A high capacity, reusable oligodeoxythymidine affinity column.
A high capacity, reusable oligodeoxythymidine (oligo(dT)) affinity column suitable for purification of polyadenylated mRNA is described. Oligo(dT) molecules, 30 nucleotides in length, were chemically synthesized on an Applied Biosystems DNA synthesizer. Deprotection of the oligo(dT) molecules was achieved by thiophenol/dioxane/triethylamine treatment. Since no hot ammonia treatment was required, the oligo(dT) molecules remained covalently attached to the support matrix. The resulting column has very high oligo(dT) density and can be used directly for purifying polyadenylated mRNA. Column-purified mRNAs were shown to be intact and functional in Northern blot analysis, in vitro translation, and reverse transcription assays.
Chow
T
T
Biotechnology Research Institute, National Research Council Canada, Montreal, Quebec.
Juby
C
C
Yuen
L
L
D000818:Animals; D015152:Blotting, Northern; D002846:Chromatography, Affinity / Q000295:instrumentation*; D066298:In Vitro Techniques; D009838:Oligodeoxyribonucleotides / Q000138:chemical synthesis; D011061:Poly A / Q000235:genetics / Q000302:isolation & purification*; D014176:Protein Biosynthesis; D012333:RNA, Messenger / Q000235:genetics / Q000302:isolation & purification*; D012194:RNA-Directed DNA Polymerase
D009838:Oligodeoxyribonucleotides; D012333:RNA, Messenger; C027903:oligo (dT); D011061:Poly A; D012194:RNA-Directed DNA Polymerase