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Agrobacterium-Mediated Stable Transformation of Medicinal Plant Andrographis paniculata Callus Expressing β-glucuronidase (GUS) Gene Marwani, Erly; Tangapo, Agustina; Dwivany, Fenny Martha
Indonesian Journal of Biotechnology Vol 18, No 2 (2013)
Publisher : Universitas Gadjah Mada

Show Abstract | Download Original | Original Source | Check in Google Scholar | Full PDF (406.629 KB)

Abstract

This study was carried out to establish a stable genetic transformation in callus culture of Andrographispaniculata mediated by Agrobacterium tumefaciens. The leaf disks of A. paniculata were infected with A. tumefaciensLBA4404 carrying a binary vector pCAMBIA1304 that contain β-glucuronidase (GUS) and hygromycinphosphotransferase (hpt) genes. The infection was conducted by dipping method for one hour, followed byco-cultivation in the dark for three days. To examine transient GUS expression, the co-cultivated leaf disks wereassayed for β-glucuronidase activity and to obtain stable transformed callus, the co-cultivated leaf disks wereselected on the callus induction medium which contain 20 mg/l hygromycin for selection. The transformedcallus was periodically subcultured every three weeks into the fresh selection medium over the 15 weeksperiod. To test a stable transformation, the callus was subjected to PCR analysis for GUS gene detection. Theresults indicated that the co-cultivated leaf disks expressed GUS activity and proliferated to produce callus onthe selective medium. Analysis of PCR on the transformed callus indicated the presence 976 bp fragment thatconfi rmed the presence of β-glucuronidase gene. These fi ndings imply that the β-glucuronidase was stablyintegrated into A. paniculata callus culture.Keywords: Andrographis paniculata, Agrobacterium tumefaciens, andrographollide, transformed callus,β-glucuronidase gene.
PENGARUH KERAPATAN AGROBACTERIUM TERHADAP EFISIENSI TRANSFORMASI PADA PISANG CAVENDISH (Musa acuminata colla) Apriyani, Ria Khoirunnisa; Esyanti, Rizkita Rachmi; Dwivany, Fenny Martha
BIOTIKA Jurnal Ilmiah Biologi Vol 17, No 1 (2019): BIOTIKA JUNI 2019
Publisher : Universitas Padjadjaran

Show Abstract | Download Original | Original Source | Check in Google Scholar | DOI: 10.24198/bjib.v17i1.21898

Abstract

Transformasi genetik melalui perantara Agrobacterium tumefaciens merupakan salah satu carauntuk meningkatkan kualitas buah Pisang Cavendish (Musa acuminata colla (AAA Group))yang memiliki susunan gen triploid. Efisiensi transformasi melalui A. tumefaciens terhadapembrio Pisang Cavendish dilakukan dengan mengoptimasi berbagai kerapatan optik kulturbakteri. A. tumefaciens, yaitu strain AGL1 dan GV3101 (yang masing-masing membawaplasmid ganda pBI121), mengandung gen gusA (pengode enzim β-Glucuronidase) digunakandalam proses transformasi ini. Perbedaan kerapatan optik (0,6; 0,8; 1,0) suspensi bakteri digunakan untuk proses transformasi. Persentase efisiensi transformasi dihitung berdasarkanjumlah embrio pisang yang positif pada uji GUS dibagi dengan jumlah total embrio pisang yangdiuji, dikali 100%. Efisiensi transformasi tertinggi diperoleh dari embrio pisang yangdiinokulasi oleh A.tumefaciens strain AGL1/pBI121 pada OD600nm = 1 (96%) dan efisiensitransformasi terendah diperoleh dari embrio pisang yang diinokulasi oleh A.tumefaciens strainGV3101/pBI121 pada OD600nm = 0.6 (40%).
Agrobacterium-Mediated Stable Transformation of Medicinal Plant Andrographis paniculata Callus Expressing β-glucuronidase (GUS) Gene Erly Marwani; Agustina Tangapo; Fenny Martha Dwivany
Indonesian Journal of Biotechnology Vol 18, No 2 (2013)
Publisher : Universitas Gadjah Mada

Show Abstract | Download Original | Original Source | Check in Google Scholar | Full PDF (406.629 KB) | DOI: 10.22146/ijbiotech.7873

Abstract

This study was carried out to establish a stable genetic transformation in callus culture of Andrographispaniculata mediated by Agrobacterium tumefaciens. The leaf disks of A. paniculata were infected with A. tumefaciensLBA4404 carrying a binary vector pCAMBIA1304 that contain β-glucuronidase (GUS) and hygromycinphosphotransferase (hpt) genes. The infection was conducted by dipping method for one hour, followed byco-cultivation in the dark for three days. To examine transient GUS expression, the co-cultivated leaf disks wereassayed for β-glucuronidase activity and to obtain stable transformed callus, the co-cultivated leaf disks wereselected on the callus induction medium which contain 20 mg/l hygromycin for selection. The transformedcallus was periodically subcultured every three weeks into the fresh selection medium over the 15 weeksperiod. To test a stable transformation, the callus was subjected to PCR analysis for GUS gene detection. Theresults indicated that the co-cultivated leaf disks expressed GUS activity and proliferated to produce callus onthe selective medium. Analysis of PCR on the transformed callus indicated the presence 976 bp fragment thatconfi rmed the presence of β-glucuronidase gene. These fi ndings imply that the β-glucuronidase was stably integrated into A. paniculata callus culture.Keywords: Andrographis paniculata, Agrobacterium tumefaciens, andrographollide, transformed callus,β-glucuronidase gene.
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.
Isolation of MA-ACS Gene Family and Expression Study of MA-ACS1 Gene in Musa acuminata Cultivar Pisang Ambon Lumut LISTYA UTAMI KARMAWAN; SONY SUHANDONO; FENNY MARTHA DWIVANY
HAYATI Journal of Biosciences Vol. 16 No. 1 (2009): March 2009
Publisher : Bogor Agricultural University, Indonesia

Show Abstract | Download Original | Original Source | Check in Google Scholar | Full PDF (440.641 KB) | DOI: 10.4308/hjb.16.1.35

Abstract

Musa acuminata cultivar pisang ambon lumut is a native climacteric fruit from Indonesia. Climacteric fruit ripening process is triggered by the gaseous plant hormone ethylene. The rate limiting enzyme involved in ethylene biosynthesis is ACC synthase (ACS) which is encoded by ACS gene family. The objective of this study is to identify MA-ACS gene family in M. acuminata cultivar pisang ambon lumut and to study the MA-ACS1 gene expression. The result showed that there were nine M. acuminata ACS gene family members called MA-ACS1–9. Two of them (MA-ACS1 and MA-ACS2) were assessed using reverse transcriptase PCR (RT-PCR) for gene expression study and it was only MA-ACS1 correlated with fruit ripening. The MA-ACS1 gene fragment has been successfully isolated and characterized and it has three introns, four exons, and one stop codon. It also shows highest homology with MACS1 gene from M. acuminata cultivar Hsian Jien Chiao (GenBank accession number AF056164). Expression analysis of MA-ACS1 using quantitative PCR (qPCR) showed that MA-ACS1 gene expression increased significantly in the third day, reached maximum at the fifth day, and then decreased in the seventh day after harvesting. The qPCR expression analysis result correlated with the result of physical analysis during fruit ripening. Key words: pisang ambon lumut, MA-ACS gene family, gene characterization, gene expression, quantitative PCR (qPCR)