Judul: Optimasi Transformasi Eksplan Tebu menggunakan Gen P5CS melalui Agrobacterium tumefaciens
Abstrak: Genetic transformation is one of the attempt in generating sugarcane (Saccharum officinarum L.) that are tolerant to drought stress. P5CS gene has a role in prolin biosynthesis, the amino acid that accumulating in water stress condition. Transformation of a P5CS gene construct into plant cells in conjunction with regeneration for transgenic plantlets should develop sugarcane tolerant to drought stress. The aim of this research is to generate drought-tolerant sugarcane through optimization transformation which includes the strain of Agrobacterium tumefaciens, a good source of sugarcane eksplant and varieties. This study was done through verification of IBRIEC (Indonesian Biotechnology Research Institute for Estate Crops) P5CS gene collection A. Tumefaciens transformation (Wang 2006), sugarcane explants transformation (Sain 1994) and sugarcane transformant regeneration. Verification of P5CS gene collection was PCR analysed by specific primers P5CS full lenght. PCR and gel electrophoresis showed a single band agarose and the same size of 2.4 kb with positive control. It is convinced that the P5CS gene has been isolated and transformed into A. tumefaciens. IBRIEC collection P5CS genes in plasmid pBI121 were arranged in a recombinant construct of PBI-P5CS transformed into A. tumefaciens strain GV3101, LBA4404 and AGL1. Recombinant constructs PBI- P5CS from Vigna aconitifolia (V. aconitifolia) successfully transformed into cell A. tumefaciens were grown with appropriate antibiotics. Strain GV3101 and LBA4404 used 50 ppm kanamycin and rifampicin respectively, while antibiotics were added in AGL1 strain is 50 ppm kanamycin, rifampicin and ampicillin respectively. The existence of P5CS gene in A. tumefaciens was tested by observing the growth of transformed colonies in media selection and colony PCR analysis using specific primers P5CS full lenght. The results showed that A. tumefaciens transformants were grown in selection media. PCR and gel electrophoresis analysis showed a single band agarose and same size of 2.4 kb with positive control. This showed the success of the transformation A. tumefaciens and used to explants sugar cane transformation. Sugarcane explants used in this research was included calli grown in solid medium, embryogenic calli and somatic embryos with MS medium derived from Temporary Immersion System (TIS) culture with three varieties Kidang Kencana, PS 881 and PS 891. Co-cultivation medium used solid MS medium with acetosyringon in room temperature for two days. Transgenes selection was done in solid MS with antibiotics 50 ppm kanamycin and 500 ppm cefotaxime. Explants were not transformed cultured in MS media as positive control negative control while planted in the media selection. Observation of sugarcane transforman in selection media was calculated the percentage growth in 16th weeks and 32th weeks. The existence of P5CS gene in transgenic shoots was tested with GUS histochemical assay, PCR using conserved regions of specific primers P5CS and NPTII. GUS histochemical test for the presence of P5CS gene constructs showed positive results in the presence of blue staining. DNA amplification showed that expected bent size at 1.2 kb and 700 bp for primers NPTII. The results of this examination proved that both transgenes were inserted in the sugarcane genomes. The test results are stable sugarcane transformants through histochemical GUS test and PCR analysis showed stable transformants sugarcane. Growth
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Judul: Transformasi Genetik Padi (Oryza sativa L.) dengan Gen PaCS Penyandi Sitrat Sintase Menggunakan Perantara Agrobacterium tumefaciens
