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Identification of Potential Bacteria on Several Lakes in East Java, Indonesia Based on 16S rRNA Sequence Analysis Achmad Rodiansyah; Ainul Fitria Mahmudah; Mastika Marisahani Ulfah; Uun Rohmawati; Dwi Listyorini; Eko Agus Suyono; Sitoresmi Prabaningtyas
HAYATI Journal of Biosciences Vol. 28 No. 2 (2021): April 2021
Publisher : Bogor Agricultural University, Indonesia

Show Abstract | Download Original | Original Source | Check in Google Scholar | DOI: 10.4308/hjb.28.2.136

Abstract

Four bacterial isolates from Ranu Pani and Ranu Grati in east java had been revealed to be potentials to produce IAA (PIS isolate), phosphate solubilizer (GPS isolate), cellulose hydrolysis (PSS isolate) and, amylum hydrolysis (PAS), two dominant bacterial isolates from Rani Pani (PØD isolate) and Ranu Grati (GØD isolate) which were co-cultured with microalgae promoted microalgae growth, yet its taxonomical position has not been clearly known. The aim of this study was to identify those bacterial isolates using 16S rRNA barcode. This research conducted by gDNA isolation, the 16S rRNA sequence was amplified using 27F and 1492R primers. Reconstructed phylogenetic trees and genetic distance analysis showed that the isolate PIS and PSS identified as Bacillus cereus Group closely related to Bacillus paramycoides. PAS isolate identified as Bacillus subtilis Group closely related to Bacillus amyloliquefaciens, GPS isolate identified as novel species in genus Enterobacter, and two dominant isolates (PØD and GØD) identified as Enterobacter cloacae complex closely related to Enterobacter cloacae. The genomic approach and additional phenotypes-examination are required to clarify its taxonomical position.
Modification of recombinant human epidermal growth factor (rh-EGF) expression vector by site-directed mutagenesis for therapeutic protein production Achmad Rodiansyah; Riyona Desvy Pratiwi; Sabighoh Zanjabila; Asrul Muhamad Fuad
Indonesian Journal of Biotechnology Vol 24, No 1 (2019)
Publisher : Universitas Gadjah Mada

Show Abstract | Download Original | Original Source | Check in Google Scholar | DOI: 10.22146/ijbiotech.41859

Abstract

Recombinant human epidermal growth factor (rh-EGF) has high value in therapies for h-EGF deficiency-related diseases. The expression of the h-EGF gene was designed by using the pET21b(+) vector and Escherichia coli BL21(DE3) as the expression host. In a previous study, the sequence of a 6xHis tag without any restriction sites was fused to the h-EGF gene, yet it was not possible to obtain a purified and single rh-EGF by this approach. In this study, we modified the rh-EGF expression vector using site-directed mutagenesis (SDM) to remove the sequence of the 6xHis tag. The vector modification was carried out by inserting a stop codon and the EcoRI restriction site, along with deleting the 6xHis tag sequence. The results of PCR showed non-specific bands, while 2-step cycles PCR produced one non-specific band, and 3-step cycles PCR produced two non-specific bands. After purification of the PCR products, the SDM-recombinant plasmids treated for template plasmid-free product were transformed into E. coli DH5a. Even though the transformation efficiency was low, the planned gene mutations including the deletion of the 6xHis tag and insertion of the stop codon and EcoRI restriction site in plasmid pET21b(+) were successfully carried out. When using this modified vector in expression studies, rh-EGF of a similar size to that of the rh-EGF standard and approximately 1 kDa smaller than the rh-EGF-6xHis of the previous study was obtained.
Isolation and characterization of α ‐amylase encoding gene in Bacillus amyloliquefaciens PAS Achmad Rodiansyah; Sitoresmi Prabaningtyas; Mastika Marisahani Ulfah; Ainul Fitria Mahmuda; Uun Rohmawati
Indonesian Journal of Biotechnology Vol 26, No 4 (2021)
Publisher : Universitas Gadjah Mada

Show Abstract | Download Original | Original Source | Check in Google Scholar | DOI: 10.22146/ijbiotech.61425

Abstract

Amylolytic bacteria are a source of amylase, which is an essential enzyme to support microalgae growth in the bioreactor for microalgae culture. In a previous study, the highest bacterial isolate to hydrolyze amylum (namely PAS) was successfully isolated from Ranu Pani, Indonesia, and it was identified as Bacillus amyloliquefaciens. That bacterial isolate (B. amyloliquefaciens PAS) also has been proven to accelerate Chlorella vulgaris growth in the mini bioreactor. This study aims to detect, isolate, and characterize the PAS’s α‐amylase encoding gene. This study was conducted with DNA extraction, amplification of α‐amylase gene with polymerase chain reaction (PCR) method with the specific primers, DNA sequencing, phylogenetic tree construction, and protein modeling. The result showed that α‐amylase was successfully detected in PAS bacterial isolate. The α‐amylase DNA fragment was obtained 1,468 bp and that translated sequence has an identity of about 98.3% compared to the B. amylolyquefaciens α‐amylase 3BH4 in the Protein Data Bank (PDB). The predicted 3D protein model of the PAS’s α‐amylase encoding gene has amino acid variations that predicted affect the protein’s structure in the small region. This research will be useful for further research to produce recombinant α‐amylase.
Screening and Prediction of Potential Compounds from Virgin Olive Oil Acting on Proteins Associated with Cancer Disease Achmad Rodiansyah
Majalah Obat Tradisional Vol 26, No 1 (2021)
Publisher : Faculty of Pharmacy, Universitas Gadjah Mada

Show Abstract | Download Original | Original Source | Check in Google Scholar | DOI: 10.22146/mot.52737

Abstract

Virgin olive oil contains phenolic compounds that were potential for anti-inflammatory and cancer treatment. Computational biology is a beneficial method to understand how this compound can affect the biological process in humans. This research is conducted by the potential screening of VOO compounds, constructing the pharmacological network and enrichment, and docking simulation. The enrichment result showed that the EGFR, BRAF, MAPK1, CCND1, and MDM2 protein have multiple cancer contributions and related pathways. The docking simulation result showed that the interaction of EGFR-luteolin, BRAF-luteolin, MAPK1-luteolin, CCND1-apigenin, and MDM2-1-hydroxypinoresinol has the highest binding affinity. Further research with the in-vitro method is required to check the reliable mechanisms of each compound to their protein target.