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Indonesian Journal of Biotechnology

ijbiotech
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Published by Universitas Gadjah Mada

ISSN : 08538654 EISSN : 20892241 DOI : -

Core Subject : Science,

Biochemistry, Genetics & Molecular Biology Immunology & microbiology Materials Science & Nanotechnology

The Indonesian Journal of Biotechnology (IJBiotech) is an open access, peer-reviewed, multidisciplinary journal dedicated to the publication of novel research in all aspects of biotechnology, with particular attention paid to the exploration and development of natural products derived from tropical—and especially Indonesian—biodiversity. IJBiotech is published biannually and accepts original research articles featuring well-designed studies with clearly analyzed and logically interpreted results. A strong preference is given to research that has the potential to make significant contributions to both the field of biotechnology and society in general.

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Articles 8 Documents

Search results for , issue "Vol 24, No 1 (2019)" : 8 Documents clear

Isolation, identification, and detection of ACC deaminase gene-encoding rhizobacteria from rhizosphere of stressed pineapple Dori Kusuma Jaya; Giyanto Giyanto; Novik Nurhidayat; Sarjiya Antonius
Indonesian Journal of Biotechnology Vol 24, No 1 (2019)
Publisher : Universitas Gadjah Mada

ACC deaminase is a microbial cytoplasmic enzyme that cleaves ACC, a precursor of ethylene, in the stressed plant. The aims of this study were to isolate, identify, and detect the presence of ACC deaminase gene-encoding rhizobacteria from the rhizospheric soil of pineapple plants that have been exposed to abiotic and biotic stress, specifically herbicide, flooding, and Phytophthora spp. stress. A total of 49 rhizobacterial isolates were obtained, seven of which were observed for their growth on DF medium containing 3 mM L-1 ACC. The four best-growing isolates were selected for genomic DNA extraction. They were molecularly identified as Stenotrophomonas maltophilia (3), Burkholderia territorii (2A), Pseudomonas oryzihabitans (5B), and Bacillus tropicus (1E). A set of primers, 105F-acdS 5’-TGCCAAGCGTGAAGACTGC-3’ and 244R-acdS 5’-GGGTCTGGTTCGACTGGAT-3’, were constructed to amplify the ACC deaminase gene (acdS). Based on melt peak curve analysis, four products appeared to show a specific single peak at 86, 89, 87, and 89.5°C, indicating a single product was produced. In addition, a Blast search showed that these four products met the ACC deaminase feature and their acdS sequences were clustered into an ancestral group compared with the bacterial strains deposited in GenBank. These results suggest that ACC deaminase gene-encoding rhizobacteria from a pineapple plantation of tropical origin may affect the acdS sequences and may contribute to the host plant’s stress tolerance.

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

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.

Identification of single nucleotide polymorphisms in GDF9 gene associated with litter size in Garut sheep Resti Yuliana Rahmawati; Sumadi Sumadi; Tety Hartatik
Indonesian Journal of Biotechnology Vol 24, No 1 (2019)
Publisher : Universitas Gadjah Mada

The growth differentiation factor 9 (GDF9) gene has been regarded as having major impacts on ovulation rate and litter size in sheep. The aim of this study was to identify the single nucleotide polymorphisms (SNPs) of the GDF9 gene and their association with litter size in Garut sheep. For this purpose, a total of 60 ewes of Garut sheep were included in this study. Based on the sheep GDF9 reference sequences (Genbank Acc. No. AF078545.2), one pair of primers (5’-CTGCTGTTTAACCTGGATCGTG-3 5’-GGAGAGCCATACCGATGTCC-3 as forward and reverse, respectively) was used for PCR amplification. The results revealed that four SNPs (g.54C>T, g.60G>A, g.304G>A, and g.333G>A) were found in Garut sheep by direct sequencing. For SNP g.54C>T, the sheep exhibited the highest frequency of allele C and genotype CC. On the other hand, SNPs g.60G>A, g.304G>A, and g.333G>A showed a higher frequency of allele G than allele A, and the GG genotype was predominant in the population. SNP g.333G>A had a significant effect on litter size (p < 0.05), and ewes with the GG genotype had a higher litter size than those with the GA genotype. Genotype distributions for all identified SNPs were in agreement with Hardy-Weinberg equilibrium. We highlight that SNP g.333G>A may be useful as a genetic marker for litter size in Garut sheep.

