Judul: Construction and Subcloning of the GOx-Xho Gene Encoding Glucose Oksidase Enzyme into the Plasmid pPICZα
Abstrak: Glukosa oksidase (GOx; EC 1.1.3.4) adalah enzim yang mengoksidasi Dglukosa menjadi D-glukono-d-lakton dan menghasilkan hidrogen peroksida. Aktivitas enzim tersebut pada organisme asalnya, yaitu Aspergillus niger IPBCC 08.610, diketahui lebih tinggi secara intraseluler dibanding ektraselular sehingga upaya memproduksi GOx secara masif kurang efisien. Salah satu solusi dari permasalahan tersebut adalah dengan metode alternatif, yaitu produksi enzim GOx secara rekombinan. Penelitian ini bertujuan mengonstruksi ulang fragmen gen GOx dengan menambah situs restriksi XhoI menjadi GOx-Xho, mensubkloning gen tersebut ke dalam plasmid pPICZα, dan mentransformasi plasmid rekombinan ke dalam sel Escherichia coli DH5α. Konstruksi gen GOx dilakukan menggunakan metode PCR dalam dua tahap. Ligasi gen GOx dengan plasmid pPICZα dilakukan dengan dua perlakuan, yaitu dengan rasio 1:1 dan 1:3 antara plasmid dan gen. Hasil penelitian menunjukkan perbandingan plasmid dan gen yang optimal dalam subkloning adalah 1:3. Hasil subkloning berhasil ditransformasi ke dalam sel E. coli DH5α dan sel pembawa plasmid rekombinan pPICZα-GOx-Xho berhasil diperoleh.
Keyword: glucose oxidase enzyme, recombinant plasmid, reconstruction gene, subcloning gene

Judul: Construction of Glucose Oxidase Gene Without Sequence That Encodes Spacer Peptide and Subcloning into pPICZα Plasmid
Abstrak: Glukosa oksidase (GOx) (EC 1.1.3.4) berfungsi mengkatalisis reaksi oksidasi dari β-D-glukosa menjadi D-glukono-δ-lakton dan hidrogen peroksida (H2O2). Enzim ini diketahui dihasilkan oleh beberapa jenis kapang. Aspergillus niger diketahui memproduksi enzim GOx yang lebih stabil dan digunakan sebagai sumber enzim GOx yang tersedia secara komersial. Isolat lokal A. niger IPBCC 08.610 yang digunakan pada penelitian ini diketahui menghasilkan kadar enzim GOx intraseluler yang lebih tinggi dibandingkan ekstraselulernya. Hal ini menjadi suatu kendala karena ekstraksi enzim intraseluler membutuhkan proses hilir yang lebih rumit. Produksi enzim GOx secara ekstraseluler pada Pichia pastoris dapat mengatasi masalah ini dan secara bersamaan dapat meningkatkan hasil produksi GOx rekombinan. Penelitian ini bertujuan mendapatkan gen penyandi GOx tanpa sinyal peptida aslinya yang akan disubklon ke dalam plasmid pPICZα pada sisi restriksi enzim XhoI dan XbaI agar terfusi secara in frame dengan sinyal sekresi α-mating factor (α-MF) tanpa menyertakan sekuen penyandi spacer peptida EAEA pada ujung-5’. Gen GOx diamplifikasi melalui dua tahapan reaksi PCR dari plasmid pGEM®-T Easy‒GOx. Amplikon gen GOx yang diperoleh berukuran 1775 pb yang terdiri atas situs restriksi XhoI (ujung-5’), sekuen penyandi gen GOx, dan situs restriksi XbaI (ujung-3’). Subkloning amplikon ini ke dalam plasmid pPICZα diharapkan akan menghasilkan plasmid rekombinan dengan ukuran 5269 pb., Glucose oxidase (GOx) (EC 1.1.3.4) catalyzes the oxidation of β-D-glucose to form D-glucono-δ-lactone and hydrogen peroxide (H2O2). The enzyme is well known to be produced from several species of fungis. Aspergillus niger is known to produced a more stable GOx and used as source of commercially available GOx. A local isolate of A. niger IPBCC 08.610 used in this study produced higher level of intracellular GOx enzyme than its extracellular. This fact presents a problem because intracellular enzyme extraction will need a more complicated downstream processing. Production of extracellular GOx in Pichia pastoris would overcome this problem and concomitanly could increase the yield of recombinant GOx production. This study aims to obtain the GOx gene without the original signal peptide to be subcloned into the pPICZα plasmid, fused in frame with the secretion signal α-mating factor (α-MF) without the sequences that encodes spacer peptide EAEA at its 5’-end. The GOx gene construction was carried out using a two steps PCR amplification method from pGEM®-T Easy‒GOx plasmid. The GOx amplicon is 1775 bps in length consisting of the XhoI restriction site (5’-end), the GOx gene encoding sequence, and the XbaI restriction site (3’-end). Subcloning this amplicon into the pPICZα plasmid is expected to produce a recombinant plasmid that have a length of 5269 bps.
