MAELITA RAMDANI MOEIS
Research Group of Genetics and Molecular Biotechnology, School of Life Sciences and Technology, Institute of Technology Bandung

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Analisis DNA Bacillus sp. BAC4 Hasil Polymerase Chain Reaction (PCR) dengan Primer-Primer yang Dirancang dari Gen pga

Jurnal Kedokteran Maranatha Vol 8, No 1 (2008)
Publisher : Universitas Kristen Maranatha

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Abstract

Bacillus sp. BAC4 DNA was amplified using PCR method employing primers designed from pga genes of 5 bacteria (Arthrobacter viscosus, Bacillus megaterium, Escherichia coli, Kluyvera citrophila, and Providentia rettgeri). Analysis  of DNA base direct sequencing of PCR products showed the highest homology of Bacillus sp. BAC4 DNA with Bacillus subtilis DNA at the region from citG to yirG.  This fact states that primers designed from pga genes of 5 bacteria has not been successful in amplifying Bacillus sp. BAC4 pga gene, and that Bacillus sp.  BAC4 pga gene is predicted to be quite different from the existing data of pga genes available at Genbank.  Analysis of DNA base direct sequencing amplified using the pga primers shows the highest homology to the DNA of Bacillus subtilis.

Cloning and Expression of Endoglucanase Gene from Thermophilic Bacteria Bacillus sp. RP1

Microbiology Indonesia Vol 8, No 4 (2014): December 2014
Publisher : Indonesian Society for microbiology

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An endoglucanase gene from glycoside hydrolase family 5, had been isolated from Bacillus sp. RP1 and cloned into Escherichia coli. The cloned gene comprised the promoter, coding sequence and terminator of the gene.  This gene encoded a protein with 499 amino acid residues (Mr=55.2 kDa) with a typical Bacillus signal peptide. The recombinant endoglucanase (EG) had optimum activity at pH 5.0 and 50 °C. The recombinant EG was expressed in the extracellular, intracellular, and periplasmic fractions with the highest total activity (60.15%) in the intracellular fraction, measured at three hours after isopropyl-β-Dthiogalactopyranoside (IPTG) induction. Three hours after the addition of 1% carboxymethyl cellulose (CMC), there was a two-fold increase in intracellular EG specific activity compared to the uninduced cells. Three hours after the addition of 1 mM IPTG, 1% glucose, 1% galactose or 1% cellobiose the intracellular EG specific activity decreased compared to the uninduced cells.

Face Shape Variation Among Sundanese People from Western Java, Indonesia

HAYATI Journal of Biosciences Vol 22, No 1 (2015): January 2015
Publisher : Bogor Agricultural University, Indonesia

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Abstract

The face is an important visual stimulus in daily life and each face identifies a particular person. The bone structure of the skull along with various soft tissues and coloration influence perception of the face. Facial averageness, and bilateral symmetry are the two most commonly used criterion of facial attractiveness, yet, both may be perceived differently based on hormonal status of the person observed. Facial perceptions may also differ according to cultural norms. In this research, we examined variations in face-shape among Sundanese male and female adults aged 18 to 40. We applied geometric-morphometric methods to analyze the landmark-based morphological variations in the frontal and lateral views of subjects’ faces. We identified five types of female frontal face views and four of male. We also identified five types each of female and male lateral face views. The trichion, gonion and gnathion were three most variable landmarks among the face views in our study, and highly determined the shape of the individuals’ faces. Multiple face type variation may refer to many categories of attractive faces since there is no exactly perfect category in the assessment of facial attractiveness by the viewers. Therefore, we believe that the configuration of facial features cannot constitute the sole visual criterion of facial attractiveness.

OPTIMASI PROSES UNTUK EKSPRESI GEN ENDOGLUKANASE DARI Bacillus sp. RP1 OLEH Escherichia coli BL21 (DE3)/ egc

Jurnal Bioteknologi & Biosains Indonesia (JBBI) Vol 5, No 1 (2018): June 2018
Publisher : Badan Pengkajian dan Penerapan Teknologi (BPPT)

