Transcription of cloned tRNA gene fragments and subfragments injected into the oocyte nucleus of Xenopus laevis

Abstract

Cloned 3.18 kilobase fragments of Xenopus laevis DNA containing genes for tRNAMet1 and for at least one other 4S RNA species are transcribed rapidly after their injection into the nucleus of X. laevis oocytes. The newly synthesized RNA can be resolved by gel electrophoresis into a few predominant 4S RNA species and a series of slower migrating components. One of the 4S RNA species appears to be identical, by fingerprint analysis, to the tRNAMet1 isolated by hybridization of somatic cell RNA to this cloned tRNA gene fragment (tDNA). Thus, the tRNAMet1 produced after injection can be both fully processed and modified. Its rate of synthesis is estimated to be about 6 x 10(9) molecules/hr in each oocyte injected with 2 ng of tDNA. When the tDNA fragment is cleaved into two halves with restriction endonuclease Sst I, each injected half gives rise to a subset of the RNAs produced after injection of the intact fragment. This experiment thus suggests the presence of at least two transcriptional units on this cloned tDNA. This simple way of biologically testing defined restriction fragments may be of value for analyzing the functional organization of other cloned eukaryotic DNA units.