Development Of Dna Based Molecular Methodologıes Use Of Dose
Estımatıon In Irradıated Fısh
In this study, RAPD and real-time PCR were used for the development of metodologies that can able to estimate irradiation dose was done and distinguishing between irradiated and non irradiated fish. Trout (Oncorhynchus mykiss) samples were exposed to radiation doses of 0.250, 0.500, 1, 3, 5, 7 and 9 kGy in gamma cell. DNAs were extracted from irradiated samples before and after storage. Primers were designed on regions with different lengths of both nuclear and mitochondrial DNA and each primer was used to amplify DNA of irradiated samples. The amplicon curves of mitochondrial and nuclear DNA and the values of the correlations among the amplicon curves were obtained. 519 bp reagon of the 18S RNA gene on nuclear DNA provided the best correlation values. Radiation doses applied to the fish fillets were estimated using the standard curve data obtained from correlation values, and DNA damage caused by each dose was calculated. ERP primers were designed to make randomly amplifications on the DNA of the irradiated samples, and RAPD PCR was applied and obtained the agarose gel profiles of amplicons. In addition, DNA fragmentation occurring in each dose was determined by Komet test for the purpose of verification of molecular methodologies developed in this study. As a result, two molecular methodologies enabling to easily qualitative analysis of irradiated trout meat and enables quantitative estimations of the administered dose were developed. These methodologies allow the analysis of the trout flesh irradiated and stored for a period of three months up to the dose limit of around 0.5 kGy.
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