12 Water is involved, for instance, by providing hydroxyl or hydronium ions or by allowing proton transfer. 11 RNase A and some ribozymes share this mechanism. For reviews, see Emilsson et al 10 and Oivanen et al. A large variety of agents such as specific acids and bases as well as Brønsted acids and base acting as catalysts can be involved.
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However, the main degradative event is the spontaneous cleavage of the phosphodiester linkage through transesterification resulting from a nucleophilic attack of the phosphorus atom by the neighboring 2′OH.
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9 Degradation can also occur through the activity of some metallic complexes catalyzing the hydrolytic cleavage of the phosphodiester bond or by contaminating nucleases. 7, 8 Oxidation could also result from attacks by ozone, an atmospheric pollutant that rapidly reacts with RNA either in solution or in the solid state. 6 Outside the cell, they can be generated by mechanisms generally involving metallic ions. In vivo, they are produced by respiration. First, it is very sensitive to oxidation by reactive oxygen species. However, this molecule can be affected by multiple degradation reactions. For reviews, see Baginsky et al, 3 Malone et al, 4 Wang et al 5.įor most of these uses, integrity of RNA is required and must be maintained during storage. In addition, specific RNA species are used as clinical standards 1, 2 or as agents for controlling gene expression (siRNA, ribozymes). For instance, RNA populations (whole or subsets) are quantitatively analyzed for studying cellular functions, elucidating normal or pathological mechanisms or seeking molecular signatures for diagnostic purposes (microarrays, RNAseq). RNA is a tool used in many fields, from molecular and cellular biology to medicine and nanotechnology. At last, our data are consistent with a sequence-independent degradation rate of RNA in the solid state.
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No significant change in the C q values was observed over a simulated period of several decades. In addition, we showed that stored RNA is compatible for further analyses, such as reverse transcription-quantitative PCR. In these conditions, it is expected that an RNA molecule will be subjected to 0.7–1.3 cut every 1000 nucleotides per century. The degradation rate dependence in temperature fitted an Arrhenius model, with an activation energy of 28.5 kcal/mol and an extrapolated room temperature degradation rate of 3.2 10 −13/nt/s (95% confidence interval: 2.3–4.2/nt/s). Through the evaluation of RNA integrity over time at room temperature or 90 ☌, we identified atmospheric humidity as a major deleterious factor. These air- and water-tight capsules isolate RNA from the atmosphere and maintain an anhydrous and anoxic environment. Here, we report a new room temperature technology that consists in drying RNA samples in the presence of a stabilizer in stainless steel minicapsules.
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Even though freezing is currently the storage method of choice, the increasing number of samples to be stored and the costly use of a cold chain have highlighted the need for room temperature preservation methods. For most of these uses, the integrity of RNA is required and must be maintained during storage.