Long-term Stability of Total RNA in RNAstable^® as Evaluated by Expression Microarray

Abstract

Storage of labile RNA in laboratories is accomplished through ultra-low freezing of the nucleic acids. This however requires expensive freezers, convenient storage, reliable electrical power, and increased shipping costs, thereby making it a less viable option. Biomatrica (San Diego, CA) has created RNAstable Ò , a stabilization reagent that is used to store RNA in a dehydrated state at room temperature (RT) and protects the RNA from degradation. Our objective was to investigate the sequence integrity and suitability of RNA when stored in RNAstable at extended time periods and at varying temperatures through use of Illumina and Agilent RNA expression microarrays. We observed in Bioanalyzer electropherograms that total RNA extracted from 293 cells stored at RT in RNAstable for 4.5 and 11.5 months is similar in quality to RNA stored at -80°C. Illumina mRNA expression array QC metrics and gene expression patterns from RNAstable-protected RNA, in contrast to RNA stored without RNAstable, correlated well with those of freezer controls. Significantly, when RNA was stored in RNAstable at 45°C for 4.5 months, equivalent to 22 months RT storage, RNA quality, microarray probe signal intensities, probe detection rates, and expression profiles remained similar between RNAstableprotected RNA at RT and the -80°C controls. At 10.5 months, miRNA levels were compared among the storage conditions using miRNA expression arrays. Here too we found strong concordance between miRNA expression patterns when total RNA was stored in RNAstable or at -80°C. Further, Bioanalyzer electrophoresis of RNAstable-protected samples stored at RT for a relative total of 33 months or 50.5 months showed comparable integrity scores to those of -80°C controls. We conclude that use of RNAstable holds promise as an effective stabilization reagent for total RNA and should be useful in situations where shipping and storage options are limited resources.

Respiratory tract samples collected from patients in a region of Quebec, Canada, indicate the absence of early circulation of SARS-CoV-2 infection

“…RNA extracts degrade over time, but we used purified RNA extracts that were stored at 80°C, and we performed SARS-CoV-2 RNA testing 3-4 months after the RNA was extracted. In this environment, RNA can be stable for more than 50 months (12). Our PCR assay included an internal control to confirm the stability of frozen extracts.…”

Distinct promoter regions of the oxytocin receptor gene are hypomethylated in Prader-Willi syndrome and in Prader-Willi syndrome associated psychosis

“…This contradicts the assumption that the observed hypomethylation points to an increased OXTR expression. However, RNA is known to be unstable, which specifically makes it hard to accurately examine expression levels in post mortem tissue [ 42 ]. Furthermore, it is important to keep in mind that hypomethylation at specific single-CpG sites could also lead to decreased OXTR expression depending on the effect that methylation has on the interaction of DNA-binding proteins and the respective DNA at the specific CpG site.…”

Long term storage of miRNA at room and elevated temperatures in a silica sol–gel matrix

“…Recent advances in room temperature storage include commercial products such as Biomatrica (San Diego, CA, USA), GenTegra (IntegenX, Pleasanton, CA, USA) and Imagene (Evry Cedex, France) that employ Anhydrobiosis ("life without water") however are constrained with multi-step rehydration and drying cycles of their ultra-dry bio specimens, prior to downstream processing. [12][13][14][15] The potential for using hybrid functional materials to stabilize biological samples has been recognized by several scientists. 16 Encapsulation in silica sol gels, in particular, has been explored due to the relative simplicity and biocompatibility of the gels, and the ability to control its relative water content and salinity.…”

“…Recent advances in room temperature storage include commercial products such as Biomatrica (San Diego, CA, USA), GenTegra (IntegenX, Pleasanton, CA, USA) and Imagene (Evry Cedex, France) that employ Anhydrobiosis (“life without water”) however are constrained with multi-step rehydration and drying cycles of their ultra-dry bio specimens, prior to downstream processing. 12–15 …”