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IDENTIFIKASI POPULASI BAKTERI DALAM SPONS PENCUCI PIRING DENGAN METODE PCR-RFLP Shabarni Gaffar; Iman Permana Maksum; Euis Julaeha
Chimica et Natura Acta Vol 2, No 2 (2014)
Publisher : Departemen Kimia

Show Abstract | Download Original | Original Source | Check in Google Scholar | Full PDF (293.895 KB) | DOI: 10.24198/cna.v2.n2.9154

Abstract

Spons pencuci piring 200.000 kali lebih kotor dibanding dudukan toilet. Berbagai bakteri penyebab penyakit seperti Eschericia coli, Pseudomonas dan Staphylococcus, berkembang biak di permukaan yang basah. Selain itu, 500 ribu bakteri hidup di dalam saluran pembuangan bak cuci piring. Jika spons dalam kondisi tidak kering (lembab karena direndam), maka akan menjadi markas semua bakteri. Penelitian ini bertujuan untuk menentukan populasi bakteri yang terdapat pada spons cuci piring yang basah. Identifikasi dilakukan dengan metoda molekular berbasis DNA yaitu teknik PCR-RFLP gen 16s rDNA. Metoda PCR-RFLP merupakan genetik fingerprinting yang akurat, cepat dan sederhana. Koloni bakteri yang ditumbuhkan dari spons pencuci piring di lisis dan digunakan sebagai templat untuk PCR. Hasil PCR gen 16s rDNA yang berukuran 1400 pb, dipotong dengan enzim restriksi, Hinf1. Hasil penelitian menunjukkan terdapat empat pola pemotongan yang berbeda-beda. Pola yang berbeda ini menunjukkan terdapat empat populasi yang berbeda hidup pada spons basah. Kami mengidentifikasi bahwa salah satu mikroba yang tumbuh adalah E. coli yang merupakan bakteri patogen.
PHYLOGENETICS ANALYSIS OF MARINE AND COASTAL SPECIES USING 18S rRNA SEQUENCE Shabarni Gaffar -; Linawati Hardjito -; Endang Srieatimah -
Bionatura Vol 11, No 2 (2009): Bionatura Juli 2009
Publisher : Direktorat Sumber Daya Akademik dan Perpustakaan

Show Abstract | Download Original | Original Source | Check in Google Scholar | Full PDF (242.962 KB)

Abstract

Phylogenetics analysis using 18S rRNA sequence was done on some marine and coastal organisms that have been used as traditional medicine in Indonesia. Proper and accurate identification of coastal and marine organisms producing bioactive compound is important to support drug discovery. Ribosomal RNA has been described as one of most useful and most used for molecular chronometer. In this experiment a part of 18S rRNA gen was amplified using PCR employing 300F and 1400R primers. PCR product then was cloned using pGEM-T vector (Promega) and transformed into Eschericia coli. The recombinant Eschericia coli were sequenced by applying universal primer and internal primer. The alignment of sequenced product and phylogenetic analysis indicated that sample 1 closely related to Chaetomorpha crassa, sample 2 closely related to Ptilophora pinnatifida, sample 3 closely related to Flagellaria indica, and sample 4 closely related to Wallemia ichthyophaga. Keywords: phylogenetic analysis, 18S rRNA sequence.
Ekspresi Prethrombin-2 Manusia Recombinan dalamPichia pastoris dan Optimasi Kondisi Ekspresinya Shabarni Gaffar; Purba Upay; Iman Permana Maksum; Khomaini Hasan; Toto Subroto; Sutarya Enus; Soetijoso Soemitro; Wulan Pertiwi
Indonesian Journal of Pharmaceutical Science and Technology Vol 4, No 1 (2017)
Publisher : Indonesian Journal of Pharmaceutical Science and Technology

Show Abstract | Download Original | Original Source | Check in Google Scholar | Full PDF (566.296 KB) | DOI: 10.15416/ijpst.v4i1.9766

