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Background: Background: Ursodeoxycholic acid (UDCA), a secondary bile acid, is an acidic steroid synthesized from cholesterol in hepatocytes. UDCA is widely used for the treatment of various diseases related to liver injury. The use of UDCA to treat non-liver diseases has also been developed recently, such as neurodegenerative diseases, cancer, and obesity. Due to the important role of UDCA on human health, numerous studies in understanding its chemical and pharmacological properties have been published. Methods: Literature sources were obtained from online databases such as Science Direct, Google Scholar, Scopus and PubMed using keywords relating to the purpose of study. Critical analysis and review were performed for all literatures. Results: UDCA is a steroid compound with pharmacological properties. Seventeen enzymes are involved in its biosynthesis, which has been proposed in four pathways: classic, alternative, the Yamazaki, and 25-hydroxylation pathways. UDCA can be isolated from bovine bile, bear bile or all Ursidae, human, rabbit, cow, rat, hamster, sheep, pig, and plant. UDCA has been used in the treatment of several diseases such as primary biliary cirrhosis, intrahepatic cholestasis of pregnancy, hepatolithiasis associated with Caroli syndrome gallstones, cystic fibrosis, hepatitis C virus, chronic heart failure, neurodegenerative diseases, and obesity, as well as in the prevention of cancer. Conclusion: UDCA has a wide range of the pharmacological properties. Further investigations on its efficacy and safety on humans are required before it could be used for several indications. All genes which are responsible for UDCA biosynthesis have been elucidated. That said, further genetic engineering studies in order to find other prospective sources of UDCA could be a challenge for the future research.

Surfactants are amphiphilic molecules, which have hydrophilic and hydrophobic groups. Surfactants have an important role in various fields including agriculture, cosmetics, pharmaceuticals, bioremediation, and the petroleum industry especially EOR but because synthetic surfactants are not biodegradable, it is necessary to produce biodegradable surfactants such as rhamnolipid biosurfactants. Rhamnolipid is a glycolipid biosurfactant produced by Pseudomonas aeuruginosa. This species is a pathogen, so it is needed to overcome this problem by cloning the rhamnolipid gene into Escherichia coli for large-scale production. Rhamnolipid biosynthesis includes three main genes, rhlA, rhlB, and rhlC. The rhlAB produces mono-rhamnolipid and rhlABC produces di-rhamnolipid. The construction involved one plasmid pPM RHLABC (di-rhamnolipid) with T7lac promoter. Characterization of surfactants by E24, IFT, and CMC analysis showed that di-rhamnolipid biosurfactant has the best activity (70%, 0.8 mN/m, and 300 mg/L) than chemical surfactant, sodium dodecyl sulfate (46%, 4.7 mN/m, and 2000 mg/L) at pH 7, 25 °C, and 0% salinity. The conclusion from this research shows that the characteristics of di-rhamnolipid are very promising in the utilization of industrial-scale including EOR technology, agriculture, and pharmacy

Reducing oxidative species to non- or less-reactive matter is the principal function of an antioxidant. Plant-based food is the main external source of antioxidants that helps protect our cells from oxidative damage. During postharvest storage and distribution, fruits and vegetables often increase ROS production that is quenched by depleting their antioxidant pools to protect their cells, which may leave none for humans. ROS are molecules produced from oxygen metabolism; some of the most widely analyzed ROS in plants are singlet oxygen, superoxide, hydrogen peroxide, and hydroxyl radicals. ROS concentration and lifetime are determined by the availability and composition of the antioxidant system that includes enzymatic components such as SOD, CAT, and APX and nonenzymatic components such as vitamins, polyphenols, and carotenoid. Depending on its concentration in the cell, ROS can either be harmful or beneficial. At high concentrations, ROS can damage various kinds of biomolecules such as lipids, proteins, DNA, and RNA, whereas at low or moderate concentrations, ROS can act as second messengers in the intracellular signaling cascade that mediates various plant responses. Novel postharvest methods are sought to maintain fruit and vegetable quality, including minimizing ROS while preserving their antioxidant content.

