Journal: Nucleic Acids Research
Article Title: Small circular DNAs for synthesis of the human telomere repeat: varied sizes, structures and telomere-encoding activities
Figure Lengend Snippet: Extension of a human telomeric primer using C-rich DNA nanocircles. A radioactive primer composed of three repeats of the G-rich human telomeric sequence (GGGTTA) 3 was incubated with ( A ) Klenow Fragment of E.coli DNA polymerase I, lacking 3′–5′-exonuclease activity; ( B ) calf thymus polymerase alpha; ( C ) human polymerase beta; and ( D ) HeLa nuclear cell extracts. Reactions further contained 100 nM DNA nanocircle, traces of 5′- 32 P-labeled primer (GGGTTA) 3 , and 1 mM dNTPs, and were incubated at 37°C for 1 h. Lane 1, control reaction lacking DNA nanocircle; lane 2, 36 nt circle; lane 3, 42 nt circle; lane 4, 48 nt circle; lane 5, 54 nt circle; lane 6, 60 nt circle; and M, size marker.
Article Snippet: A total of 8 U of Klenow Fragment lacking 3′-5′-exonuclease activity (New England Biolabs) were used in 50 mM Tris buffer, pH 7.5, 5 mM MgCl2 and 7.5 mM DTT.
Techniques: Sequencing, Incubation, Activity Assay, Labeling, Marker