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All Journal Microbiology Indonesia Pharmacon Jurnal Bioteknologi & Biosains Indonesia (JBBI) BIOEDUSCIENCE Berkala Penelitian Hayati Makara Journal of Science Jurnal Bioteknologi & Biosains Indonesia (JBBI) Jurnal Bioteknologi & Biosains Indonesia
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Agriculture, Biological Sciences & Forestry Biochemistry, Genetics & Molecular Biology Education Environmental Science Health Professions Immunology & microbiology Materials Science & Nanotechnology Medicine & Pharmacology

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Pengaruh Ekstrak Etanol Daun Murbei (Morus Alba L.) dengan Glibenklamid Terhadap Ekspresi Gen CYP3A4 pada Kultur Sel HepG2 Nuralih, Nuralih; Churiyah, Churiyah; Pambudi, Sabar; Tamat, Swasono R.; Meila, Okpri
Pharmacon: Jurnal Farmasi Indonesia Vol 15, No 1 (2018)
Publisher : Universitas Muhammadiyah Surakarta

Show Abstract | Download Original | Original Source | Check in Google Scholar | DOI: 10.23917/pharmacon.v15i1.5766

Abstract

Mulberry leaf is a traditional herb and predicted has ecdysteronecompound which act as antihyperglicemid. Glybenclamide is a synthetic medicine used to cure diabetes mellitus type 2. The leaf reported as competitive inhibitor of CYP3A4 enzyme, which metabolizingglibenclamide. However, in many cases, combination of herbs with synthesis drugs causes interaction if used at the same time. This research aimed to see interaction of ethanol mulberry leaf extract with glibenclamide through CYP3A4 gene expression in HepG2 cell culture.Sample of mulberry extract, glibenclamide, and combination both sample were tested into cell HepG2 culture. Then RNA were isolated and purification using real time PCR to see the gene CYP3A4 expression. As a result, mulberry extract acts as inhibitor enzyme CYP3A4, while glibenclamide is enzyme substrate.The combination of mulberry and glibenclamide showed increased of expression of CYP3A4 gene, means greater enzyme produced, and lower medicine on blood plasma.
EKPRESI ANTIGEN Ag85B DARI Mycobacterium tuberculosis PADA GALUR SEL MAMALIA Pambudi, Sabar; Widayanti, Tika; Stephanie, Nadya
Jurnal Bioteknologi & Biosains Indonesia (JBBI) Vol. 7 No. 1 (2020): June 2020
Publisher : Balai Bioteknologi, Badan Pengkajian dan Penerapan Teknologi (BPPT)

Show Abstract | Download Original | Original Source | Check in Google Scholar | Full PDF (354.863 KB) | DOI: 10.29122/jbbi.v7i1.3840

Abstract

Expression of Mycobacterium tuberculosis Ag85B Antigen in Mammalian Cell CultureTuberculosis (TB) continues to be a major health problem worldwide, affecting millions of people each year. The only vaccine approved for the prevention of TB is Bacillus Calmette-Guérin (BCG). However, one of the limitations of BCG is that its preventive effect against pulmonary TB varies from person to person. Therefore, there arises a need for a new TB vaccine to replace BCG. This study aims to obtain the Ag85B recombinant protein which has characteristics similar to the native Ag85B antigen from Mycobacterium tuberculosis. In this study, we cloned and expressed recombinant Ag85B in mammalian cell culture. In the initial step, we cloned synthetic Ag85B into mammalian expression vector pFLAG-CMV4 and expressed the gene in CHO-K1 cells. Interestingly, a specific band around 30 kDa was observed in the culture media of transfected cells by Western blot analysis. The results from our research showed the potency of mammalian expression system to produce recombinant protein Ag85B for new TB vaccine candidate.Keywords: Ag85B, mammalian cells, tuberculosis, vaccine, expression ABSTRAKTuberkulosis (TB) terus menjadi salah satu masalah kesehatan dunia yang mempengaruhi jutaan manusia setiap tahun. Satu-satunya vaksin untuk TB yang ada adalah Bacillus Calmette-Guérin (BCG). Namun demikian, vaksin BCG ini memiliki kelemahan berupa terjadi efek preventif yang bervariasi dari satu individu terhadap individu lainnya.  Oleh sebab itu diperlukan pengembangan vaksin TB yang dapat menggantikan vaksin BCG yang sudah ada. Penelitian ini bertujuan memperoleh protein rekombinan Ag85B yang memiliki karakteristik mirip dengan antigen Ag85B native dari Mycobacterium tuberculosis. Pada penelitian ini, telah dilakukan kegiatan pengklonaan dan ekspresi gen Ag85B pada galur sel mamalia.  Pada tahap awal dilakukan pengklonaan gen sintesis Ag85B ke dalam plasmid pada sel mamalia pFLAG-CMV4 dan diekspresikan gennya pada sel CHO-K1. Hasil analisis Western blot menunjukan tersekresinya gen target berukuran 30 kDa pada media kultur dari sel mamalia yang ditransfeksi. Hasil dari penelitian ini menunjukkan potensi dari sistem ekpresi untuk protein rekombinan Ag85B pada galur sel mamalia sebagai kandidat vaksin TB yang baru.
EKPRESI ANTIGEN Ag85B DARI Mycobacterium tuberculosis PADA GALUR SEL MAMALIA Sabar Pambudi; Tika Widayanti; Nadya Stephanie
Jurnal Bioteknologi & Biosains Indonesia (JBBI) Vol. 7 No. 1 (2020): June 2020
Publisher : Balai Bioteknologi, Badan Pengkajian dan Penerapan Teknologi (BPPT)

