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An affinity electrophoretic method has been developed to study the state of terminal phosphorylation of RNAs and the presence of the hypermodified base Q in tRNA. It is based on the copolymerization of acryloylaminophenylboronic acid into standard polyacrylamide gels and the interaction of this derivative with free cis-diol groups present in the RNA. In the case of terminal phosphorylation, free ribose groups are present either as such, or may be introduced by enzymatic reactions specific for a particular phosphorylation pattern (e.g. using T4 RNA ligase or guanylyltransferase). Additionally, tRNA species containing the Q base may be resolved from Q-lacking tRNAs by boronate affinity electrophoresis. The introduction of a non-destructive, one-step electrophoretic procedure not only offers an alternative to classical analytical methods, but also provides a means of isolating such populations of RNAs for which other methods are unavailable or are less convenient.

nucleic acids research

Affinity electrophoresis for monitoring terminal phosphorylation and the presence of queuosine in RNA. Application of polyacrylamide containing a covalently bound boronic acid