Abstract

Pretrombin merupakan prekursor dari trombin yang memiliki aktivitas proteolitik. Trombin merubah fibrinogen menjadi benang fibrin yang salah satu aplikasinya adalah dapat digunakan sebagai lem untuk menggantikan teknik jahitan pasca bedah. Aplikasi trombin untuk pembuatan lem fibrin menuntut diproduksinya trombin rekombinan. Tujuan dari penelitian ini adalah ekspresi gen pretrombin-2 (PT2) manusia rekombinan menggunakan sistem ekspresi Pichia pastoris. Gen pengode PT2 dirancang sesuaidengan kodon preferensi P. pastoris. Fragmen PT2 diamplifikasi dengan metoda PCR dengan penambahan sisi restriksi EcoR1 pada ujung 5’ dan sisi restriksi SacII pada ujung 3’. Produk PCR yang berukuran 924 pb diligasi dengan vektor ekspresi pPICZaB untuk P. pastoris dan disubkloning dalam inang Escherichia coli. Urutan nukleotida dikonfirmasi dengan metoda dideoxy Sanger. Plasmid rekombinan pPICZaB-PT2 kemudian digunakan untuk mentransformasi P. pastoris SMD1168 defisien protease dengan metoda elektroporasi. Hasil penelitian menunjukkan bahwa gen PT2 berhasil diamplifikasi dan dikloning dalam E. coli. Analisis restriksi dan penentuan urutan DNA menunjukkan bahwa PT2 rekombinan 100% homologi dengan hasil rancangan. Hasil ekspresi PT2 oleh P. pastoris menggunakan metanol sebagai inducer memperlihatkan bahwa PT2 dengan berat molekul 35 kDa berhasil diekspresikan. Optimasi kondisi ekspresi melalui variasi konsentrasi metanol sebagai inducer dan sorbitol sebagai sumber karbon tambahan menunjukkan bahwa metanol 2% dan sorbitol 2% merupakan kondisi optimum ekspresi PT2.
Mutation and Phylogenetic Analysis of Spike Glycoprotein of Indonesian Isolates of Severe-Acute-Respiratory-Syndrome-Coronavirus-2 (SARS-CoV-2) Shabarni Gaffar; Syifa Al Fauziah Rahmani; Ari Hardianto
Majalah Kedokteran Bandung Vol 53, No 1 (2021)
Publisher : Faculty of Medicine, Universitas Padjadjaran

Show Abstract | Download Original | Original Source | Check in Google Scholar | DOI: 10.15395/mkb.v53n1.2230

Abstract

Coronavirus disease-2019 (COVID-19) is an infectious acute respiratory disease caused by SARS-CoV-2. The protein that plays a role in the entry of SARS-CoV-2 into human cells is the surface protein, or the Spike, which is thought to be the effective vaccine target to prevent SARS-CoV-2 infection. Until December 2020, Indonesia has reported 106 SARS-CoV-2 genome sequences identified from COVID-19 positive patients. The purpose of this study was to analyze the phylogenetic relationship of the Spike protein of the Indonesian isolates of SARS-CoV-2 Indonesian, as well as the virus mutations and their effects on changes in the amino acid. The 106 Indonesian SARS-CoV-2 genomes were downloaded from GISAID and  the Spike nucleotide and amino acid sequences were analyzed by multiple sequence alignment (MSA) and mutation analysis using the ClustalW method. Phylogenetic trees were created using the Neighbor-Joining method in MEGA-X software. The results showed that 30 of the 106 Indonesian isolate SARS-CoV-2 Spike were 100% identical to the Wuhan-Hu-1, while the remaining 76 had experienced mutations at 1-4 sites. There were 43-point mutations in the Spike gene, 27 of which led to amino acid changes and four had not been reported in other countries. The global mutation D614G was found in 60 Indonesian isolates , of which West Java was the province with the most reports. The phylogenetic of Spike showed that the Indonesian samples have been divided into several branches that are far from Wuhan-Hu-1. This study indicates the possibility of differences in the protein structure of Indonesian isolate SARS-CoV-2 Spike that need to be further studied to manufacture a vaccine against the Indonesian strain of SARS-CoV-2.
Review: Sintesis Dan Karakterisasi Nanopartikel Emas (AuNP) Serta Konjugasi AuNP Dengan DNA Dalam Aplikasi Biosensor Elektrokimia Egista Istioka Fazrin; Annisa Ilma Naviardianti; Santhy Wyantuti; Shabarni Gaffar; Yeni Wahyuni Hartati
PendIPA Journal of Science Education Vol 4, No 2 (2020): MARCH - JUNE
Publisher : University of Bengkulu

