ogy,10 nanotechnology applications such as DNA comput-
ing,11 DNA nanostructures using self-assembled branched
units,12 DNA sensors,13 and nanoelectronic devices.14 The
past 10 years have seen remarkable success in the construc-
tion of DNA nanoarchitectures. For example, one- and two-
dimensional DNA lattices15 and dendrimerlike DNA16 have
been constructed from a rich set of branched DNA. Various
autonomous DNA walker devices based on DNA cleavage
and the ligation of branched DNA were explored experi-
mentally. Various template-directed enzymes and the intro-
duction of a branch structure via RCU have also been
explored theoretically (Scheme 3). Figure 6 shows the results
Scheme 3
Figure 6. Capillary gel electrophoresis of photoreaction of ODN
1 (30 µM) and ODN 7 (30 µM) in the presence of template ODN
3 (33 µM) (a) before photoirradiation, (b) after irradiation at 366
nm for 2 h at 0 °C (30% yield), and (c) after irradiation at 366 nm
for 6 h at 0 °C (82% yield).
of capillary gel electrophoresis of the photoirradiated mixture
of 5′-d(AAAAAATGCGTG)-3′ (ODN 7) and ODN 1 in the
presence of template ODN 3, with the clean and efficient
formation of the expected ligated 18-mer ODN 9 and the
disappearance of ODN 7 and ODN 1. MALDI-TOF MS
indicated that ODN 9, purified by HPLC, was the ligated
product of ODN 7 and ODN 1.17 Enzymatic digestion of
isolated ODN 9 showed the formation of dA, dG, dT, and
dC in a ratio of 6:5:3:2, together with a d(RCU-T) photo-
adduct. We observed no formation of ODN 9 after photoir-
radiation of ODN 7 and ODN 1 in the absence of template
ODN 3. These results clearly indicated that ODN 9 was a
branched ODN formed by template-directed cross-linking
between the thymine of ODN 7 and the RCU of ODN 1.
In conclusion, we have demonstrated that an ODN
containing RCU at the 3′ end can be employed in the
photoligation of DNA by irradiation at 366 nm in the
presence of a template DNA, with no side reaction. This
method has the same photosensitivity as the usual method
using âCU. Using this novel photoligation method, we have
demonstrated a convenient and versatile method of generating
branched oligonucleotides, which should be particularly
useful in DNA nanotechnology.
(10) (a) Urdea, M. S. Bio/Technology 1994, 12, 926-928. (b) Colling,
M. L.; Fine, E.; Zayati, C.; Horn, T.; Ahle, D.; Detmer, L. P.; Shen,;
Kolberg, J.; Bushnell, S.; Urdea, M. S.; Ho, D. D. Nucleic Acids Res. 1997,
25, 2979-2984.
(11) Aldyen, P.; Jonoska, N.; Seeman, N. C. J. Am. Chem. Soc. 2004,
126, 6648-6658.
(12) Scheffler, M.; Dorenbeck, A.; Jordan, S.; Wustefeld, G.; Kiedrowski,
G. Angew. Chem., Int. Ed. 1999, 38, 3312-3315.
(13) Nakamura, F.; Ito, E.; Sakou, Y.; Ueno, N.; Gatuna, I. N.; Ohuchi,
F. S; Hara, M. Nano Lett. 2003, 3, 1083-1086.
(14) Becerril, H. A.; Stoltenberg, R. M.; Wheeler, D. R.; Davis, R. C.;
Hard, J. N.; Woolley, A. T. J. Am. Chem. Soc. 2005, 127, 2828-2829.
(15) Winfree, E.; Liu, F.; Wenzler, L. A.; Seeman, N. C. Nature 1998,
394, 539-544.
(16) (a) Shchepinov, M. S.; Udalova, I. A.; Bridgman, A. J.; Southern,
E. M. Nucleic Acid. Res. 1997, 25, 4447-4454. (b) Shi, J.; Bergstrom, D.
E. Angew. Chem., Int. Ed. Engl. 1997, 36, 111-113.
(17) MALDI-TOF MS: calculated for ODN 9 (C180H223N72O106P17) [M
+ H]+ 5618.7373, found 5618.7555.
Supporting Information Available: Experimental pro-
cedures for the synthesis, purification, and analysis of ODNs
2-7, oligonucleotide digestion, and melting studies. This
material is available free of charge via the Internet at
OL050709G
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Org. Lett., Vol. 7, No. 14, 2005