Show Abstract | Download Original | Original Source | Check in Google Scholar | Full PDF (354.863 KB) | DOI: 10.29122/jbbi.v7i1.3840

Abstract

Expression of Mycobacterium tuberculosis Ag85B Antigen in Mammalian Cell CultureTuberculosis (TB) continues to be a major health problem worldwide, affecting millions of people each year. The only vaccine approved for the prevention of TB is Bacillus Calmette-Guérin (BCG). However, one of the limitations of BCG is that its preventive effect against pulmonary TB varies from person to person. Therefore, there arises a need for a new TB vaccine to replace BCG. This study aims to obtain the Ag85B recombinant protein which has characteristics similar to the native Ag85B antigen from Mycobacterium tuberculosis. In this study, we cloned and expressed recombinant Ag85B in mammalian cell culture. In the initial step, we cloned synthetic Ag85B into mammalian expression vector pFLAG-CMV4 and expressed the gene in CHO-K1 cells. Interestingly, a specific band around 30 kDa was observed in the culture media of transfected cells by Western blot analysis. The results from our research showed the potency of mammalian expression system to produce recombinant protein Ag85B for new TB vaccine candidate.Keywords: Ag85B, mammalian cells, tuberculosis, vaccine, expression ABSTRAKTuberkulosis (TB) terus menjadi salah satu masalah kesehatan dunia yang mempengaruhi jutaan manusia setiap tahun. Satu-satunya vaksin untuk TB yang ada adalah Bacillus Calmette-Guérin (BCG). Namun demikian, vaksin BCG ini memiliki kelemahan berupa terjadi efek preventif yang bervariasi dari satu individu terhadap individu lainnya.  Oleh sebab itu diperlukan pengembangan vaksin TB yang dapat menggantikan vaksin BCG yang sudah ada. Penelitian ini bertujuan memperoleh protein rekombinan Ag85B yang memiliki karakteristik mirip dengan antigen Ag85B native dari Mycobacterium tuberculosis. Pada penelitian ini, telah dilakukan kegiatan pengklonaan dan ekspresi gen Ag85B pada galur sel mamalia.  Pada tahap awal dilakukan pengklonaan gen sintesis Ag85B ke dalam plasmid pada sel mamalia pFLAG-CMV4 dan diekspresikan gennya pada sel CHO-K1. Hasil analisis Western blot menunjukan tersekresinya gen target berukuran 30 kDa pada media kultur dari sel mamalia yang ditransfeksi. Hasil dari penelitian ini menunjukkan potensi dari sistem ekpresi untuk protein rekombinan Ag85B pada galur sel mamalia sebagai kandidat vaksin TB yang baru.
Pengaruh Ekstrak Etanol Daun Murbei (Morus Alba L.) dengan Glibenklamid Terhadap Ekspresi Gen CYP3A4 pada Kultur Sel HepG2 Nuralih Nuralih; Churiyah Churiyah; Sabar Pambudi; Swasono R. Tamat; Okpri Meila
Pharmacon: Jurnal Farmasi Indonesia Vol 15, No 1 (2018)
Publisher : Universitas Muhammadiyah Surakarta

Show Abstract | Download Original | Original Source | Check in Google Scholar | DOI: 10.23917/pharmacon.v15i1.5766

