Indonesian Journal of Biotechnology
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Biochemical properties of crude extracellular proteases from Chromohalobacter salexigens BKL5 and Micrococcus luteus 11A
No full textIn this work, we have reported an enzymatic activity and biochemical properties of extracellular proteases from Chromohalobacter salexigens BKL5 and Micrococcus luteus 11A. C. salexigens BKL5 and M. luteus 11A were previously isolated from Bledug Kuwu mud volcano and dietary industry wastewater treatment, respectively. Both bacterial strains were able to produce extracellular proteases, when grown on minimal agar medium supplemented with 1% of skim milk. Proteolytic indexes of C. salexigens BKL5 and M. luteus 11A were 2.5±0.14 and 2.9±0.42, respectively. Both extracellular proteases exhibited optimum enzymatic activity at pH 7, with specific activity of C. salexigens BKL5 was 13.3% higher than that of M. luteus 11A. Optimum temperature for enzymatic activity of both proteases was 45°C. Metal cofactor preferences assay showed that extracellular protease from C. salexigens BKL5 preferred Zn2+, meanwhile extracellular protease from M. luteus 11A mainly preferred Ca2+ ion. Metal cofactor preferences assay also suggested that crude extracellular protease from C. salexigens BKL5 was categorized as metalloprotease, meanwhile crude extracellular protease of M. luteus 11A was common neutral protease. The enzymatic stability assay against various salt concentrations showed that crude extracellular protease from C. salexigens BKL5 was more stable than that of M. luteus 11A
The growth improvement of Grammatophyllum scriptum (Lindl.) Bl. in vitro plantlet using photoautotrophic micropropagation system
No full textTo improve the growth of Grammatophyllum scriptum (Lindl.) Bl. in vitro plantlet, a photoautotrophic micropropagation system (PMS) was developed by growing in vitro plantlet on VW medium with varying concentration of sucrose (0, 5, 10, and 20 g/L) and additional carbon dioxide from the air (bottle covered with cap or filter). The result showed that the leaf length would increase up to 6.5 cm with PMS and it would keep growing by the adding of 5 g/L sucrose. Average number of leaves increased by 6.7 strands with PMS and the addition of sucrose increased the average quantity of leaves up to 7.7 strands. Average number and root length would increase with PMS and would even increase more with 5 g/L sucrose addition. PMS with 5 g/L sucrose can increase chlorophyll a and b concentration. The number of stomata per unit area in PMS was lower than closed culture. This shows that PMS can increase the growth of G. scriptum in vitro plantlet and the growth increase would be effective if it is combined with sucrose addition
Study of creatinine transport through chitosan/pectin/poly(vinyl alcohol) blend membranes
No full textCreatinine was final product of creatine metabolism inskeletal muscle. Increasing of creatinine showed decreasing of kidney function. Kidney fuction decreased could be treated by hemodialysis (HD). One of the natural polymer was cellulose which often used as hemodialysis membrane. Chitosan as natural membrane was used as membrane in this research because the structure was almost similar to cellulose. Chitosan was complexed by pectin and poly(vinil alcohol) (PVA) to fixed mechanical characteristic of membrane. The objective of this research were to synthesize, characterize, and know efficiency of creatinine transport using chitosan/pectin/PVA blend membranes. The functional groups of synthesized membrane were characterized by FTIR spectrophotometer. Efficiency of optimum creatinine transport was known by using membrane with chitosan and pectin ratio70:30 while PVA used 0.5%; 1.0% and 1.5%. The source and acceptor phase resulted were complexed by picric acid and analyzed by Ultraviolet-Visible (UV-Vis) Spectrophotometer. The result of membrane synthesized which was analyzed by FTIR Spectrophotometer shows that there is a band broadening on wave number 3448.72 cm-1. It is indicated that there are an overlapped stretching of hydrogen bond -OH on PVA and NH2 on chitosan. The optimum creatinine 70 ppm transported using membrane with 1.5% PVA addition is 59%.