Background: Immediate and accurate diagnosis of malaria is essential for effective control of this disease. Immunochromatographic based rapid diagnostic tests (RDTs) are economical, simple to perform, and provide results in a relative short time, can be useful to assist effective management of malaria. The commercially available malaria RDT in Indonesia is still imported. Therefore, an effort to produce malaria RDT independently is necessary. One of the biomarkers used in RDTs is Plasmodium lactate dehydrogenase pLDH. The production and accumulation of pLDH during asexual stage or blood-stage in all human infected malaria parasites can be used to indicate parasites viability, which is correlated with the number of parasites present in the plasma of infected patients. Objective: The aim of this research is to produce recombinant PfLDH in Escherichia coli BL21(DE3). Methods: PfLDH gene was cloned into pET30a expression vector to obtain a 6.2 kbp recombinant plasmid pET30a-PfLDH. E. coli BL21(DE3) was transformed with pET30a-PfLDH using the heat shock method. Then, E. coli BL21(DE3)- pET30a-PfLDH was cultured in LB broth containing 50 mg/mL kanamycin and was induced by 1mM IPTG at 37oC. Results: SDS-PAGE and Western Blot analysis showed that recombinant PfLDH was expressed with molecular mass ~30 kDa. Conclusion: Recombinant PfLDH is expressed in E. coli BL21(DE3) and can be used in further research for producing rPfLDH as a biomarker for malaria RDT development.

Introduction. A halophilic bacterium of theHalomonas elongataBK-AG25 has successfully produced ectoine with high productivity. To overcome the drawbacks of high levels of salt in the production process, a nonhalophilic bacteria ofEscherichia coli(E. coli) was used to express the ectoine gene cluster of the halophilic bacteria, and the production of ectoine by the recombinant cell was optimized.Methods. The ectoine gene cluster from the halophilic bacterium was isolated and inserted into an expression plasmid of pET30(a) and subsequently transformed intoE. coliBL21 (DE3). Production of ectoine from the recombinantE. coliwas investigated and then maximized by optimizing the level of nutrients in the medium, as well as the bioprocess conditions using response surface methodology. The experimental designs were performed using a central composite design.Results. The recombinantE. colisuccessfully expressed the ectoine gene cluster ofHalomonas elongataBK-AG25 under the control of theT7promoter. The recombinant cell was able to produce ectoine, of which most were excreted into the medium. The optimization of ectoine production with the response surface methodology showed that the level of salt in the medium, the incubation temperature, the optical density of the bacteria before induction, and the final concentration of the inducer gave a significant effect on ectoine production by the recombinantE. coli. Interestingly, the level of salt in the medium and the incubation temperature showed an inverse effect on the production of intracellular and extracellular ectoine by the recombinant cell. At the optimum conditions, the production yield was about 418 mg ectoine/g cdw (cell dry weight) after 12 hours of incubation.Conclusion. This study is the first report on the expression of an ectoine gene cluster ofHalomonas elongataBK-AG25 inE. coliBL21, under the control of theT7promoter. Optimization of the level of nutrients in the medium, as well as the bioprocess condition using response surface methodology, has successfully increased the production of ectoine by the recombinant bacteria.

Rhamnolipid is a potent biodegradable surfactant, which frequently used in pharmaceutical and environmental industries, such as enhanced oil recovery and bioremediation. This study aims to engineer Escherichia coli for the heterologous host production of rhamnolipid, to characterize the rhamnolipid product, and to optimize the production using autoinduction medium and POME (palm oil mill effluent). The construction of genes involved in rhamnolipid biosynthesis was designed in two plasmids, pPM RHLAB (mono-rhamnolipid production plasmid) and pPM RHLABC (di-rhamnolipid production plasmid). The characterization of rhamnolipid congeners and activity using high-resolution mass spectrometry (HRMS) and critical micelle concentration (CMC). In order to estimate rhamnolipid yield, an oil spreading test was performed. HRMS and CMC result show E. coli pPM RHLAB mainly produced mono-rhamnolipid (Rha-C14:2) with 900 mg/L and 35.4 mN/m of CMC and surface tension value, whereas E. coli pPM RHLABC mainly produced di-rhamnolipid (Rha-Rha-C10) with 300 mg/L and 34.3 mN/m of CMC and surface tension value, respectively. The optimum condition to produce rhamnolipid was at 20 h cultivation time, 37 oC, and pH 7. In this condition, the maximum rhamnolipid yield of 1245.68 mg/L using autoinduction medium and 318.42 mg/L using 20% (v/v) of POME. In conclusion, the characteristics of the rhamnolipid by recombinant E. coli is very promising to be used in industries as the most economical way of producing rhamnolipid.