Abstract

Mulberry leaf is a traditional herb and predicted has ecdysteronecompound which act as antihyperglicemid. Glybenclamide is a synthetic medicine used to cure diabetes mellitus type 2. The leaf reported as competitive inhibitor of CYP3A4 enzyme, which metabolizingglibenclamide. However, in many cases, combination of herbs with synthesis drugs causes interaction if used at the same time. This research aimed to see interaction of ethanol mulberry leaf extract with glibenclamide through CYP3A4 gene expression in HepG2 cell culture.Sample of mulberry extract, glibenclamide, and combination both sample were tested into cell HepG2 culture. Then RNA were isolated and purification using real time PCR to see the gene CYP3A4 expression. As a result, mulberry extract acts as inhibitor enzyme CYP3A4, while glibenclamide is enzyme substrate.The combination of mulberry and glibenclamide showed increased of expression of CYP3A4 gene, means greater enzyme produced, and lower medicine on blood plasma.
VERIFIKASI cDNA T29 SEBAGAI KANDIDAT GEN PENGKODE PROTEIN Toxoplasma gondii DENGAN METODE SDS PAGE Ira Djajanegara; Wayan Artama; Retno Lestari; Sabar Pambudi
JURNAL PENELITIAN BIOLOGI BERKALA PENELITIAN HAYATI Vol 11 No 1 (2005): December 2005
Publisher : The East Java Biological Society

Show Abstract | Download Original | Original Source | Check in Google Scholar | DOI: 10.23869/460

Abstract

The process of cDNA construction from mRNA isolated from Toxoplasma gondii has been done. There were 7 candidates cDNA which one of them is called T29. Since Toxoplasma gondii is the cause of toxoplasmosis infection, cloning the gene encoding protein from this parasite provides an important tool for developing diagnostic kit for detection of toxoplasmosis. Digestion of the cDNA T29 with EcoRI which is the restriction site where the cDNA was inserted yielded a 1.862 bp fragment. The fragment was subcloned into E. coli expression vector pMal-p2x and transformed into E.coli strain TB1. Colonies of TB1 were grown on ampicillin plates and the recombinant plasmid was extracted using the standard procedure. The plasmid was digested using EcoRI and PstI, checked by PCR amplification using malE and M13/pUC primers. The recombinant plasmid was expressed in TB1 and the protein extracted was ran in SDS PAGE to observe the presence of the expressed protein. Based on the data from this experiment, there was no expression result of the expressed cDNA which was confirm by the PCR result. Therefore, it was concluded that cDNA T29 was not carrying the gene coding for protein from parasite Toxoplasma gondii.
MUTATION DETECTION OF MULTIDRUG-RESISTANT TUBERCULOSIS BY RT-PCR METHOD AS THE DIAGNOSTIC TOOL OF MDR-TB Titta Novianti; alfero Putra Iryanto; feby feby; callista marsya; putri mega utami; febriana dwi wahyuni; henny saraswati; seprianto; adri nora; roaslein putri; nie nie; sabar pambudi
Jurnal Bioteknologi & Biosains Indonesia (JBBI) Vol. 10 No. 1 (2023): Juni 2023
Publisher : Balai Bioteknologi, Badan Pengkajian dan Penerapan Teknologi (BPPT)

Show Abstract | Download Original | Original Source | Check in Google Scholar | DOI: 10.29122/jbbi.v10i1.5653

Abstract

Eight percent of tuberculosis (TB) cases worldwide are resistant to rifampicin, with mutations occurring in the rpoB and katG genes. It is necessary to develop a specific multidrug-resistant (MDR) diagnostic technique using the RT-PCR method in Indonesia to aid in rapid and accurate diagnosis. In-silico testing using SnapGene software resulted in the design of DNA primers for the katG and rpoB genes, plasmids, and specific probes. This study employed a cross-sectional design using 30 non-MDR-TB and MDR-TB samples from RSUD Sitanala, Tangerang Banten, which were tested for amplification of the katG and rpoB genes using Sybr green RT-PCR. Validity testing was conducted using specific probes for the katG and rpoB genes. The amplification results showed that MDR-TB samples and MDR-TB plasmids required a longer time compared to non-MDR-TB samples and non-MDR-TB plasmids. The Quantification Cycle (Cq) value in non-MDR-TB samples was lower than the Cq value in MDR-TB samples. A t-test revealed a significant difference in Cq values of the rpoB and katG genes between MDR-TB and non-MDR-TB patients (p-value < 0.005). These differences in Cq values indicate that the findings of this study can serve as an initial reference for the development of an RT-PCR-based diagnostic kit for MDR-TB.
Cloning and expression of NS2B/NS3 protein of DENV3 Indonesia strain in Saccharomyces cerevisiae expression system for the development of Dengue vaccine ASRI SULFIANTI; SABAR PAMBUDI; ISMA NUR AZIZAH; DODDY IRAWAN SETYO UTOMO; ABINAWANTO ABINAWANTO
Microbiology Indonesia Vol. 12 No. 2 (2018): June 2018
Publisher : Indonesian Society for microbiology

