Accurate in vitro transcription of plant promoters with nuclear extracts prepared from cultured plant cells.
A simple method is presented for the preparation of nuclear extracts from suspension cultures of rice, wheat and tobacco cells. These extracts are shown to be capable of RNA Polymerase II-dependent transcription from two plant promoters in vitro; a 250 bp fragment of a wheat gliadin promoter containing sequences from -167 bp to +83 relative to the in vivo transcriptional initiation site and two fragments of the CaMV 35S promoter, containing sequences from -419 to +17, and from -90 to +17. Using the rice extract, transcription is shown to be extract-dependent, DNA-dependent, alpha-amanatin-sensitive, promoter-dependent, and accurate with respect to initiation site selection on the gliadin promoter and the -90 to +17 35S promoter, but not accurate on the -419 to +17 35S promoter.
Roberts
M W
MW
Institute of Biological Chemistry, Washington State University, Pullman 99163-6340.
Okita
T W
TW
D001483:Base Sequence; D002457:Cell Extracts; D002467:Cell Nucleus; D002469:Cell Separation; D002478:Cells, Cultured; D004247:DNA; D005903:Gliadin / Q000235:genetics; D008969:Molecular Sequence Data; D012275:Oryza; D010942:Plant Viruses / Q000235:genetics; D010944:Plants; D010957:Plasmids; D011401:Promoter Regions, Genetic; D014158:Transcription, Genetic
D002457:Cell Extracts; D004247:DNA; D005903:Gliadin