Method development for detection of E545A mutation PIK3CA gene in breast cancer patients using Tm Shift SYBR Green I qPCR
No full textE545A is one of the point mutations, and its frequency is high in PIK3CA gene (3.8%), particularly breast cancer patients in Singapore (13.8%) and Mexico (11.5%). In addition to induce breast cancer, the mutation also caused resistance of anti-HER2 in HER2 cancer subtype. The tremendous effect of this mutation was not supported by affordable detection method. This study aimed to develop a feasible and sensitive method of E545A detection. The developing method used Tm and Ct to identify samples. Based on optimization, the best condition was obtained at optimization 2 at annealing temperature of 65°C. At this condition, Tm and Ct of each sample were (a) exon 9 (78.4°C and 13.005±0.007) and (b) E5454A (80.4°C and 10.07±0.1). This method also demonstrated good precision as observed in variance coefficient of intra and inter assay (0). Thus, method for E5454A detection mutation was successfully developed
Determination of allelopathic potential in mahogany (Swietenia macrophylla King) leaf litter using sandwich method
No full textThe sandwich method is a reliable screening bioassay that can be utilized to investigate allelopathic activity of leaf litter leachates. Screening the allelopathic potential of mahogany (Swietenia macrophylla King) leaf litter in plant–plant interaction using the sandwich bioassay method has not been reported. The research objectives were to determine and categorize allelopathic potential of S. macrophylla leaf litter using the sandwich bioassay method, and to determine specific activity (EC550). S. macrophylla leaf litter. The results showed that S. macrophylla leaf litter exhibited strong allelopathic activity when compared with 46 leaf litter species and was included in the top ten of allelopathic leaf litter species. Increasing S. macrophylla leaf litter concentration was concomitant with inhibition of radicle lettuce seedling growth compared with the control. According to the linear regression analysis, the effective concentration (EC50) of S. macrophylla was estimated to be 3.25 mg D.W. eq. mL-1 and was considered to have strong growth-inhibitory activity on lettuce radicle elongation. The results suggest the possibility of allelopathic potential of leaf litter in plant–plant interaction under S. macrophylla trees
The synergistic effect of doxorubicin and ethanolic extracts of Caesalpinia sappan L. wood and Ficus septica Burm. f. leaves on viability, cell cycle progression, and apoptosis induction of MCF7 cells
No full textCaesalpinia sappan L. and Ficus septica Burm. f known asa potential plant with wide variety of medicinal properties, including anticancer. Present study was aimed to explore cytotoxic effect ofsappan wood (ECS) and awar-awar leaves (EFS), and its combination with doxorubicin (dox) on MCF-7 cells focusing on cell cycle progression and apoptosis induction.The result of MTT assay showed that single treatment of ECS and dox performed cytotoxic effect with the IC50 value of 32 µg/mL and 6 µM respectively, while EFS performed low cytotoxic effect with the IC50 value of 282 µg/mL. The combination of ECS with EFS and doxorubicin showed synergistic cytotoxic effect. Flow cytometry analysis revealed that combination of ECS (16 µg/mL) with EFS (8 µg/mL) and doxorubicin (2 µM) induced apoptosis, and cell accumulation at sub-G1 and G2/M phases.Immunoblotting assay confirmed the apoptosis induction of this combination through increasing of cleavage of PARP-1. Based on these results, the synergistic cytotoxic effect of this combinationwas through G2/M phase accumulation and apoptosis inductionand potentially to be developed as co-chemotherapeutic agent
The genetic variations and relationship of Madura tobacco (Nicotiana tabacum L.) based on molecular characteristics
No full textMadura has at least 22 genotypes of local tobaccos (Nicotiana tabacum L.). This diversity could potentially produce new genotype of tobaccos with superior characters. However, information of the genetic diversity of Madura tobaccos is still limited. The aim of this study was to determine the genetic variation and relationship of 24 genotypes of Madura tobaccos with Random Amplified Polymorphic DNA (RAPD) analysis. In this research we were used 6 single primers for amplification: (OPA-18, OPB-12, OPB-14, OPC-1, OPC-8 and OPC-19) and 2 mixture primers ((OPB-12+OPC-8) and (OPC-1+OPC-19)). Genetic similarity and clustering was analyzed with Unweighted Pair Group Method Arithmetic (UPGMA) method with Numerical Taxonomy and Multivariate Analysis System (NTSYS) version 2.10 software. From this research we found that OPA18425, OPB12450, OPC8500, (OPC19+OPC1)550 and OPC8800 can be used as specific markers. Polymorphic bands percentage with mixture primers was relatively equal with single primers (<60%). The dendogram showed that Madura tobacco genotypes consist of 2 main clusters: cluster A (22 genotypes) and cluster B (2 genotypes: Bukabu Sa’ang and Prancak-95). Madura tobaccos had high genetic similarity between genotypes ranging from 0.80-1.00