Show Abstract | Download Original | Original Source | Check in Google Scholar | Full PDF (3936.725 KB) | DOI: 10.5454/mi.12.2.4

Abstract

NS3 protein is 618 amino acids (aa) in length containing serine protease and helicase domains required for DENV replication. Alignment of consensus amino acid sequences from all four DENV serotypes demonstrated that this protein is more conserved (78%) among the different dengue serotypes, which elicits a strong cellular immune response after viral infection in humans and animal models. Present study, a central hydrophilic region of NS3 cofactor, NS2B (NS2BH) with full length of NS3 genes DENV3 Indonesian strain were amplified from CDNA following PCR, and inserted to PYES2/CT shuttle vector. The recombinant plasmid was transformed and expressed in Saccharomyces cerevisiae expression system. As result, detection with Anti-His detector and Anti-NS3 shown NS2BH/NS3 was expressed as 83 KDA protein band. We found that addition of NS2BH on NS3 full length construction plasmid increase the yield and activity of protein expression in S. cerevisiae. In future study, our recombinant NS2B/NS3 protein can be used as recombinant protein in Dengue vaccine development.
The Potency of Aluminum Hydroxide Nanoparticles for Dengue Subunit Vaccine Adjuvant SABAR PAMBUDI; ETIK MARDLIYATI; SILMI RAHMANI; DAMAI RIA SETYAWATI; TIKA WIDAYANTI; ANGELINA GILL; ASRI SULFIANTI; WHINIE LESTARI
Microbiology Indonesia Vol. 12 No. 3 (2018): September 2018
Publisher : Indonesian Society for microbiology

Show Abstract | Download Original | Original Source | Check in Google Scholar | Full PDF (984.8 KB) | DOI: 10.5454/mi.12.3.5

Abstract

The potency of aluminum hydroxide as an adjuvant in vaccine development is considered to depend on its particle size. In previous studies, we have successfully prepared two size particle, micro, and nano, aluminum hydroxide gel (alum) adjuvants. The potency of those particles as a candidate of adjuvant is needed to be characterized. In this study, we formulated our adjuvants with purified DENV3 pre Membrane Envelope (prM-E) recombinant protein and evaluated the induction of nitric oxide level in mouse macrophage RAW 264.7 cells. We prepared the alum adjuvant by precipitation-homogenization methods with an agitation rate at 11,000xg. Secreted prM-E recombinant protein was collected from Pichia pastoris X-33 fermentation which produced using bioreactor. Recombinant protein purification was carried out by anion exchange chromatography followed with size exclusion chromatography. The purified prM-E recombinant protein was observed as a single band around 70 -1k Da with a concentration of 105 mg mL . Complex nanoparticles alum with prM-E protein significantly (p<0.05) induced the nitric oxide level. Further analysis should be conducted in order to discover the detail molecular mechanism of nanoparticle alum adjuvant, recombinant protein, and cellular immune response.
CLONING OF DENGUE VIRUS TYPE 3 (INDONESIAN STRAIN D3-1703) NON STRUCTURAL-1 GENE INTO pYES2/CT VECTOR Narita, Vanny; Widyanto, Rahma Micho; Pambudi, Sabar; Sudiro, Tjahjani Tjahjani
Makara Journal of Science Vol. 15, No. 2
Publisher : UI Scholars Hub

