Quantification and DNase treatment of RNA Sample Clauses

Quantification and DNase treatment of RNA. DNase treatment of isolated RNA removed genomic contaminant DNA from the RNA sample. DNA-free™ kit (Applied Biosystems) was used to treat RNA according to the manufacturer’s instructions. Briefly, 5 μl of 10× DNase buffer and 1 μl rDNase were added to the RNA and mixed gently. The mixture was then incubated at 37 oC for 30 min. To stop the reaction, 5 μl of DNase inactivation reagent was added while gently mixing and incubating for 2 min at room temperature. The DNase and inactivation reagents were removed by centrifugation (10, 000 g for 1.5 min) and the resultant RNA was transferred to a new tube. RNA concentration was determined with Nanodrop spectrophotometer (Thermo Scientific) using the manufacturer’s software. The pedestal was first cleaned using lint-free cloth. The machine was self-caliberated after each use and zeroed using appropriate buffer in which the nucleic acid is stored. 1.5µl of each undiluted total RNA samples was analysed, recording the concentration in ng/µl. Also, the OD260/OD280 readings were between 1.8 and 2.1 meaning that there is little or no protein contamination in a RNA sample.
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