ENZYMATIC INCORPORATION OF 3’-O-FLUORESCENTLY MODIFIED NUCLEOTIDES FOR SEQUENCING-BY-SYNTHESIS

Abstract

Incorporation of 3’-O-fluorescently modified nucleotides by a DNA polymerase was examined. Among the dye-labeled dTTPs tested, modified nucleotides with Pacific Blue and Bodipy-FL could be completely incorporated. Due to the blocking groups on the 3’-OH, termination of DNA synthesis was observed after extension of a single nucleotide in the primer. Since the linker is chemically cleavable under DNA compatible conditions, those modified nucleotides may be used as reversible terminators in sequencing-by-synthesis (SBS)