Global demand of antimalarial drug artemisinin has a gap with production capacity from existing sources since the low content of this compound from Artemia annua L. Genetic engineering-based strategy for A. annua plant on key enzymes in artemisinin biosynthetic pathway is needed. Artemisinic aldehyde ∆ 11 (13) double bond reductase (dbr2) is one of the key enzyme on artemisinin biosynthesis which was studied in this research. Agrobacterium tumefaciens -mediated transformation of A. annua using dbr2 was carried out. Synthetic dbr2 was ligated into pCAMBIA1303 and transformed into Escherichia coli DH5α. pCAMBIA1303- dbr2 plasmid was transformed to A. tumefaciens AGL1. Leaves of A. annua were infected by positive transformant of recombinant A. tumefaciens (OD600 ≈ 1) supplemented with acetosyringone 50 ppm, and Silwet S-408 0.02%. Samples were incubated in desiccators connected with vacuum pump, this method is called infiltration vacuum. Leaves were covered in dark for 45 min, and co-cultivated on MS co-cultivation media for 3 days. All leaves were washed in 300 ppm cefotaxime and divided into 2 parts; 3 leaves for GUS histochemical assay and 300 mg of leaves for HPLC analysis. Transient transformation was done in triplicate. In GUS histochemical assay, pCAMBIA1303 and pCAMBIA- dbr2 showed positive blue spot where coefficient of variance was less than 5%. PCR analysis for genomic DNA of transformed A. annua showed a positive result of inserted dbr2 recombinant indicated by migration profile and direct sequencing analysis. It could be concluded that pCAMBIA- dbr2 construct and transformation into A. annua have been successfully performed.

The development of synthetic biology will shape the new era of science and technology. It is an emerging bioengineering technique involving genetic engineering which can alter the phenotype and behavior of the cell or the new product. Synthetic biology may produce biomaterials, drugs, vaccines, biosensors, and even a recombinant secondary metabolite used in herbal and complementary medicine, such as artemisinin, a malaria drug which is usually extracted from the plant Artemisia annua. The power of synthetic biology has encouraged scientists in Indonesia, and is still in early development. This paper also covers some research from an Indonesian research institute in synthetic biology such as observing the production of bio surfactants and the enhanced production of artemisinin using a transient expression system. Synthetic biology development in Indonesia may also be related to the iGEM competition, a large synthetic biology research competition which was attended by several universities in Indonesia. The ...

Artemisinin, a secondary metabolite from Artemisia annua L. is a sesquiterpene lactone that has antimalaria activity but produced at low quantities by the plant. Low levels of artemisinin in the plant is related to the biosynthetic pathways influenced by specific enzymes that play role in the formation of artemisinin. Farnesyl diphosphate (FDP), which is the main precursor of artemisinin, also known as the precursor for the formation of sterols. Compared with the other compounds, sterol biosynthetic pathway is the biggest competitor of the artemisinin production since sterols are needed by plants to regulate membrane fluidity and permeability. This research aimed to study the effect of terbinafin and DMSO as sterol synthesis inhibitors on the regulation of the artemisinin biosynthetic pathway by analyzing the expression level of two genes linked, squalene synthase (SQS) and amorpha-4,11-diene synthase (ADS) using quantitative PCR (qPCR) and the amount of artemisinin is determined using high performance liquid chromatography (HPLC). The results showed that at mRNA levels, terbinafin 30 μM had no significant effect on SQS and ADS expression levels, but it increased the amount of artemisinin at 50 hours incubation up to 1.36 times higher than control. DMSO increased the expression level of ADS up to 3-4 times and increased the content of artemisinin up to 2.42 times higher than control at 50 hours incubation time

Luwak coffee is a highly-priced coffee produced exclusively by the palm civet or luwak ( Paradoxurus hermaphrodites ssp.). The purpose of this study was to determine the diversity of culturable bacteria in the gastro intestinal tract of luwak. The bacterial isolates were phenotypically characterized by their morphology and molecularly by analysis of their1,500bp 16s rDNA sequence. The results showed that Enterobacter cloacae and Lactobacillus brevis were found all over luwak’s digestive tract . Enterobacter cloacae was the most common species. The most diverse bacterial population was found in small intestine. Seven bacterial generawere successfully identified from the small intestine and colon, compared to only five genera found in the stomach.

Amplification of putative potBCD genes from Serratia rubidae was conducted by PCR using a pair of primers FERC and RERC. A fragment with ~1800 bp size was ligated to pGEM-T Easy vector then cloned to competent Escherichia coli DH5α. The recombinant plasmid was sequenced using SP6, T7 and two internal primers, FPC and RPC. A sequence similarity search and analysis was performed with the BLASTN program. The sequence was found to have 83% similarity to potABCD genes from E. coli. Those genes encoded a spermidine-preferential uptake system that consists of four kinds of protein: PotA is a membrane-associated ATPase, PotB and PotC are transmembrane proteins that form channels, and PotD is a periplasmic substrate-binding protein. Alignment analysis showed that the isolated clone consisted of potB (partial), potC (full length) and potD (partial). The sequences for potBCD genes from S. rubidae are not available in the NCBI database. Furthermore, we have submitted this sequence, potBCD from S. rubidae , on GeneBank with Acc. number FJ447342.