Abstrak: Peningkatan produksi padi (Oryza sativa L.) dapat dilakukan dengan menerapkan intensifikasi dan ekstensifikasi pertanian. Penerapan ekstensifikasi pertanian terkendala oleh berkurangnya lahan pertanian akibat konversi lahan pertanian menjadi lahan perindustrian, perumahan, dan perkantoran. Pemanfaatan lahan marginal terutama tanah jenis ultisol dapat menjadi alternatif untuk menerapkan ekstensifikasi pertanian. Tetapi, tanah ultisol bersifat masam dan berpotensi mengandung Al yang dapat menghambat pertumbuhan tanaman. Perakitan tanaman padi yang toleran cekaman aluminium dapat dilakukan dengan mengintroduksi gen yang berkaitan dengan aktifitas toleransi terhadap cekaman aluminum. Sitrat sintase (CS) terlibat dalam biosintesis asam sitrat. Sitrat memiliki kemampuan mengkelat Al paling kuat diantara asam organik lain. Pada beberapa tanaman, ekspresi gen CS meningkatkan toleransinya terhadap cekaman Al. Penelitian ini bertujuan untuk mentransformasi genetik padi dengan gen penyandi sitrat sintase menggunakan perantara Agrobacterium tumefaciens. Transformasi genetik dilakukan dengan perendaman selama 10 menit kalus embriogenik umur 3 minggu yang berasal dari embrio biji padi kultivar Kasalath dan Nipponbare dalam suspensi Agrobacterium tumefaciens strain LBA4404 yang mengandung plasmid pMSH1-PaCS dengan gen penanda seleksi higromisin. Kokultivasi dilakukan selama tiga hari pada kondisi gelap. Seleksi kalus transgenik menggunakan media yang mengandung higromisin 20 mg/L. Kalus yang resisten higromisin diregenerasikan di media 2N6R yang mengandung 5 mg/L kinetin. Subkultur dilakukan setiap 2 minggu sekali pada media yang sama. Eksplan kalus yang digunakan untuk transfomasi pada padi kultivar Kasalath sebanyak 135 kalus dan Nipponbare sebanyak 240 kalus. Kalus kultivar Kasalath dan Nipponbare yang tahan higromisin sebanyak 2 % dan 35,8% dari jumlah eksplan kalus yang ditransformasi. Terdapat 13 kalus dari kultivar Nipponbare yang beregenerasi menghasilkan tunas dan masing-masing kalus menghasilkan tanaman padi transgenik putatif, sedangkan kalus padi kultivar Kasalath yang diregenerasikan tidak ada yang menghasilkan tunas. Penambahan antibiotik cefotaksim sebanyak 200 mg/L pada media regenerasi dapat menurunkan daya regenerasi dari kedua kultivar. Konfirmasi gen penanda seleksi higromisin dengan primer 35S-F dan hpt-R pada sepuluh nomor kultivar Nipponbare transgenik putatif menghasilkan amplikon dengan ukuran 1100 pb. Konfirmasi gen PaCS dengan menggunakan primer forward 35S-F dan PaCS-R menghasilkan amplikon dengan ukuran 1630 pb untuk padi kultivar Nipponbare transgenik nomor 10, sedangkan untuk padi kultivar Nipponbare transgenik nomor 1 sampai 9 menghasilkan amplikon 900 pb.
Keyword: Agrobacterium tumefacien, Oryza sativa L., sitrat sintase, transformasi genetik

Judul: Hamiltonian formulation for linear waves motion in two layers fluid
Abstrak: The surface wave could be considered as a wave that separates two fluids, namely water and air. Based on this assumption, it is introduced the interfacial wave, a wave between two layers of fluid with different density. The formulation of interfacial waves motion begins with deriving the base equation of irrotational ideal fluid. Furthermore, according to irrotational fluid assumption, the base equation can be stated in velocity potential. In this derivation, the fluids domain is assumed to be restricted by rigid lid boundary conditions, both at the upper and lower limit. Therefore, the interfacial waves motion can be explained in a hamiltonian formulation. In the hamiltonian formulation, total energy is defined as the sum of kinetic and potential energy. The hamiltonian system is obtained from reduction of kinetic energy by using the Dirichlet Neumann Operator. The resulted kinetic energy equation is nonlinear. Therefore, this form is linearized by first part of the Taylor expansion. This linearization gives a dispersion relation of linear wave. Based on this dispersion relation, the phase speed of the linear wave depends on the density ratio of the two layers fluid.
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