The characterization of bacteriocins produced by Lactobacillus plantarum strains isolated from traditional fermented foods in Indonesia and the detection of its plantaricin-encoding genes Sogandi Sogandi; Apon Zaenal Mustopa; I Made Artika
Indonesian Journal of Biotechnology Vol 24, No 1 (2019)
Publisher : Universitas Gadjah Mada

Lactobacillus plantarum is widely found in either anaerobic plant matter or fermented foods, and it has been recognized as producing antimicrobial bacteriocins. This study aimed to characterize the antimicrobial bacteriocins of L. plantarum and detect its genes that encode plantaricins. Samples were isolated from traditional fermented foods from Indonesia. Antimicrobial activity was evaluated using the agar diffusion assay procedure. The titration method applied the maximum amounts of lactic acid at 1054 mg/mL and hydrogen peroxide at 3.85 mg/mL. Based on the results, the supernatant of the L. plantarum strains appeared to have a broad spectrum of antimicrobial activity against pathogens, which would be active at pH 2.0–12.0 and stable temperature. In addition, almost all of the L. plantarum strains contained plantaricin-encoding genes (e.g. plnA, plnF,plnJK, and plnW), which were grouped into one cluster as indicated by phylogenetic analysis. Therefore, this study discovered clear evidence of the potential of some L. plantarum strains to act as antimicrobial agents.

Diagnosis and molecular characterization of Anaplasma platys in dog patients in Yogyakarta area, Indonesia Muh. Disna Faizal; Aris Haryanto; Ida Tjahajati
Indonesian Journal of Biotechnology Vol 24, No 1 (2019)
Publisher : Universitas Gadjah Mada

Anaplasma platys is a tick-borne, Gram-negative bacterium that causes anaplasmosis, a companion vector-borne disease impacting dogs. Information on this disease remains limited in Indonesia. Its symptoms are not specific, so molecular analysis is required for a rapid and accurate diagnosis. GroEL is an essential gene commonly used for classification and species identification of many groups of bacteria, including Anaplasma spp. In this study, a molecular diagnosis of anaplasmosis based on the groEL gene sequence was conducted using PCR. In addition, the genetic diversity of Anaplasma platys in infected dogs was determined. Blood samples were collected from 51 dogs suspected of anaplasmosis from Prof. Dr. Soeparwi Animal Hospital, animal clinics, and pet shops in the Yogyakarta area, Indonesia, based on anamnesis, histories of tick infestations, and clinical symptom examinations. DNA extraction and PCR targeting the groEL gene were performed, followed by sequencing. Phylogenetic tree analysis and construction were carried out using the BLAST and MEGA programs. Positive PCR sample results (amplicon length of 624 bp) were found in 6 of 51 dogs. Samples A1 (KHJ/C2), A2 (KHJ/A2), A3 (KSK/L), A4 (KHJ/L), and A5 (KNP/M2) had close ties to Anaplasma platys (AF478129.1) from GenBank. Phylogenetic analysis showed a very high homology value (100%) and bootstrap value of 100%. It can be concluded that there was no genetic diversity in the Anaplasma platys found in infected dogs in the Yogyakarta area.

Inverse correlation of kidney interstitial cells expansion with hemoglobin level and erythropoietin expression in single and repeated kidney ischemic/reperfusion injury in mice Dian Prasetyo Wibisono; Nur Arfian; Muhammad Mansyur Romi; Wiwit Ananda Wahyu Setyaningsih; Dwi Cahyani Ratna Sari
Indonesian Journal of Biotechnology Vol 24, No 1 (2019)
Publisher : Universitas Gadjah Mada

