Tri Joko Raharjo

Determination of Porcine Contamination in Laboratory Prepared dendeng Using Mitochondrial D-Loop686 and cyt b Gene Primers by Real Time Polymerase Chain Reaction

The identifications of species in meat products have created interests since these foods became the target of forgery and fraud in the market. The presence of pork in food products is not allowed for the Muslim community. Hence, an analysis is necessary to detect the presence of pork in processed meat products, such as in dendeng (dried meat) product. Real time polymerase chain reaction using mitochondrial displacement loop686 and cytochrome b (cytb) gene primers was used to identify specific pork DNA among other four types of DNA species; namely beef, chicken, goat, and horse. This method was also used to identify pork DNA in the laboratory processed pork-beef dendeng as well as commercial dendeng from market. The results showed that real time polymerase chain reaction using displacement loop686 and cytb gene primers were able specifically to distinguish between pork DNA and the other species. The lowest concentration of 0.5% of pork DNA in a mixture of pork-beef processed products of dendeng was able to be detected by both primers with the product amplification of 114 and 134 bp (base pair) for the displacement loop686 and 149 bp for cytb gene, respectively. High sensitivity was also obtained when both primers were applied with the lowest detection limit of 5 pg/µL pork DNA. The results of the six commercial dendeng amplification using both primers showed no amplified products present, meaning that these products do not contain porcine DNA.

A Coarse-Grained Molecular Dynamics Simulation Using NAMD Package to Reveal Aggregation Profile of Phospholipids Self-Assembly in Water

The energy profile of self-assembly process of DLPE, DLPS, DOPE, DOPS, DLiPE, and DLiPS in water was investigated by a coarse-grained molecular dynamics simulation using NAMD package. The self-assembly process was initiated from random configurations. The simulation was carried out for 160 ns. This study presented proof that there were three major self-assembled arrangements which became visible for a certain duration when the simulation took place, that is, liposome, deformed liposome, and planar bilayer. The energy profile that shows plateau at the time of these structures emerge confirmed their stability therein. Our findings have highlighted the idea that liposomes and deformed liposomes are metastable phases which eventually will turn into planar bilayer, the stable one.

COMPARATIVE EVALUATION OF CONVENTIONAL VERSUS RAPID METHODS FOR AMPLIFIABLE GENOMIC DNA ISOLATION OF CULTURED Azospirillum sp. JG3

As an initial attempt to reveal genetic information of Azospirillum sp. JG3 strain, which is still absence despite of the strains’ ability in producing valued enzymes, two groups of conventional methods: lysis-enzyme and columnkit; and two rapid methods: thermal disruption and intact colony were evaluated. The aim is to determine the most practical method for obtaining high-grade PCR product using degenerate primers as part of routine-basis protocols for studying the molecular genetics of the Azospirillal bacteria. The evaluation includes the assessment of electrophoresis gel visualization, pellet appearance, preparation time, and PCR result of extracted genomic DNA from each method. Our results confirmed that the conventional methods were more superior to the rapid methods in generating genomic DNA isolates visible on electrophoresis gel. However, modification made in the previously developed DNA isolation protocol giving the simplest and most rapid method of all methods used in this study for extracting PCR-amplifiable DNA of Azospirillum sp. JG3. Intact bacterial cells (intact colony) loaded on electrophoresis gel could present genomic DNA band, but could not be completely amplified by PCR without thermal treatment. It can also be inferred from our result that the 3 to 5-min heating in dH2O step is critical for the pretreatment of colony PCR of Azospirillal cells.

Partial purification and biochemical characterization of extracellular lipase from Azospirillum sp. JG3 bacteria

An extracellular lipase from Azospirillum sp.JG3 bacteria was partially purified by ammonium sulfate fractionation. Production of crude extract lipase in Nutrient Broth medium fortified with 1 % olive oil as inducer, fractionation of lipase using ammonium sulfate, and characterization of the highest specific activity of lipase fraction have been achieved in this study. The ammonium sulfate lipase fraction which has highest specific activity is F60 with its value is 12,283.33 U/mg protein. The optimum lipase activity of F60 was achieved at 40° C and pH 7.0. The addition of 1.0 mM EDTA strongly decreased its activity. It considered that this lipase was a metalloenzyme. The addition of 1.0 mM Ca2+ solution showed that this metal ion increased lipase activity, it indicated that Ca2+ ion is an activator of this lipase. Whereas addition of Mg2+, Zn2+, Co2+, and Cu2+ metal ions decreased its activity. Effect of surfactants and commercial detergents addition towards lipase activity showed that all treatments decr...