Show Abstract | Download Original | Original Source | Check in Google Scholar | Full PDF (726.164 KB) | DOI: 10.33369/pendipa.4.2.21-39

Abstract

The use of gold nanoparticles (AuNP) is an appropriate one in the development of biosensors because it has a unique, strong adsorption, high biocompatibility, and large surface area. AuNPs can be synthesized by various methods with the same synthesis principle by reducing Au3+ to Au. This review explains the various methods of AuNP synthesis and their characteristics, the interaction of AuNP with biomolecules including DNA, and the application of AuNP-DNA bioconjugates in biosensors. Several applications of AuNP-DNA bioconjugates as electrochemical biosensors in the past two years are proposed in this review.
An Electrochemical Immunosensor for the Detection of B-Type Natriuretic Peptide (BNP) Based on Sandwich ELISA Using Screen Printed Carbon Electrodes Yeni Wahyuni Hartati; Muhammad Hilman Daniswara; Ratna Nurmalasari; Shabarni Gaffar; Toto Subroto
Molekul Vol 13, No 1 (2018)
Publisher : Universitas Jenderal Soedirman

Show Abstract | Download Original | Original Source | Check in Google Scholar | Full PDF (448.537 KB) | DOI: 10.20884/1.jm.2018.13.1.379

Abstract

B-type natriuretic peptide (BNP) is a 32-amino acid polypeptide, a cardiac neuro hormone that specifically secreted from heart ventricle as a response towards the increase of volume and pressure in the heart. The determination of BNP concentration in patients blood is one of the method used to diagnose heart failure. An electrochemical immunosensor modified with the streptavidin/biotin system on screen printed carbon electrodes for the detection of the BNP antigen was developed in this study. Monoclonal anti-BNP capture antibody was immobilized on streptavidin-modified SPCEs to give a well oriented of antibody. Furthermore, a biotinylated anti-BNP that conjugated with horseradish peroxidase (HRP) was used as secondary antibody. The electrochemical signal produced by redox activity of substrate 3,3,5,5'-tetramethybezidine dihydrochloride (TMB/H2O2) was measured by differential pulse voltammetry. The BNP immunosensor showed a linear response between 1.0×10-2 and 1.0×102 ng/mL, and the limit of detection was 3.3 ng/mL. BNP immunosensor is a promising technology for the rapid and convenient detection of heart failure.
Expression of Recombinant Antibody Fragment, Anti BNP-SCFV on the Periplasm of Escherichia Coli for the Detection of Heart Failure Shabarni Gaffar; Sofyan Multazam N Aji; Yeni W Hartati; Safri Ishmayana; Toto Subroto
Molekul Vol 12, No 1 (2017)
Publisher : Universitas Jenderal Soedirman

Show Abstract | Download Original | Original Source | Check in Google Scholar | Full PDF (448.733 KB) | DOI: 10.20884/1.jm.2017.12.1.288

Abstract

Basic natriuretic peptide (BNP) is a polypeptide hormone consist of 32 amino acids that secreted by the heart ventricle to respond the excessive stretching of heart muscle cells. BNP can be used as prognostic marker for patients with heart failure. The presence of BNP in blood can be detected by BNP antibody, which is anti BNP-single chain variable fragment (Anti BNP-SCFV). The antibody is a combination of polypeptides between varying region on the heavy chain (VH) and the light chain (VL) of immunoglobulin. Anti BNP-SCFV will bind to BNP through the antigen-antibody interaction. Concentration of BNP in a patient’s blood can be detected through the interaction of BNP with Anti BNP-SCFV using immunosensor method. Production of recombinant Anti BNP-SCFV in Escherichia coli as host is reported in the present study. Anti BNP-SCFV was expressed in fusion form with OmpC signal peptide that direct the protein to a periplasmic space. Expression was performed under RhaBad promoter as control using L-rhamnose as inducer. SDS-PAGE characterization showed consistent band at 28 kDa, which was assumed as Anti BNP-SCFV. The optimum expression was found at four hours after induction with 4 mM inducer. Anti BNP-SCFV was secreted from the cell as characterized by the presence of the protein on periplasmic membrane and extracellular fraction.
PENGARUH KONSENTRASI PENGINDUKSI METANOL SERTA"SUMBER KARBON SORBITOL DAN MONITOL TERHADAP PRODUKSI a-AMILASE Saccharomycopsis fibuligera R64 DALAM Pichia pastoris Shabarni Gaffar; Dani Permana; Diana P Rahmawati; Triana N Meirina; Abu Bakar M.I. Syihab; Safrl Ismayana; Toto Subroto; O Suprijana
Jurnal Kimia Terapan Indonesia Vol 13, No 2 (2011)
Publisher : Research Center for Chemistry - LIPI