Show Abstract | Download Original | Original Source | Check in Google Scholar

Abstract

Dengue is an infectious disease caused by dengue virus. Dengue endemic region includes America, Western Pacific, Africa, East Mediterranian, and South East Asia including Indonesia. An early diagnostic system specific for Indonesia is needed to control dengue in Indonesia. In this research, cloning of Non Structural 1 (NS1) gene from dengue virus type 3 (Indonesian strain D3-1703) into pYES2/CT vector was performed. In the long run, NS1 recombinant protein will be expressed in Saccharomyces cerevisiae for diagnostic materials. Polymerase Chain Reaction (PCR) amplification of NS1 gene fragments were done with optimal annealing temperature at 55 ºC. NS1 gene fragment and pYES2/CT were cut by BamH I and Not I enzymes. The digested pYES2/CT was dephosphosrylated using Calf Intestine Alkaline Phospatase enzyme. Ligation with the vector:insert ratio of 1:12 and 1:20 resulted in 6 and 5 recombinant colony candidates respectively. Restriction enzyme and PCR verifications showed that 5 recombinant plasmids contained NS1 gene. Sequencing of the first 600 bp of one recombinant plasmid was performed. The blastn analysis showed that it had a 99% identity with dengue virus type 3 strain FW06. Finally, it was shown that NS1 clone within pYES2/CT was in the correct Open Reading Frame and ready to be expressed in S. cerevisiae.
MUTATION DETECTION OF MULTIDRUG-RESISTANT TUBERCULOSIS BY RT-PCR METHOD AS THE DIAGNOSTIC TOOL OF MDR-TB Sabar Pambudi
Jurnal Bioteknologi dan Biosains Indonesia Vol. 10 No. 1 (2023)
Publisher : BRIN - Badan Riset dan Inovasi Nasional

Show Abstract | Download Original | Original Source | Check in Google Scholar

Abstract

Eight percent of tuberculosis (TB) cases worldwide are resistant to rifampicin, with mutations occurring in the rpoB and katG genes. It is necessary to develop a specific multidrug-resistant (MDR) diagnostic technique using the RT-PCR method in Indonesia to aid in rapid and accurate diagnosis. In-silico testing using SnapGene software resulted in the design of DNA primers for the katG and rpoB genes, plasmids, and specific probes. This study employed a cross-sectional design using 30 non-MDR-TB and MDR-TB samples from RSUD Sitanala, Tangerang Banten, which were tested for amplification of the katG and rpoB genes using Sybr green RT-PCR. Validity testing was conducted using specific probes for the katG and rpoB genes. The amplification results showed that MDR-TB samples and MDR-TB plasmids required a longer time compared to non-MDR-TB samples and non-MDR-TB plasmids. The Quantification Cycle (Cq) value in non-MDR-TB samples was lower than the Cq value in MDR-TB samples. A t-test revealed a significant difference in Cq values of the rpoB and katG genes between MDR-TB and non-MDR-TB patients (p-value < 0.005). These differences in Cq values indicate that the findings of this study can serve as an initial reference for the development of an RT-PCR-based diagnostic kit for MDR-TB.
Co-Authors A.A. Ketut Agung Cahyawan W Abinawanto Alfan Danny Arbianto alfero Putra Iryanto ANGELINA GILL ANOM BOWOLAKSONO Arbianto, Alfan Danny ASRI SULFIANTI Astari Dwiranti Azzizah, Isma Nur Bachti Alisjahbana callista marsya CHURIYAH CHURIYAH Churiyah Churiyah DAMAI RIA SETYAWATI Dewi Esti Restiani DODDY IRAWAN SETYO UTOMO Etik Mardliyati Febriana Dwi Wahyuni feby feby HENI RACHMAWATI Ira Djajanegara Ira Djajanegara Ira Djajanegara Nadya Stephanie nie nie Nina Artanti Nina Artanti Nora, Adri Novianti, Titta Nuralih Nuralih Nuralih Nuralih, Nuralih Okpri Meila Okpri Meila, Okpri putri mega utami Rahma Micho Widyanto Rahmana E. Kartasasmita Rahmana E. Kartasasmita Rajagukguk, Selly Setiati Restiani, Dewi Esti Retno Lestari Retno Lestari roaslein putri Rukmi, Ai Saraswati, Henny Seprianto, Seprianto SILMI RAHMANI Sri Darmawati Stephanie, Nadya Sudiro, Tjahjani Tjahjani Sukmadjaja Asyarie Sukmadjaja Asyarie Sulaiman Ngongu Depamede Swasono R. Tamat Swasono R. Tamat Tarwadi Tarwadi Tarwadi, Tarwadi Tika Widayanti TIKA WIDAYANTI Tika Widayanti Utomo, Doddy Irawan Setyo Vanny Narita Wayan Artama Whinie Lestari Yuwita, Anita