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When using phiX174 RFI DNA as a template, invitro, E. coli RNA polymerase synthesizes four major purine triphosphate-containing 5' end sequences. RNase A digests of alpha(32)P labeled RNA were further digested with spleen exonuclease to remove the bulk of the oligonucleotides with 5' hydroxyls and then chromatographed on DEAE cellulose to resolve the remaining 5' terminal oligonucleotides. By application of standard separation and sequence techniques, the major 5' end sequences were shown to be: pppApUp(Cp), pppApApApUp(Cp), pppApApApApUp(Cp), and pppGpApUp(Gp).

Nucleotide sequences of the 5′ termini of øX174 mRNAs synthesized in vitro