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Total tRNA Shipping Requirements
If you have contacted the lab and are preparing total tRNA for shipment and analysis, please follow the protocol that can be downloaded here.
CMCT Derivatization Conditions
We have identified a typo in an earlier paper on CMCT derivatization for pseuduridine detection by mass spectrometry (Durairaj, A.; Limbach, P.A. “Improving CMC Derivatization of Pseudouridine in RNA for Mass Spectrometric Detection”, Anal. Chim. Acta 612 (2008) 173-181. PMID: 18358863). This paper stated in the Results & Discussion that an 8000:1 mol ratio was optimum, although one can confirm that was not the mole ratio described in the Experimental section. We apologize for this error.
Our “best” conditions, which are gauged based on compatibility with mass spectrometry (not primer extension assays), are typically around 100 milligrams of CMCT for every 5 milligrams of tRNA. Under those conditions, we can limit non-specific derivatization yet still adequately derivatize pseudouridine for detection by MS.
Prof. Addepalli in the laboratory has been examining a number of other reaction conditions for RNAs beyond tRNAs, including more recent efforts identifying any off-target effects of CMCT. Although his work remains unpublished, you can contact him for more details by email.