[DNA sequencing by hybridization with oligonucleotides immobilized in a gel. Chemical ligation as a method of expanding the prospects for the method].
Chemical ligation was used to obtain a series of modified DNA duplexes containing a substituted pyrophosphate internucleotide bound in a given site of the sugar-phosphate backbone. The efficiency of chemical ligation was shown to depend on the structure of the synthesis center for the modified internucleotide bond, thermosiability of the initial DNA complexes, as well as the nature of the non-nucleotide substituent. Maximal yields (up to 77%) were obtained with DNA complexes of regular structure using for condensation the H-butylamide oligonucleotide derivatives on the terminal phosphate group. Conditions were selected for rapid quantitative cleavage of the substituted pyrophosphate bond in aqueous medium (pH 8.0). It was established that modified oligonucleotides which have a phosphoester bond between the phosphate group and the non-nucleotide substituent are more labile than the corresponding compounds with a phosphoamide bond. The results obtained may be useful for DNA sequencing by hybridization with a set of oligonucleotides. This will allow one to achieve unambiguous recognition of nucleotide sequences containing DNA with repeats and thus extend considerably the sphere of application of the method.
Kuznetsova
S A
SA
Kanevskiĭ
I E
IE
Florent'ev
V A
VA
Mirzabekov
A D
AD
Shabarova
Z A
ZA
D001483:Base Sequence; D004247:DNA / Q000737:chemistry*; D006868:Hydrolysis; D008969:Molecular Sequence Data; D009693:Nucleic Acid Hybridization; D009838:Oligodeoxyribonucleotides / Q000737:chemistry*; D017422:Sequence Analysis, DNA
D009838:Oligodeoxyribonucleotides; D004247:DNA
Sekvenirovanie DNK gibridizatsieĭ s oligonukleotidami, immobilizirovannymi v gele. Khimicheskoe ligirovanie kak sposob rasshireniia vozmozhnosteĭ metoda.