Keyword: Amplification, Aspergillus niger, glucose oxidase, pPICZα, spacer peptide

Judul: Thermal Characteristics and Dynamics of Tropical Cyclone over Western North Pacific Ocean: A Case Study of Typhoon Songda (Chedeng) 2011
Abstrak: The frequent occasion of Tropical Cyclone (TC) over Western North Pacific (WNP) Ocean lies from 10˚N to 26˚N and 121˚E to 170˚E. Sources of TC development come from warm ocean temperature where latent heat is released during condensation processes and subsequently concentrated in the boundary layer. Relative humidity at 850-700 hPa is responsible to produce the most intense storm. Meanwhile, weak vertical wind shear between 850-200 hPa is one of the dynamical factors that lead to the development of TC. Pressure level of 850-200 hPa is chosen to minimize the thermodynamic effect in the lower troposphere. Typhoon Songda (Chedeng in the Philippines) or TSC lasted from May 18th to 30th, 2011 and strengthened into powerful Super Typhoon stage between May 24th and 27th. Long curving track and the exhibition of all sequences of TC life cycle, TSC is then considered as an ideal case of TC. The objective of the study with case of TSC is to investigate the role of thermal characteristics and dynamics on TC intensity. The research is carried out by using Advanced Research of Weather Research and Forecasting model (WRF ARW model) version 3.3. Two domains are employed in the model. The first domain situate from 02˚N to 40˚N and 118˚E to 149˚E and cover spatial resolution () of 33.1 km. The second domain situate from 09˚N to 27˚N and 119˚E to 132˚E with of 11 km. Timestep of the model () is 120 seconds. The model is supported by the following physical parameterization, i.e., Yonsei University Scheme (YSU) in the boundary layer, Kain-Fritsch scheme (KF) in the cumulus, and WRF Single-Moment 3-class scheme in the microphysics. Initial meteorological condition prior to the formation of the cyclone is generated using WRF ARW standard-release model. Sensitivity of the model output is obtained from a selected single-parameter change scenarios, namely, 10 % to 20 % decrease and increase of wind shear initial data at 850-200 hPa, and by 10 % to 20 % decrease of relative humidity initial data at 850-700 hPa. The result showed that the negative correlations between wind shear and intensity are found in wind shear scenarios. Wind shears of 10 ms-1 which lead to strengthening of the wind intensity in the early period of Typhoon development are found in wind shear scenarios. The effect of wind shear changes on wind intensity is statistically significant in the 20 % wind shear scenarios. During mature stage, weak wind intensity occurred when wind shear increase as given in the WRF standard-release and the 20 % decreasing scenario. Different situation is found in the 20 % increasing scenario where wind shear tend to weak and lead to strong wind intensity at this stage. Meanwhile, the reduction of moisture supply at 850-700 hPa resulting from 20 % decreasing relative humidity play a major role in the weakening of wind intensity and reduces mechanical energy of the cycle process as much as 300 J kg-1.
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