Simple and efficient preservation of fish environmental RNA in filtered water samples via RNAlater

Abstract

Abstract Environmental RNA (eRNA) analysis has recently received attention as a better means to infer the physiological status of a community and living biotic assemblages than environmental DNA (eDNA). However, eRNA is thought to be degraded more rapidly than eDNA, increasing the risk of false-negative detection and complicating large-scale eRNA sampling in the field. In addition, the need for a deep freezer (− 80°C or below) further limits the practical application of eRNA analysis in places that are not accessible by vehicle. Here we focused on two types of reagents (RNAlater and LBP buffer) and assessed their performance for eRNA preservation. We found that very high concentrations of ayu (Plecoglossus altivelis) eRNA were quantifiable from filter samples collected from an aquarium by using RNAlater preservation at − 20°C for at least a week. Compared with the sample stored at − 20°C without any preservative, the filter samples preserved in RNAlater had eRNA concentrations that were tens to hundreds of times higher. Although further technical refinement is needed, our findings have provided valuable information to enhance the methodology for improving eRNA quality and quantity in environmental samples. This will boost the practical application of eRNA-based meta-transcriptomics targeting macro-organisms.