1987 Volume 102 Issue 5 Pages 1289-1301
An in vitro splicing system was constructed using portions of chicken δ-crystallin pre-mRNA synthesized in vitro and a HeLa nuclear extract. Analysis of the reaction products revealed that about 25 of the pre-mRNA was precisely spliced at 30°C in 2h under the standard conditions. The other major products of the reaction detected were a 5'-exon fragment and three RNA species showing unusual electrophoretic mobilities on polyacrylamide gels. Structural analyses showed that these three RNAs contain a branch (lariat) structure as seen in the in vitro splicing reactions of human β-globin, adenovirus, and yeast pre-mRNAs. In addition, methylation at the N-7 position of the blocking guanosine of the 5'-terminal cap structure of pre-mRNA has been suggested to play an important role in the splicing reaction.