Evaluation of rapid detection kit against avian influenza A virus and H5 subtype for field Sample
No full textAvian influenza virus is poultry viral disease, which causes high economic losses. Various efforts have been made to control the disease. One effort is required screening fast and precise diagnostic test. This study was aimed to determine the potential of rapid test kit of AIV/H5 Anigen Rapid Test for the detection of AI virus types A and subtype H5 in the field. Some tests were carried out, e.g.: the potential test, cross-reaction test, sensitivity and specificity test. Potency test was done to evaluate potential of detection limits of the kit, by having the test of serial dilution of AI virus positive control. Cross-reaction test was done to detect antigens other than AI virus H5N1, e.g.: IB virus of Massachuset strain, IBV strain 4-91, Newcastle Disease virus, and Escherichia coli. Sensitivity and specificity test were applied to the filed samples which clinically and laboratory were confirmed as H5N1 positive. To confirm the result of rapid test was then being done by reverse transcriptase polymerase chain reaction. Based on these results it can be concluded that, Anigen Kit AIV/H5 Ag Rapid Test can detect antigen-containing samples having AI virus HA titer up to 26of type A virus, and up to 25 for subtype H5 virus. Anigen Kit AIV/H5 Ag Rapid Test showed no cross-reactions with Infectious Bronchitis virus, Newcastle Disease virus, and Escherichia coli. Anigen A Rapid Test Kit AIV Ag showed a sensitivity of 50% and specificity of 100%, while Anigen Ag Rapid Test Kit AIV/H5 showed a sensitivity of 25% and specificity is 100%
Analysis of whole cell protein profiles by SDS-PAGE to identify indigenous cellulose-producer acetic acid bacteria
No full textThis study was carried out to analyze the suitability of the identification of four indigenous cellulose-producing acetic acid bacterial isolates (ANG29, KRE65, ANG32 and SAL53) based on the analysis of whole cellular protein profiles against identification based on phenotypic traits. Whole cellular protein profiles were determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) method. The whole cellular protein profiles obtained from sample isolates, were compared with reference isolates for species identification. The results showed that based on visual observations can be determined as much as 12 bands of protein with a molecular weight of 19,099 KDa up to 132.182 KDa. Based on the analysis of protein bands were detected visually, fourth indigenous cellulose- producing acetic acid bacterial isolates in the study had a higher similarity profile to the reference strain Gluconacetobacter xylinus BTCC 769 compared with other reference strains namely G. hansenii NBRC 14820T. This condition is consistent with the results of the identification of fourth cellulose producing acetic acid bacterial isolates based on phenotypic traits. Thus, the whole cellular protein profiles by SDS-PAGE technique can be used as a one of method to identification of cellulose producing acetic acid bacterial isolates
Identification of excretory secretory (ES) liquid antigen protein Fasciola gigantica with polyethilene glycol (PEG) separation
No full textFasciola gigantica diagnosis usually performed by detection of worm eggs presence in the feces,but this conventional method has many disadvantages. Early diagnosis (early detection) cannot be performed in conventional methods because the worms in the host's body began to lay eggs at the age of 8–12 weeks of patency. The current detection method that is based on antibodyantigen reactions using excreted/secreted (ES) liquid by adult F. gigantica, is believed to be used for the early detection of fasciolosis. This study aimed to characterize the antigenic components of F. gigantica extretory/secretory products that could be used as a vaccine candidate development for early fasciolosis diagnostics. ES products were separated by PEG4000 at various concentrations (8%, 16%, 24%), then precipitates (pellets) obtained were dialyzed and characterized using SDSPAGE and Western blotting. Results from SDSPAGE showed that there were 18 proteins bands with 7–70 kDa molecular weights. Western blotting on pellets derived from PEG separation at various concentrations affirmed that the proteins of 50, 25 and 20 kDa were antigenic at 8% PEG concentration, the 25 kDa and 50 kDa were antigenic at 16% PEG concentrations and the 25 kDa was antigenic at 25% PEG concentratio