Ischemic/reperfusion injury (IRI) causes acute kidney injury that may lead to chronic kidney disease. We investigated the correlation between kidney interstitial cells expansion, hemoglobin level, and erythropoietin expression as the chronic effects of single and repeated kidney IRI in mice. We created an IRI model using male Swiss mice by clamping the bilateral renal pedicles. Subjects were divided into four groups that contained six mice each: control/sham operation, single acute IRI, single chronic IRI, and repeated IRI. Our results showed that the single chronic and repeated IRI groups significantly increased the tubular injury score, decreased the hemoglobin level, and increased erythropoietin expression compared with the control. Lower hemoglobin levels in all of the groups compared with the control was associated with erythropoietin resistance. In single chronic and repeated kidney IRI, there were decreased creatinine levels compared with the control. The decreased creatinine levels from the single acute IRI group to the single chronic IRI group, suggesting a repair phase of IRI starting on day 7 occurred in the single chronic IRI group. A macrophage marker, CD68, and an inflammatory mediator marker, MCP-1, significantly increased in all IR groups, indicating inflammation occurred due to IRI. In conclusion, chronic and repeated kidney IRI induced interstitial cells expansion and inflammation associated with anemia.

The establishment of PCR amplification, cloning, and sequencing of bovine herpesvirus 1 (BHV-1) glycoprotein D gene isolated in Indonesia Dewi Noor Hidayati; Eko Agus Srihanto; Tri Untari; Michael Haryadi Wibowo; Koichi Akiyama; Widya Asmara
Indonesian Journal of Biotechnology Vol 24, No 1 (2019)
Publisher : Universitas Gadjah Mada

Considering the increasing incidence of infectious bovine rhinotracheitis (IBR) in Indonesia, it was necessary to conduct a more in-depth study of bovine herpesvirus-1 (BHV-1) as the causative agent of IBR disease. Previous research reports indicate that the BHV-1 subtypes found in Indonesia are subtype 1.1. Currently, IBR field case detection in Indonesia still uses the serological method (ELISA), which has the potential to give false positive results and cannot explain the virus subtype. Other detection methods, such as viral isolation, take longer and require adequate resources. This study aimed to determine the BHV-1 subtypes of Indonesian isolates using molecular techniques. Nested PCR using two pairs of primers was successfully used to amplify the glycoprotein D (gD) gene. The gD gene fragment was cloned into the pGEM-T plasmid. Analysis of the gD gene sequence was subsequently carried out to determine the BHV-1 character of the Indonesian isolates. The results indicated that the isolates were different from the previous isolates, and had similarities (100%) with subtype 1.2 strain SP1777 and SM023.

Expression and purification of recombinant coat protein of sugarcane mosaic virus from Indonesian isolate as an antigen for antibody production Natalia Tri Astuti; Nurmalasari Darsono; Suvia Widyaningrum; Widhi Dyah Sawitri; Sri Puji Astuti; Win Darmanto
Indonesian Journal of Biotechnology Vol 24, No 1 (2019)
Publisher : Universitas Gadjah Mada

Sugarcane mosaic virus (SCMV, genus Potyvirus, family Potyviridae) is a prominent pathogen of sugarcane (Saccharum sp. hybrids). It can cause losses in susceptible varieties, in crop as well as sugar production, economically. Although it has been studied in major sugar-producing countries, research on the definement of SCMV from Indonesian isolates based on molecular study has been very limited. This study aimed to obtain a proper recombinant antigens emanating from coat protein of SCMV from Indonesian isolate in order to produce polyclonal antibodies that cann be used for immunodiagnosis assays in a subsequent study. A gene-encoding coat protein of SCMV (CP-SCMV) was amplified using RT-PCR and cloned into vector pJET1.2. The cDNA was inserted into 6X His-tag expression plasmid of pET28a(+) and over-expressed in Escherichia coli BL21(DE3) to produce a recombinant protein. The highest expression was found in 0.1M IPTG induction media for 5 h at 37oC. SDS-PAGE analysis clarified that the recombinant CP-SCMV remained as an insoluble fraction. Purifications was carried out by the affinity Ni-NTA resin, followed by electroelution to obtain a highly purified protein. To meet the quality requirements of a proper antigen, the highly purified protein was concentrated. A molecular weight of the rCP-SCMV (approximately 40 kDa) was clearly observed by 10% SDS-PAGE at the concentration of 16.184 mg/mL.

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