Human papillomavirus (HPV) type 52 is the most prevalent type for causing cervical cancer in Indonesian population. Cervical cancer becomes the most common cancer suffered by Indonesian women. Prevention of HPV infection can be achieved using HPV virus-like particle (VLP) vaccine derived from L1 major capsid protein. This study aimed to clone and analyze HPV-52 L1 gene. DNA obtained from biopsy of a cervical cancer patient was amplified using specific primers designed from Asian originated HPV-52 L1 gene available in the GenBank. The isolated HPV-52 L1 gene sequence was submitted to GenBank with accession number [KF225497]. Expression of HPV-52 L1 gene was performed using pRSET/EmGFP Escherichia coli expression vector. We analyzed and compared the HPV-52 L1 gene expressions from recombinant E.coli BL21 (DE3) that had been induced for 3 hours with 1 mM IPTG and without induction. The protein was expressed in insoluble form. We performed the following bioinformatic analyses: construction of phlyogenetic tree, T-cell epitopes prediction and 3D proteins structure modelling. We utilized the following softwares: MEGA5 for phylogenetic tree, IEDBann for MHC prediction, CLC DNA Workbench 6.5 for hydrophobicity analysis and PDB-Viewer Deep for 3D protein structure analysis. The phylogenetic tree which was developed based on [KF225497] sequence showed that it shared a branch with Asian countries (Philippines and Thailand). The deduced amino acid sequences of the predicted epitopes that were consistent in all of the programs were 259GTLGDPVPGDLYIQGS274 and 345KKESTYKNE353. This information may be useful to design diagnostic strategies and vaccine suitable for Indonesian population.

Klebsiella oxytoca produces sucrose isomerase which catalyses the conversion of sucrose to isomaltulose, a new generation of sugar. From the previous study, palI gene from Klebsiella oxytoca was succesfully isolated from sapodilla fruit (Manilkara zapota). The full-length palI gene sequence of Klebsiella oxytoca was cloned in E. coli DH5α. The deduced amino acid sequence shows 498 residues which includes conserved motif for sucrose isomerisation 325RLDRD329 and 97% identical to palI gene from Klebsiella sp. LX3 (GenBank:AAK82938.1). This fragment was succesfullly ligated into the expression vector pET-32b using overlap-extension PCR and cloned in Escherichia coli BL21 (DE3) pLysS. DNA sequencing result shows that palI gene of Klebsiella oxytoca was inserted in-frame in pET-32b. This is the first report on cloning of palI gene from Klebsiella oxytoca.

Leaf colors of a plant can be used to identify stress level due to its adaptation to environmental change. For most leaves green-related colors are sourced from chlorophyll a and b. Chlorophyll concentration is normally measured using a spectrophotometer in laboratory. In some remote observation places, it is impossible to collect the leaves, preserve them, and bring them to laboratory to measure their chlorophyll content. Based on this need, measurement of chlorophyll content is observed through its color. Using CIE chromaticity diagram leaf color information in RGB is transformed into wavelength (in nm). Paddy seed with variety name IR-64 is used in observation during its vegetation stage t (age of 0-10 days). Light exposure time {\tau} is chosen as environmental change, which normally should be about 12 hours/day, is varied (0-12 hours/day). Each day sample from different exposure time is taken, its color is recorded using HP Deskjet 1050 scanner with 1200 dpi, and its chlorophyll content is obtained from absorption spectrum measured using Campspec M501 Single Beam UV/Vis Spectrophotometer after it is rinsed in 85 % acetone solution and the information from the spectrum is calculated using Arnon method. It has been observed that average wavelength of leaf color {\lambda}avg is decreased from 570.55 nm to 566.01 nm as is measured for t = 1 - 10 days with {\tau} = 9 hours/day, but chlorophyll concentration C is increased from 0.015 g/l to 3.250 g/l and from 0.000 g/l to 0.774 g/l for chlorophyll a and b, respectively. Other value of {\tau} gives similar results. Based on these results an empirical relation between concentration of chlorophyll a Cc-a and its wavelength {\lambda}avg can be formulated., Comment: 8 pages, 5 figures, conferene paper to be presented in Padjadjaran International Physics Symposium 2013 (PIPS 2013), 7-8 May 2013, Jatinangor, Indonesia