Show Abstract | Download Original | Original Source | Check in Google Scholar | Full PDF (5297.927 KB) | DOI: 10.14203/jkti.v13i2.152

Abstract

Pichia pastoris has been widely used as host for heterologous protein expression for commercial purposes. The advantages of P. pastoris as protein expression host is its ability to grow with high cell density and the presence of a gene promoter that canbe induced tightly, named AOXI encoding alcohol oxidase. Improvement of recombinant protein production under control of AOXI promoter in P. pastoris expression system is still a major concern. Optimization of methanol concentration as inducerand addition of carbon sources, is one of the strategies to improve the expression level. This research aims to study the effect of methanol as inducer as well as sorbitol and mannitol as additional carbon source to the expression level of recombinantSaccharomycopsis fibuligera a-amylase (Sfamy) by P. pastoris (Mut). Sorbitol and manuot known as non-repressive carbon source, to increase the growth ofP. past oris, but not inhibit the AOX1 promoter and foreign proteins expression. Sfamy was expressed in P. pastoris GS115 (His-, Mut) with addition ofcarbon source, sorbitol and mannitol saparately to the expression medium. The result showed that, the optimum concentration" of methanol inducer for Sfamy production is 0.75%. The addition of sorbitol or mannitol increased Sfamy production. Concentrations of sorbitol and mannitol 2% with0.75% methanol inducer increase the secretion level of recombinant Sfamy 2.13 times and 1.94 times respectively, compared with no additional carbon source. This result indicated that addition of carbon source can improved recombinant protein production by P. pastoris, and the used of sorbitol as additionalcarbon source is more effective compared to mannitol.Key words: a-amylase. Pichid pastoris, methanol, sorbitol,mannitol.
Temperature effect on expression of recombinant human prethrombin-2 in Escherichia coli BL21(DE3) ArcticExpress Saronom Silaban; Murniaty Simorangkir; Shabarni Gaffar; Iman Permana Maksum; Toto Subroto
Jurnal Pendidikan Kimia (JPKim) Vol 11, No 3 (2019): December
Publisher : Pascasarjana Universitas Negeri Medan

Show Abstract | Download Original | Original Source | Check in Google Scholar | Full PDF (263.103 KB) | DOI: 10.24114/jpkim.v11i3.15779

Abstract

Many proteins produced in E. coli accumulate in inclusion bodies. This study aims to detect the role of temperature in reducing the formation of inclusion bodies during recombinant human prethrombin-2expressed in E. coli BL21 (DE3) Arctic Express host. In this study, we created a host growth curve to find out the right time to add an inducer. The inducer used in our experiment was IPTG 0.1 mM. The fermentation process use a temperature of 12°C and 22°C. The results showed that recombinant human prethrombin-2 was successfully expressed as protein soluble using an optimum temperature of 12°C in E. coli BL21 (DE3) Arctic Express. It was indicated from the 63kDa protein band obtained from the soluble fraction on SDS-PAGE. The higher temperature of fermentation increased the amount of protein in the insoluble fraction due. It concluded that the fermentation temperature affect the rate of prethrombin-2 expression.Keywords: E. coli BL21(DE3) ArcticExpress, prethrombin-2, soluble, temperature
A Rapid and Sensitive Diagnosis of Typhoid Fever Based On Nested PCR-Voltammetric DNA Biosensor Using Flagellin Gene Fragment Yeni Wahyuni Hartati; Santhy Wyantuti; M. Lutfi Firdaus; Nurul Auliany; Rini Surbakti; Shabarni Gaffar
Indonesian Journal of Chemistry Vol 16, No 1 (2016)
Publisher : Universitas Gadjah Mada

