EKSPLORASI BAKTERI TERMOHALOFILIK POTENSIAL PENGHASIL L-ASPARAGINASE SEBAGAI ANTIKANKER DI SUMBER AIR PANAS WAWOLESEA

ABSTRAK Tujuan penelitian ini adalah untuk memperoleh isolat bakteri termohalofilik penghasil enzim L-aparaginase dari sumber air panas Wawolesea. Bakteri termohalofilik penghasil L-asparaginase asal sumber air panas Wawolesea diperoleh dengan tahapan: survey dan pengukuran parameter lingkungan sumber air panas Wawolesea; isolasi bakteri pada media NA (Nutrient Agar) dan seleksi bakteri penghasil enzim L-asparaginase pada media M-9. Hasil isolasi menunjukkan adanya 52 isolat bakteri termohalofilik dan 14 isolat diantaranya mampu menghasilkan L-asparaginase. Kata Kunci: Bakteri Termohalofilik, L-Asparaginase, Sumber Air Panas Wawolesea ABSTRACT  The objective of this study was to obtain isolates of thermohalophilic bacteria producing L-asparaginase enzyme from Wawolesea hot spring. L-asparaginase producing thermohalophilic bacteria from the Wawolesea hot spring are obtained by steps; survey and environmental parameters measurement of Wawolesea hot spring, isolation of bacteria on NA (Nutrient agar) medium and selection of L-asparaginase producing bacteria on M-9 medium. The isolation results showed 52 isolates of thermohalophilic bacteria and 14 isolates of them capable of producing L-asparaginase. Keywords: Thermohalophilic bacteria, L-asparaginase, Wawolesea hot sprin

Karakterisasi Morfologi Phytophthora sp. Asal Buah Kakao Desa Olo-oloho, Kabupaten Konawe, Sulawesi Tenggara

This study aimed to determine the morphological characteristics of Phytophthora sp. isolated from cocoa fruits from Olo-oloho Village, Konawe Regency, Southeast Sulawesi. Isolation of Phytophthora sp. carried out by the point method using V4 (Vegetable Juice Agar) media incubated at 27ºC for 24 hours. Morphological characterization of Phytophthora sp. included characterization of colony morphology and cell morphology. The results showed that the colony morphological characteristics were white colonies, cotton-like textures, the uneven edge of the colony, zoning and radial lines. The morphological characteristics of the cell had asexual spores in the form of sporangium and chlamydospores, hyphae are not aseptic, greenish-black zoospores, zoospores are round and double-flagged, and have sporangiophores. Keywords: Phytophthora sp., colony morphology, cell morpholog

Isolasi dan Pengklonan Fragmen CDNA Gen Penyandi H+-ATPase Membran Plasma dari Melastoma Malabathricum L.

Melastoma malabathricum L. grows well in acid soil with high level of soluble aluminum. One of the important proteins in the detoxifying acid and aluminum stress is a plasma membrane H+ -ATPase protein encoded by PMA gene. The objective of this research was to isolate and clone the cDNA fragment of MmPMA encoding plasma membrane H+ -ATPase from M. malabathricum L. By reverse transcription, total cDNA had been synthesized from the total RNA as template. The fragment of MmPMA cDNA had been successfully isolated by PCR by using total cDNA as template and PMA primer designed from conserved region for corresponding gene. This fragment had been successfully inserted into pGEM-T Easy and the recombinant plasmid was successfully introduced into E. coli DH5". Nucleotide sequence analysis showed that the length of MmPMA fragment is 806 bp encoding 268 amino acids. Local alignment analysis based on nucleotide of mRNA showed that MmPMA fragment was 81% identical to part of PMA of Sesbania rostrata, Juglans regia, and Prunus persica. Based on deduced amino acid sequence, MmPMA was 94% identical to part of PMA of Juglans regia; 93% to PMA of S. rostrata, and Arabidopsis thaliana. MmPMA fragment has phosphorylation intermediate domain (DKTGT) and ATP binding domain (KGAP, DPPR, MITGD, and GDGVN)

Isolasi dan Pengklonan Gen Penyandi H+-ATPase Membran Plasma dari Melastoma Malabathricum L.

