RNAlater Tissue Collection

Telomeres are the non-coding, repetitive DNA sequences at the ends of chromosomes that are required for chromosome positioning in the nucleus and for DNA protection from recombination, degradation, and replicative damage. In cellular senescence research, the length of the telomeres can be used as a marker for biological age, where older cells have shorter telomeres. With the hope that various tissue archives could be used in aging studies, Dr. Koppelstaetter and colleagues sought to assess whether different types of sample preservation methods would still allow accurate determination of telomere length by a commonly accepted technique used with small sample sizes (qPCR: ratio of the amount of amplicon from telomere sequences to that of a single copy gene).

Normal renal cortical tissue was obtained from a kidney removed due to carcinoma. DNA immediately isolated from one piece served as the reference sample. Triplicate samples, preserved in either RNA later or 4% formaldehyde, were stored in darkness at room temperature for one of six different time periods (range: 3–168 hr). DNA was isolated from the samples and qPCR assays were run with telomere specific primers, as well as primers specific for three single copy genes (36B4, GAPDH, b-actin).

At all time points, RNA later-treated samples gave results essentially identical to the reference sample for relative telomere length. In contrast, formaldehyde fixation caused an apparent time-dependent increase in relative telomere length, which was mainly due to inconsistencies in the amplification of the single copy gene PCR products. Formaldehyde, which is known to denature double-stranded DNA at AT-rich regions, destabilized genomic DNA in the samples, so that amplification of the longer PCR products assessed in this study had the most variability. In contrast, RNA later has been proven to stabilize RNA and is now shown to preserve DNA within tissues. For additional details, see Koppelstaetter, et al. [1].