Show Abstract | Download Original | Original Source | Check in Google Scholar | Full PDF (333.05 KB) | DOI: 10.22146/ijc.21182

Abstract

Typhoid fever caused by Salmonella typhi is an important issue for public health in the world. Laboratory methods for rapid and sensitive diagnosis are very important for disease management. The purpose of this study was to determine the performance of nested PCR–voltammetric DNA biosensor using flagellin gene (fla) of S. typhi as a marker. The differential pulse voltammetry using pencil graphite electrode was applied to measure the guanine oxidation signal of probes vs synthetic target stDNA and probes vs fla PCR product hybridizations. The probe DNA selectivity was examined by hybridized probes vs non-complementary sequence. The result showed that the first round nested PCR product can not be visualized by agarose electrophoresis, whereas using the voltammetric biosensor methods can be detected both for the first or second round nested PCR product. The average peak current of hybridized probe vs first and second round of PCR product was 2.32 and 1.47 μA respectively, at 0.9 V. Detection of the DNA sequences of the infectious diseases from PCR amplified real sample was also carried out using this voltammetric DNA biosensor methods.
Co-Authors Aathirah, A Sayyidatina Abu Bakar M.I. Syihab Ade Rizki Ridwan Firdaus Aga Adi Masyhuri Agus Safari Al Arofatus Naini Albayyinah, Dyandra Hera Amalia, Riezki Anni Anggraeni, Anni Annisa Ilma Naviardianti Annisa Ilma Naviardianti Ari Hardianto Arum Kurnia Sari Bacthi Alisjahbana Bayu Shiddiq Widhi Pratama Dani Permana Darwati Darwati Desi Harneti Putri Huspa DESSY NATALIA Diana P Rahmawati Egista Istioka Fazrin Ekawardani, Savira Endang Srieatimah - Erina Hilmayanti Ersanda Hafiz Euis Julaeha Euis Julaeha Fajriyah, Maulida Ghina Nur Fadhilah Ghina Uli Felicia Tambunan Hafiz, Ersanda Hesti Lina Wiraswati Iman Permana Maksum Iqbal Wahyu Mustaqim Irkham Irkham KHOMAINI HASAN Kindi Farabi Korry Novitriani, Korry Lia Faridah LINAWATI HARDJITO Lubis, Rubianto A. M. Lutfi Firdaus Ma'ruf, Ilma Fauziah Masyhuri, Aga Adi Mohamad Nurul Azmi Muhammad Hilman Daniswara Muhammad Yudha Nugraha Muhammad Yusuf Murniaty Simorangkir Nayla Haraswati Nurlelasari Nurlelasari Nurmalasari, Ratna Nurul Auliany O Suprijana Oo Suprijana Permadi, Nandang Pertiwi, Wulan Purba Upay Purnama Purnama PUTRI, RAFIKA Putri, Rafika Nanda R. Ukun M.S. Soedjanaatmadja Rani Maharani Rima Melati Rini Surbakti Riza Apriani Riza Apriani Riza Apriani Rubianto A. Lubis Rudi Hartono Rustaman Rustaman Saadah D. Rachman Safri Ishmayana Safrl Ismayana Santhy Wyantuti Sari Syahruni SARONOM SILABAN Saronom Silaban Savira Ekawardhani Shabrinna, Hanif Soetijoso Soemitro SOETIJOSO SOEMITRO Soetijoso Soemitro Sofa Fajriah Sofyan Multazam N Aji Supriyadi, Isma Yustifania Sutarya Enus SUTARYA ENUS Syifa Al Fauziah Rahmani Tambunan, Ghina Uli Felicia Tati Herlina Tati Herlina Toto Subroto Tri Mayanti Triana N Meirina Umi Baroroh Unang Supratman Upay, Purba Vicki Nishinarizki Wulan Pertiwi Yeni W Hartati Yeni Wahyuni Hartati Yenni Wahyuni Hartati, Yenni Wahyuni Yohan Yohan Yohan Yohan Yoshihito Shiono