Synthesis of novel poly(dG)-poly(dG)-poly(dC) triplex structure by Klenow exo- fragment of DNA polymerase I

Alexander Kotlyar*, Natalia Borovok, Tatiana Molotsky, Dmitry Klinov, Benjamin Dwir, Eli Kapon

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

12 Scopus citations

Abstract

The extension of the G-strand of long (700 bp) poly(dG)-poly(dC) by the Klenow exo- fragment of DNA polymerase I yields a complete triplex structure of the H-DNA type. High-performance liquid chromatography analysis demonstrates that the length of the G-strand is doubled during the polymerase synthesis. Fluorescence resonance energy transfer analysis shows that the 5′ ends of the G- and the C-strands, labeled with fluorescein and TAMRA, respectively, are positioned close to each other in the product of the synthesis. Atomic force microscopy morphology imaging shows that the synthesized structures lack single-stranded fragments and have approximately the same length as the parent 700 bp poly(dG)-poly(dC). CD spectrum of the polymer has a large negative peak at 278 nm, which is characteristic of the poly(dG)-poly(dG)-poly(dC) triplex. The polymer is resistant to DNase and interacts much more weakly with ethidium bromide as compared with the double-stranded DNA.

Original languageEnglish
Pages (from-to)6515-6521
Number of pages7
JournalNucleic Acids Research
Volume33
Issue number20
DOIs
StatePublished - 2005

Funding

FundersFunder number
European grant for Future & Emerging TechnologiesIST-2001-38951
H2020 Future and Emerging Technologies

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