Effects of Postmortem Interval and Preservation Method on RNA Isolated from Field-Preserved Avian Tissues

Abstract

Studies of gene expression and transcriptome profiling often require the isolation of high-quality mRNA. Here, we examined how preservation method and the lag between collection and processing (postmortem interval; PMI) affect RNA quality and yield in field-collected samples. We subjected tissue samples of 11 House Sparrows (Passer domesticus) to two preservation treatments (flash freezing in liquid nitrogen versus treatment with Ambion's RNAlater prior to freezing) and eight PMI treatments (intervals ranging from 0 to 360 min). Mean RNA yields were significantly higher for flash-frozen tissues. Total RNA yield, as measured by a UV-vis spectro- photometer (Nanodrop), was not negatively associated with PMI in either preservation treatment, and we were able to extract RNA of quality (28S:18S fluorescence ratio >0.9) sufficient for most gene-expression applications from specimens that were held at ambient temperature for up to 6 hr postmortem. These results demonstrate that suffi- cient quantities of high-quality RNA can be extracted from avian tissues collected under typical field conditions. Although RNA remains stable for several hours postmortem, other studies have demonstrated that the transcrip- tional profile changes as a function of PMI. To increase the value of tissue samples for RNA studies we recommend that collectors include with a specimen detailed notes on the duration of the PMI as well as other extrinsic (e.g., time of day, temperature) and intrinsic (e.g., age of bird, sex) factors that influence levels of gene expression.