Synthesis of RNA from cellulose-bound complementary DNA.
Radioactively labeled RNAs were synthesized from cellulose-bound cDNA templates using Escherichia coli RNA polymerase. Hybridization of this RNA to excess unlabeled cDNA approached 100%, indicating the complementarity of product and template. The average length of the RNA product, as determined by formamide gels, was approximately 40% of the template length. Hybridization of unlabeled globin RNA produced by this technique to labeled globin cDNA indicated the population of RNA sequences represented at least 80% of the template sequences. Approximately 30% of the RNA product by mass contains poly(A) tails as determined by binding to oligo(dT)-cellulose. The template can be reused for several cycles of synthesis with little loss of synthetic capability and therefore, can amplify the amount of mRNA initially used to produce the template.
D002482:Cellulose; D004269:DNA, Bacterial / Q000378:metabolism*; D012321:DNA-Directed RNA Polymerases / Q000378:metabolism*; D004926:Escherichia coli / Q000201:enzymology; D007700:Kinetics; D009693:Nucleic Acid Hybridization; D012319:RNA Polymerase II / Q000378:metabolism*; D012329:RNA, Bacterial / Q000378:metabolism*; D013698:Templates, Genetic
D004269:DNA, Bacterial; D012329:RNA, Bacterial; D002482:Cellulose; D012319:RNA Polymerase II; D012321:DNA-Directed RNA Polymerases