The Ca. LSU intron flanking a 129 bp exon upstream and a 100 bp exon downstream was inserted into the lacZ gene on pRS426 to transform E. coli. Northern blot analysis and RT-PCR showed that splicing of Ca. LSU in E.coli is efficient upon inducible expression of the precursor RNA. In contrast, co-transcriptional self-splicing of the intron in vitro is much less active. Therefore, this E. coli splicing system can be used as a better model to investigate the effect of the ribozyme inhibitors on Ca. LSU splicing in living cell. We examined the effects of neomycin sulfate and pentamidine on Ca. LSU splicing in E. coli, and found that these drugs does-dependently inhibit the intron splicing.However, heomycin is more potent than pentamidine in this action.