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Nucleic Acids Res. 1994 Oct 11;22(20):4259-67.

Termination of DNA synthesis by novel 3'-modified-deoxyribonucleoside 5'-triphosphates.

Author information

1
Department of Molecular and Human Genetics, Baylor College of Medicine, Houston, TX 77030.

Abstract

Eight 3'-modified-dNTPs were synthesized and tested in two different DNA template assays for incorporation activity. From this enzymatic screen, two 3'-O-methyl-dNTPs were shown to terminate DNA syntheses mediated by a number of polymerases and may be used as alternative terminators in Sanger sequencing. 3'-O-(2-Nitrobenzyl)-dATP is a UV sensitive nucleotide and was shown to be incorporated by several thermostable DNA polymerases. Base specific termination and efficient photolytic removal of the 3'-protecting group was demonstrated. Following deprotection, DNA synthesis was reinitiated by the incorporation of natural nucleotides into DNA. The identification of this labile terminator and the demonstration of a one cycle stop-start DNA synthesis are initial steps in the development of a novel sequencing strategy.

PMID:
7937154
PMCID:
PMC331938
DOI:
10.1093/nar/22.20.4259
[Indexed for MEDLINE]
Free PMC Article

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