Abstrait

Quality variations of extractable RNA in frozen samples of oral cancer (21 y); optimization of the extraction protocol

Cintia Chamorro M, Mario Perez-Sayans, Elena Padi

Access to sufficient tissue samples to develop a RNA study is complicated and in most cases a biobank is needed to preserve samples. Due to this it is necessary to assess how frozen tissue samples are influenced by the time they have been kept in storage. We have extracted RNA from 19 samples of oral squamous cell carcinoma (frozen from 1994 to 2015) and 31 samples of oral mucosa from healthy patients (extracted in 2015). The miRVana miRNA Isolation Kit (Ambion) was used to extract RNA. The extraction was performed following the manufacturer's instructions, except for the first part, sample disruption, for which we used a modified process. We determined the concentration of all samples, using NanoDrop 2000; while the RNA Integrity Number (RIN) was determined by NanoChip 6000 and the Agilent 2100 Bioanalyzer. The distribution of RIN and concentration is the same among location categories (p=0.696 and p=0.774, respectively). Results revealed a mean of RNA concentration lower in samples conserved for longer periods of time (p=0.006) but the RIN did not change with time storage (p=0.142). Our method was efficient to disrupt the tissue and to extract total RNA with good RIN and enough concentration. Although the concentration obtained from frozen tissue samples decreased with the passage of time, the RIN did not change.