Melastoma malabathricum L. is an Al-accumulating plant that grows well in acidic soils with high level of soluble aluminum in the tropics. One of the important proteins in the detoxifying Al stress is a plasma membrane H+-ATPase, a most abundant protein on the plasma membrane, encoded by PMA gene. The objective of this research was to isolate and characterize the gene encoding plasma membrane H+-ATPase from M. malabathricum L. Full length cDNA of MmPMA had been successfully isolated through a gradual isolation of the gene. The 5' end and middle part of the MmPMA gene had been successfully isolated by PCR by using total cDNA as template and pma primers designed from some plants, while the 3' end of Mmpma had been isolated by 3' RACE. The parts of the gene had been successfully joined by PCR. The joining product was successfully inserted into pGEM-T Easy and the recombinant plasmid was successfully introduced into E. coli DH5α. Nucleotide sequence analysis showed that the length of MmPMA coding sequence was 2,871 bp encoding 956 amino acids with molecular weight of 105.29 kDa and a predicted pI value of 6.84. Local alignment analysis based on nucleotide of mRNA showed that MmPMA is 82% identical to pma Vitis vinifera; 81% to pma Juglans regia, pma Populus trichocarpa, pma Sesbania rostrata, and pma Prunus persica and 80% to pma Lycopersicon esculentum. Based on deduced amino acid sequence, MmPMA is 94% identical to PMA Vitis vinifera and PMA Juglans regia; 93% to PMA Populus trichocarpa; 92% to PMA Vicia faba, Lycopersicon esculentum, and Arabidopsis thaliana, AHA4. MmPMA has 10 transmembrane domains, 4 cytoplasm loops, 6 functional domains and 3 autoregulatory domains

DETEKSI MOLEKULER BAKTERI Escherichia coli SEBAGAI PENYEBAB PENYAKIT DIARE DENGAN MENGGUNAKAN TEHNIK PCR

Penelitian ini dilakukan untuk mendeteksi secara molekuler bakteri Escherichia coli sebagai penyebab penyakit diare dengan menggunakan tehnik PCR. DNA genomik Escherichia coli diekstraksi menggunakan metode boiling, kemudian diamplifikasi menggunakan primer 16E1 dan 16E2. Hasil PCR positif Escherichia coli ditunjukkan dengan adanya pita fragmen DNA pada ukuran sekitar 584 pasang basa pada gel elektroforesis. Dalam penelitian ini sampel yang digunakan adalah sebanyak delapan titik dengan distribusi empat titik dari sampel air sumur bor dan empat dari air sumur galian. Dari delapan titik sampel, ada satu sampel yang terdeteksi secara molekuler yaitu mengandung Escherichia coli yaitu dengan kode sampel ASB 3yang dibuktikan dengan adanya pita DNA. Hasil penelitian menunjukkan bahwa metode PCR dapat mendeteksi Escherichia coli secara spesifik dan lebih cepat. Kata Kunci : Air, Bakteri Escherichia coli, Isolasi DNA, Tehnik PCR

RNAi dari Fragmen 3’UTR Gen Penyandi H+ -ATPase Membran Plasma Melastoma malabathricum L. dapat Menghambat Pertumbuhan Tanaman Tersebut

The RNA silencing technique is an effective tool to examine the biological function of the target mRNA in plants. The recent development of GATEWAYTM cloning technology makes it easy to construct the RNAi vectors with trigger sequences and to analyze the function of a target gene. The objective of this research was to construct RNAi including the 3’UTR fragment of the gene coding plasma membrane H+-ATPase from Melastoma malabathricumL., 3’UTRMmpma. RNAi vector had been successfully constructed using GATEWAYTM cloning technology with the 3’UTRMmpma was used as double-stranded RNA (dsRNA) trigger sequence, pENTRTM/D-TOPO®as entry vector, and pANDA plasmid as destination vector. RNAi had been successfully introduced into M. malabathricumL. mediated by A. tumefaciensEHA101 to analyze the function of Mmpma gene in the detoxifying Al stress. Based on the test of transgenic plants tolerance to Al stress showed that in the nutrient solution including 3.2 mM Al (AlCl3.6H2O), the transgenic plants underwent growth suppression especially roots and leaves, whereas non-transgenic plants underwent growth normally. It showed that suppression of Mmpmagene expression by RNAi to M. malabathricumL. caused the plant became sensitive to Al.Keywords: 3’UTRMmpma, A. tumefaciens, Al stress, RNAi vecto

Properties and Application of Edible Modified Bacterial Cellulose Film Based Sago Liquid Waste as Food Packaging

