The expression of a nopaline synthase ? human growth hormone chimaeric gene in transformed tobacco and sunflower callus tissue

Abstract

To study whether mammalian RNA processing signals function in plants, we have constructed a chimaeric gene in which the complete human growth hormone (hGH) gene is flanked by DNA fragments containing the promoter and polyadenylation site of the nopaline synthase gene. The hGH gene used contains four introns and an additional 440 bp downstream from the hGH poly(A) addition site. The transcription of this chimaeric gene was studied following its introduction into sunflower and tobacco cells using a Ti plasmid vector. Analysis of poly(A)(+) RNA isolated from the transformed tumor tissue demonstrated the following: (1) a single polyadenylated transcript, 2700 bp in length, was transcribed from the chimaeric gene; (2) the transcription was initiated at the published start site of the nopaline synthase gene; (3) the hGH polyadenylation site was not used for processing of the 3' end; only the poly(A) addition site of the nopaline synthase gene was recognized, (4) no splicing of the hGH introns could be detected. We also demonstrate that the hGH pre-mRNA isolated from plant cells can be spliced in a HeLa cell nuclear extract, indicating that the hGH pre-mRNA was functional. These results show that processing signals of the hGH pre-mRNA are not recognized in these plant cells.