Synthesis and purification of large amounts of RNA oligonucleotides.
Biophysical studies of RNA oligonucleotides require milligram amounts of RNA of specific length and sequence. Transcription from synthetic DNA templates using T7 RNA polymerase is a convenient method for synthesis of RNA oligonucleotides ranging in size from 9 to about 45 nucleotides. Here we present methods that make the large-scale synthesis of RNA oligonucleotides practical. This paper describes a rapid method for isolating T7 RNA polymerase free from RNases for use in transcription reactions. Protocols are also described for purification of the desired RNA oligonucleotide from the other products of transcription.
Wyatt
J R
JR
University of California, Berkeley.
Chastain
M
M
Puglisi
J D
JD
D001483:Base Sequence; D002021:Buffers; D002845:Chromatography; D012321:DNA-Directed RNA Polymerases / Q000302:isolation & purification* / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics / Q000254:growth & development; D008969:Molecular Sequence Data; D009838:Oligodeoxyribonucleotides; D009843:Oligoribonucleotides / Q000138:chemical synthesis / Q000302:isolation & purification*; D013604:T-Phages / Q000201:enzymology / Q000235:genetics; D014158:Transcription, Genetic
D002021:Buffers; D009838:Oligodeoxyribonucleotides; D009843:Oligoribonucleotides; D012321:DNA-Directed RNA Polymerases