Bacterial cellulose (BC) based on sago liquid waste has been developed to be used as food packaging. This study investigated the physicochemical and mechanical properties of modified BC film and its application as food packaging. The modified BC film performed carboxymethyl cellulose (CMC) as a stabilizer and glycerol as a plasticizer. Films were prepared by casting technique using BC as the primary material and composites with various concentrations of CMC and glycerol (0.5%, 1%, and 1.5%, v/v). BC film was applied as the packaging of meat sausage, and the quality of meat sausage was measured based on weight loss, moisture content, pH, protein content, and total microbial count. The addition of CMC and glycerol influences the physical and mechanical properties of BC composites film. The best mechanical properties of edible BC film were collected by adding 1% CMC and 1% glycerol with a tensile strength of 17.47 MPa, elongation at a break of 25.60%, and Young’s modulus of 6.54 GPa. FTIR analysis showed the characteristic bands of BC, and the addition of CMC and glycerol slightly changed the FTIR spectrum of the composites. The utilization of modified BC-based sago liquid waste film as the packaging of meat sausage could maintain sausage quality during 6 days of storage at room temperature. Therefore, edible BC film has the potential to be used as food packaging. Keywords: bacterial cellulose; edible film; food packaging; sago liquid wast

Isolasi dan Pengklonan Gen Penyandi H+-ATPase Membran Plasma dari Melastoma malabathricum L.

ABSTRACT Melastoma malabathricum L. is an Al-accumulating plant that grows well in acidic soils with high level of soluble aluminum in the tropics. One of the important proteins in the detoxifying Al stress is a plasma membrane H+-ATPase, a most abundant protein on the plasma membrane, encoded by PMA gene. The objective of this research was to isolate and characterize the gene encoding plasma membrane H+-ATPase from M. malabathricum L. Full length cDNA of MmPMA had been successfully isolated through a gradual isolation of the gene. The 5’ end and middle part of the MmPMA gene had been successfully isolated by PCR by using total cDNA as template and pma primers designed from some plants, while the 3’ end of Mmpma had been isolated by 3’ RACE. The parts of the gene had been successfully joined by PCR. The joining product was successfully inserted into pGEM-T Easy and the recombinant plasmid was successfully introduced into E. coli DH5α. Nucleotide sequence analysis showed that the length of MmPMA coding sequence was 2,871 bp encoding 956 amino acids with molecular weight of 105.29 kDa and a predicted pI value of 6.84. Local alignment analysis based on nucleotide of mRNA showed that MmPMA is 82% identical to pma Vitis vinifera; 81% to pma Juglans regia, pma Populus trichocarpa, pma Sesbania rostrata, and pma Prunus persica and 80% to pma Lycopersicon esculentum. Based on deduced amino acid sequence, MmPMA is 94% identical to PMA Vitis vinifera and PMA Juglans regia; 93% to PMA Populus trichocarpa; 92% to PMA Vicia faba, Lycopersicon esculentum, and Arabidopsis thaliana, AHA4. MmPMA has 10 transmembrane domains, 4 cytoplasm loops, 6 functional domains and 3 autoregulatory domains.Keywords: aluminum, cDNA, MmPMA, PCR, RAC

Isolasi dan Pengklonan Fragmen cDNA Gen Penyandi H+-ATPase Membran Plasma dari Melastoma malabathricum L.

Melastoma malabathricum L. grows well in acid soil with high level of soluble aluminum. One of the important proteins in the detoxifying acid and aluminum stress is a plasma membrane H+  -ATPase protein encoded by PMA gene. The objective of this research was to isolate and clone the cDNA fragment of MmPMA encoding plasma membrane H+ -ATPase from M. malabathricum L. By reverse transcription, total cDNA had been synthesized from the total RNA as template. The fragment of MmPMA  cDNA  had been successfully isolated by PCR by using total cDNA  as  template and PMA primer designed from conserved region for corresponding gene. This fragment had been successfully inserted into pGEM-T Easy and the recombinant plasmid was successfully introduced into E. coli DH5". Nucleotide sequence analysis showed that the length of MmPMA fragment is 806 bp encoding 268 amino acids. Local alignment analysis based on nucleotide of mRNA showed that MmPMA fragment was 81% identical to part of PMA of Sesbania rostrata, Juglans regia, and Prunus persica. Based on deduced amino acid sequence, MmPMA was 94% identical to part of PMA of Juglans regia; 93% to PMA of S. rostrata, and Arabidopsis thaliana. MmPMA fragment has phosphorylation intermediate domain (DKTGT) and ATP binding domain (KGAP, DPPR, MITGD, and GDGVN).   Keywords: isolation, Melastoma malabathricum L., MmPMA fragment, sequencin