Optimization of DNA extraction and the position of mosquito Species from southeast minahasa in North sulawesi using NADH dehydrogenase Gene and Cytochrome oxidase Sub Unit 1 Gene
Caroline Manuahe, Mokosuli Yermia Semuel and Verawati Ida Yani Roring
A research has been done that aimed to optimize the extraction of mosquitoes’ DNA and to find out the characteristic of NADH dehydrogenase as well as cytochrome oxidase sub unit 1 (CO1) of mitochondrial genes to reveal the position of mosquito species from Southeast Minahasa. Mosquito samples retrieved from Southeast Minahasa on four villages in two subdistricts, namely Tombatu and Toluaan. The reserach was conducted in tree stages. The first stages was extraction of DNA, followed by NADH dehydrogenase gene and cytochrome oxidase sub unit 1 gene amplification and the last stage was electrophoresis and sequencing. The results showed that extraction of mitochondrial DNA from the mosquito using a fore leg organ with modified proteinase-K immersion for 24 hours was the best treatment. Mitochondrial DNA gene amplification N5 (NADH dehydrogenase) did not work while CO1 gene was successfully amplified on modified PCR condition. Mosquitoes’ CO1 gene amplified with a length of 702 bp. The frequency GC is 228 or 33.4%. The most numerous amino acid codified are asparagine and serine. The posisition of mosquito samples based on CO1was nearest to Armigeres subalbatus [KJ 410334.1] with 89% similarity level, 11% differences of nitrogen base of CO1 gene, therefore the mosquito from Southeast Minahasa conctructed a new node at phylogeny trees.
Caroline Manuahe, Mokosuli Yermia Semuel, Verawati Ida Yani Roring. Optimization of DNA extraction and the position of mosquito Species from southeast minahasa in North sulawesi using NADH dehydrogenase Gene and Cytochrome oxidase Sub Unit 1 Gene. J Entomol Zool Stud 2016;4(4):498-508.