Enzymatic Synthesis and Characterization of Unmodified Ribozymes and Substrates

This chapter provides the basic procedures for RNA handling, transcription reactions, and transcript purification (1 ,2 ). For further analysis, the RNAs can be end-labeled. Simple procedures generate a sequence ladder to confirm colinearity of template and transcript (3 ,4 ). This is not a “mere luxury,” since examples with significant template slippage have been reported (5 ). If a small labeled cleavage product is produced, also the exact cleavage sites can be determined (6 ).