The biogenesis of mature, functional mRNA in eukaryotes often requires the precise excision of noncoding, intervening sequences (i.e., introns) which are present in the precursor mRNA (pre-mRNA) molecule and subsequent ligation of the flanking, functional sequences (i.e., exons). Pre-mRNA splicing is thus a critical step in the long chain of events required for the expression of a large number of eukaryotic genes. In a number of instances, the splicing process itself is regulated such that alternatively spliced mRNAs which encode novel proteins are generated. The development of cell free splicing systems, as well as the availability of yeast genetic techniques, has greatly aided investigations of the mechanism whereby nuclear pre-mRNA is spliced. Nuclear pre-mRNA splicing involves two transesterification steps (Fig. 1; see Moore et al. 1993 for detailed review). In the first step, the 2′ hydroxyl group of an adenosine near the 3′ end of the intron (i.e., the branch point) engages in a nucleophilic attack on the phosphodiester bond at the 5′ splice site, thereby forming the intermediates of the splicing reaction, the cleaved exon 1, and the intron-exon 2 lariat. In the second step, the 3′ hydroxyl group of exon 1 carries out a nucleophilic attack on the phosphodiester bond at the 3′ splice site. This results in the ligation of the two exons and release of the intron, which is still in the form of a lariat.