An oligonucleotide hybridization approach to DNA sequencing.
We have proposed a DNA sequencing method based on hybridization of a DNA fragment to be sequenced with the complete set of fixed-length oligonucleotides (e.g., 4(8) = 65,536 possible 8-mers) immobilized individually as dots of a 2-D matrix [(1989) Dokl. Akad. Nauk SSSR 303, 1508-1511]. It was shown that the list of hybridizing octanucleotides is sufficient for the computer-assisted reconstruction of the structures for 80% of random-sequence fragments up to 200 bases long, based on the analysis of the octanucleotide overlapping. Here a refinement of the method and some experimental data are presented. We have performed hybridizations with oligonucleotides immobilized on a glass plate, and obtained their dissociation curves down to heptanucleotides. Other approaches, e.g., an additional hybridization of short oligonucleotides which continuously extend duplexes formed between the fragment and immobilized oligonucleotides, should considerably increase either the probability of unambiguous reconstruction, or the length of reconstructed sequences, or decrease the size of immobilized oligonucleotides.
Khrapko
K R
KR
V.A. Engelhardt Institute of Molecular Biology, Academy of Sciences of the USSR, Moscow.
Lysov YuP
Khorlyn
A A
AA
Shick
V V
VV
Florentiev
V L
VL
Mirzabekov
A D
AD
D001483:Base Sequence; D003201:Computers; D004247:DNA / Q000032:analysis*; D005796:Genes; D005807:Genes, Overlapping; D005821:Genetic Techniques; D015336:Molecular Probe Techniques; D009693:Nucleic Acid Hybridization; D009841:Oligonucleotides / Q000032:analysis*; D010446:Peptide Fragments / Q000032:analysis
D009841:Oligonucleotides; D010446:Peptide Fragments; D004247:DNA