Heterobifunctional modification of DNA for conjugation to solid surfaces.
Many biosensors, DNA arrays, and next-generation DNA sequencing technologies need common methods for end modification of random DNA sequences generated from a sample of DNA. Surface immobilization of chemically modified DNA is often the first step in creating appropriate sensing platforms. We describe a simple technique for efficient heterobifunctional modification of arbitrary double-stranded DNA fragments with chosen chemical groups. The modification requires the use of short (10-20 base pairs) synthetic adaptors having desired terminal functional groups and installs known sequences, which can be used for hybridization of primers in the sequencing-by-synthesis approaches. The method, based on ligation under optimized conditions, is selective and provides high yields of the target heterobifunctional DNA product. An additional two-step procedure can be applied to select further for the desired bifunctionalized product using PCR amplification with a chemically modified primer. Both functional groups in the modified DNA are chemically active and can be used in surface immobilization of the DNA strands to create the surface of a biosensor or sequencing chip.
Lim
Hana I
HI
Chemistry Department, Lehigh University, 6 E. Packer Ave., Bethlehem, PA 18015, USA.
Oliver
Piercen M
PM
Marzillier
Jutta
J
Vezenov
Dmitri V
DV
D001483:Base Sequence; D015374:Biosensing Techniques / Q000295:instrumentation*; D004247:DNA / Q000138:chemical synthesis / Q000737:chemistry* / Q000235:genetics; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction; D017422:Sequence Analysis, DNA / Q000295:instrumentation*
D004247:DNA