad; Suhandono, Sony
Indonesian Journal of Biotechnology Vol 29, No 4 (2024)
Publisher : Universitas Gadjah Mada

Show Abstract | Download Original | Original Source | Check in Google Scholar | DOI: 10.22146/ijbiotech.94653

Abstract

Plant improvement through in vitro culture and genetic engineering is a significant aspect of breeding programs aimed at producing disease‐resistant cultivars of disease‐prone red chili (Capsicum annuum L.). However, the Capsicum genus is recalcitrant to genetic transformation and in vitro regeneration. Moreover, developing a universal transformation protocol is difficult due to its highly genotype‐dependent nature. Therefore, this study aimed to develop an Agrobacterium‐mediated in planta transformation method applicable to various red chili cultivars. Two open‐pollinated varieties, Tanjung 2 and Ciko, were subjected to transformation. The young seedlings were immersed in transformation medium containing Agrobacterium tumefaciens strain GV3101 harboring the binary vector pCAMBIA1301, which carries the β‐glucuronidase (GUS) gene. GUS histochemical analysis revealed that all the primary transformants of Tanjung 2 and Ciko were identified as chimeric. The average staining in the body of the seedlings was 88.63 + 26.33% in Tanjung 2, and 90.65 + 16.77% in the Ciko variety. More than 50% of the seedlings continued to express GUS in their shoot areas 10 days after Agrobacterium infection, indicating the possibility of transgene inheritance in the following generation. The in planta transformation approach is notably genotype independent, making it a promising standard transformation protocol for different red chili varieties.