title: Long, processive enzymatic DNA synthesis using 100% dye-labeled terminal phosphate-linked nucleotides.
abstract: We demonstrate the efficient synthesis of DNA with complete replacement of the four deoxyribonucleoside triphosphate (dNTP) substrates with nucleotides carrying fluorescent labels. A different, spectrally separable fluorescent dye suitable for single molecule fluorescence detection was conjugated to each of the four dNTPs via linkage to the terminal phosphate. Using these modified nucleotides, DNA synthesis by phi 29 DNA polymerase was observed to be processive for products thousands of bases in length, with labeled nucleotide affinities and DNA polymerization rates approaching unmodified dNTP levels. Results presented here show the compatibility of these nucleotides for single-molecule, real-time DNA sequencing applications.
authors: Pacific Biosciences Inc., Menlo Park, California 94025, USA.
authors: Jonas
authors: J
authors: Korlach
authors: Arek
authors: A
authors: Bibillo
authors: Jeffrey
authors: J
authors: Wegener
authors: Paul
authors: P
authors: Peluso
authors: Thang T
authors: TT
authors: Pham
authors: Insil
authors: I
authors: Park
authors: Sonya
authors: S
authors: Clark
authors: Geoff A
authors: GA
authors: Otto
authors: Stephen W
authors: SW
authors: Turner
mesh_terms: D004247:DNA / Q000138:chemical synthesis*; D054306:Dideoxynucleotides / Q000737:chemistry; D005456:Fluorescent Dyes / Q000737:chemistry*; D007700:Kinetics; D015394:Molecular Structure; D009711:Nucleotides / Q000737:chemistry*
chemical_list: D054306:Dideoxynucleotides; D005456:Fluorescent Dyes; D009711:Nucleotides; D004247:DNA