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Process Optimization for Endoglucanase Gene Expression Derived from Bacillus sp. RP1 by Escherichia coli BL21 (DE3)/egcABSTRACTCellulases are one of the most used enzymes in industrial processes. In an effort to increase production, industries have developed strategies such as isolating new cellulase producing strains, genetic engineering and process optimization since the last 50 years. One endoglucanase producing strain, Bacillus sp. RP1 was isolated from hot springs. The ribosome binding site and coding sequence of the endoglucanase gene (egc) from Bacillus sp. RP1 was cloned into pGEM-T Easy. The recombinant plasmid was used to transform E. coli BL21 (DE3). Cloning was followed by process optimization. Medium composition was selected using Plackett-Burman design. The medium components tested were rice hull, molasses, ammonium chloride, urea and fishmeal. Rice hull and molasses were found to be the factors most influencing enzyme activity and dry cell weight, respectively. The next step involved Box-Behnken method and response surface methodology to optimize the responses against molasses concentration, rice hull concentration and fermentation time. The concentration intervals used to test were 1%, 5.5% and 10% while the fermentation time used were 24, 36 and 48 hours. The conditions which optimized both enzyme activity and dry cell weight were 7.45% molasses, 6.45% rice hull and 39.52 hours of fermentation.Keywords: Bacillus sp. RP1, E. coli BL21 (DE3), egc, Endoglucanase, optimization ABSTRAKSelulase adalah salah satu enzim yang banyak dimanfaatkan dalam berbagai industri. Sebagai upaya untuk memenuhi kebutuhan, 50 tahun terakhir dikembangkan beberapa strategi untuk meningkatkan produksi selulase yang mencakup rekayasa genetika dan optimasi proses. Karena itu, dilakukan kloning gen egc dan RBS yang berasal dari Bacillus sp. RP1 yang diisolasi dari sumber air panas ke dalam vektor pGEM-T Easy. E. coli BL21 (DE3) ditransformasikan dengan vektor yang mengandung gen egc tersebut. Setelah kloning, optimasi proses berupa desain medium turut dilakukan untuk mengoptimalkan ekspresi gen egc. Desain medium diawali dengan seleksi komposisi medium menggunakan metode Plackett-Burman. Komponen medium yang diuji adalah kulit beras, molase, amonium klorida, urea dan tepung ikan. Kulit beras dan molase diperoleh sebagai bahan yang paling berpengaruh terhadap aktivitas enzim dan berat kering sel. Tahap selanjutnya melibatkan metode statistik Box-Behnken dan metodologi respons permukaan yang bertujuan mengoptimalkan respons aktivitas enzim dan berat kering sel terhadap konsentrasi molase, konsentrasi kulit beras dan lama fermentasi. Konsentrasi yang diuji adalah 1%, 5,5% dan 10%, sedangkan lama fermentasi yang diuji adalah 24, 36 dan 48 jam. Konsentrasi optimal molase adalah 7,45% dan konsentrasi optimal kulit beras adalah 6,45% dengan lama fermentasi optimal 39,52 jam.Kata Kunci: Bacillus sp. RP1, E. coli BL21 (DE3), egc, Endoglukanase, optimasi

Expression and Purification of PhoR Sensor-Domain Histidine Kinase of Mycobacterium tuberculosis in Escherichia coli

Microbiology Indonesia Vol 9, No 2 (2015): June 2015
Publisher : Indonesian Society for microbiology

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Abstract

Globally, tuberculosis (TB) remains a leading cause of death. The emergence of multidrug-resistant strains (MDR-TB) and extensively drug-resistant strains (XDR-TB) has fuelled the discovery for novel drugs and drug targets for its successful and better treatment. One of the potential candidates for drug target is PhoR sensory protein histidine kinase, a part of the Two Component System (TCS) PhoP/PhoR in Mycobacterium tuberculosis (Mtb). This protein system was known for its role on regulating hundred of Mtb virulence factors, from genes for cell wall and lypid synthesis to genes for adaptation in human leukocyte and hypoxia response. Previous studies have successfully characterized, isolated, and cloned the putative sensory domain of PhoR protein gene into pRSET vector expression system. In this study, Escherichia coli was transformed with pRSET-SensPhoR and cultivated at 37oC under IPTG induction to express PhoR sensor-domain protein. Most of the proteins were overexpressed in the form of inclusion bodies.  Subsequent protein purification in Ni-NTA system under refolding condition on urea gradient was performed to isolate PhoR sensor-domain protein in soluble form. Arginine was supplemented in purified protein solution to prevent aggregation during long term storage.  While highly purified protein was acquired, small angle X-ray scattering (SAXS) analysis was conducted to obtain 3-dimensional (3D) protein structures in solution.    doi:10.5454/mi.9.2.1 

Transposon Insertion Phenomenon during Cloning of a Partial Fragment Derived from Metagenomic DNA Isolated from Deep-Sea Water and Sediment of Kawio Island, North Sulawesi

Squalen, Buletin Pascapanen dan Bioteknologi Kelautan dan Perikanan Vol 8, No 3 (2013): December 2013
Publisher : Research and Development Center for Marine and Fisheries Product Processing and Biotechnol

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Abstract

Transposon is well-known as mobile element found abundant both in prokaryote and eukaryote genomes. In bacteria, transposon (famous name of a transposable DNA) could jump from chromosome to plasmid and its contrary. One type of transposons in bacteria known as insertion sequence (IS), it does not contain any additional genes except a gene encoding transposase, an enzyme that correlated to transponsition activities. The finding of transposon insertion unfortunately found during cloning of a fragment derived from deep-sea metagenomic DNA in this research. In the initial, this research was aimed to clone and characterize the á-amylase encoded gene derived from metagenomic DNA isolated from deep-sea water and sediment of Kawio Island, North Sulawesi. Metagenomic DNA has been isolated from deep-sea water and sediment and by using Whole Genome Amplification (WGA) technique, the DNA it could be increased in quantities to 146,31 ng for each 1 ng of metagenomic DNA. A fragment of ~1000 bp in length was obtained by using touchdown PCR method. The presence of a transposon in this DNA fragment is proposed as a hypothesis for losing ~700 bp leaving just 310 bp cloned sequence. Analysis of sequencing result showed a highest similarity between this 310 bp partial fragment with a replication protein (Rep) encoded gene from Pseudomonas putida (Query Coverage: 88%; Max. Identity: 80%, Positive: 86%) and this protein is known to be involved in plasmid replication where transposase encoding genes known usually presence together with this gene (Rep gene) in a bacterial plasmid.