TRANSLATIONAL INHIBITION BY 5'-POLYCYTIDINE TRACTS IN XENOPUS EMBRYOSAND IN-VITRO

Introduction of in vitro transcribed mRNA into Xenopus laevis embryos is a useful technique for analyzing gene function. In order to optimiz e expression of cDNA constructs from in vitro transcribed mRNAs, we ex amined the translational efficiency of reporter genes that simulated c DNAs synthesized by tailing with deoxyguanosine (dG) before second str and cDNA synthesis. When transcribed in vitro, these cDNAs give rise t o RNAs containing a 5'-homopolymeric cytidine (poly(C)) stretch. We ob served that the presence of a 5'-poly(C) tract depressed translation o f a CAT reporter gene at least 100-fold in Xenopus embryos and up to 5 -fold in vitro. This effect was not seen when a 5'-polyadenosine tract was tested. Translational depression was dependent on the phage polym erase used for in vitro transcription: RNAs transcribed by T7 polymera se translated far more poorly than those transcribed by SP6 polymerase . These results have general implications for optimizing expression of cDNA constructs.