Dengue virus (DENV) is a significant pathogen emerging worldwide as a cause of infectious disease. Antidengue treatments are urgently required to control the emergence of dengue. DENV is a mosquito-borne disease responsible for acute systemic diseases and serious health conditions. DENVs were distributed in the tropical and sub-tropical areas and transmitted to humans by
Dengue Hemorrhagic Fever (DHF) is caused by dengue viruses that belong to Flaviviridae. The disease is known to be caused by 4 types of dengue viruses, namely DENV-1, DENV-2, DENV-3, and DENV-4 associated with antigenic. Dengue virus is a virus RNA that causes illness with clinical manifestations of Dengue Fever, Dengue Hemorrhagic Fever and Dengue Shock Syndrome. The aim of research was to determine the effectiveness of dimethyl sulfoxide, acetone, and ethanol 70% as precipitation solvent in the process of RNA isolation. The method used was Reverse Transcription - Polymerase Chain Reaction (RT-PCR) and Polymerase Chain Reaction (PCR) with specific primers for dengue virus type 1 (DENV-1). RNA isolation can be done easily using an RNA Isolation Kit. Use of RNA Isolation Kit results in a purer RNA isolate from contaminants and from RNA degradation. In generally the isolation is using cold ethanol / alcohol with concentration 90-95%. Ethanol / Alcohol does not dissolve RNA and light density of alcohol lighter than water makes RNA rise and hover on the surface. In RNA isolation solvent precipitation that used are acetone, ethanol 70%, and DMSO. In qualitative RNA measurements using agarose gel electrophoresis and was examined under the UV light-illuminator and quantitative RNA measurements using Nanodrop spectrophotometry with absorbance ratio at 260/280 and 260/230 showed a good result indicated by the appearance of the band on electrophoresis results in PCR. While the measurement quantitatively is showed that there was still protein contamination but the results are quite good because it does not much different from the ratio set in the reference. Acetone, ethanol 70%, and DMSO can be used as a substitute of 96% ethanol in the process of RNA isolation in DENV-1 virus and can also be applied to other dengue virus because the structure of the 4th antigen serotype is very similar one with the other and no effect.
Dengue virus distributed in tropical and subtropical regions in the world. DENV viruses are transmitted between humans primarily by Aedes aegypti and Aedes albopictus mosquitoes and are endemic in most areas in which the vectors occur. Four serotypes of dengue virus are DENV-1, DENV-2, DENV-3 and DENV-4. DENV-2 is comprised of six genotypes. Ethanol precipitation is a commonly used technique for concentrating and de-salting nucleic acids (DNA or RNA) preparations in aqueous solution. RNA isolation by combining Guanidinium thiocyanate and phenol reported has been reported. In this report, we investigated RNA isolation from DENV-2 using QIAamp Mini Kit with 2-Isopropanol, Methanol, Chloroform precipitation solvent. Electrophoregram showed DNA band as the result of RNA isolation with methanol and 2-isopropanol are produced quite well. Dna band of the of RNA isolation with chloroform solvent has the lowest intensity than methanol and 2-isopropanol. This study showed that methanol and 2-isopropanol can used as precipitation solvent for isolating RNA.
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