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

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.

Polymerases:

Topics:

Exonuclease Activity

Status:

new topics/pols set partial results complete validated

Results:

Polymerase Reference Property Result Context
Klenow (exo-) Kotlyar AB2005 3-5' Exonuclease (proofreading) No
Klenow (exo-) Kotlyar AB2005 5-3' Exonuclease Unspecified
Klenow fragment Kotlyar AB2005 3-5' Exonuclease (proofreading) Yes
Klenow fragment Kotlyar AB2005 5-3' Exonuclease Unspecified

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