RNA sequencing using fluorescent-labeled dideoxynucleotides and automated fluorescence detection.
Although dideoxy terminated sequencing of RNA, using reverse transcriptase and oligodeoxynucleotide primers, is now a well established method, the accuracy is limited by sequence ambiguities due to unspecific chain termination events. A protocol is described which circumvents these ambiguities by using fluorescence labels tagged to dideoxynucleotides. Only chain terminations caused by dideoxynucleotides were detected while premature terminated cDNA's remain undetectable. In addition, the remaining multiple signals at nucleotide positions can be assigned to sequence heterogeneities within the RNA sequence to be determined.
Max-Planck Institut für Biophysikalische Chemie, Abteilung Biochemische Kinetik, Göttingen, FRG.
G J
GJ
Bauer
D001483:Base Sequence; D004279:DNA, Viral / Q000235:genetics; D004854:Herpesvirus 4, Human / Q000235:genetics; D007202:Indicators and Reagents; D008969:Molecular Sequence Data; D009838:Oligodeoxyribonucleotides; D015345:Oligonucleotide Probes; D012313:RNA / Q000235:genetics*; D012335:RNA, Ribosomal / Q000235:genetics; D012367:RNA, Viral / Q000235:genetics; D012194:RNA-Directed DNA Polymerase
D004279:DNA, Viral; D007202:Indicators and Reagents; D009838:Oligodeoxyribonucleotides; D015345:Oligonucleotide Probes; D012335:RNA, Ribosomal; D012367:RNA, Viral; D012313:RNA; D012194:RNA-Directed DNA Polymerase