Spacers Applications

Spacers Applications

Some common usages of Spacer phosphoramidites are to position various tags/labels at a desired length from an oligonucleotide, to immobilize oligonucleotides to a solid phase (like a microsphere or microarray slide), or to form non-nucleoside folds and hairpin loops within an oligonucleotide (1-3). Spacers can be placed anywhere within an oligo, and multiple incorporations of a Spacer can be performed to effectively generate spacers of virtually any desired length. For example, six successive incorporations of dSpacer were used to create the optimal separation for FRET between the donor and acceptor fluorophores in energy transfer fluorescent sequencing primers (4).

In addition to being used as a spacer arm, the Spacer C3 modification also can be placed at the 3’-end of an oligonucleotide to effectively block that end from enzymatic reactions (e.g., extension by PCR) (5). dSpacer and rSpacer also can be used to mimic abasic sites in oligonucleotides slated for use in DNA damage/repair studies (6). Photocleavable linkers and spacers can be used to introduce photocleavable tags/labels onto the ends of an oligo, or link two separate oligonucleotides through a short, photocleavable spacer arm for use in photo-triggered hybridization applications (7).

References

(1) Li, H., McGall, G. Photoactivatable Silanes: Synthesis and Uses in Biopolymer Array Fabrication on Glass Substrates. In Frontiers in Biochip Technology. X, W-L., Cheng, J. (Ed.) Springer Science+Business Media, Inc. (2006), pp. 176-190.
(2) Salunkhe, M., Wu, T.F., Letsinger, R.L. Control of folding and binding of oligonucleotides by use of non-nucleotide linker. J. Am. Chem. Soc. (1992), 114: 8768-8772.
(3) Durand, M., Chevrie, K., Chassignol, M., Thuong, N.T., Maurizot, J. Circular dichroism studies of an oligodeoxyribonucleotide containing a hairpin loop made of a hexaethylene glycol chain : conformation and stability. Nucleic Acids Res. (1990), 18: 6353-6359.
(4) Ju, J., Glazer, A.N., Mathies, R.A. Cassette labeling for facile construction of energy transfer fluorescent primers. Nucleic Acids Res. (1996), 24: 1144-1148.
(5) Zhou, L., Myers, A.N., Vandersteen, J.G., Wang, L., Wittwer, C.T. Closed-Tube Genotyping with Unlabeled Oligonucleotide Probes and a Saturating DNA Dye. Clin. Chem. (2004), 50: 1328-1335.
(6) Takeshita, M., Chang, C.N., Johnson, F., Will, S., Grollman, A.P. Oligodeoxynucleotides containing synthetic abasic sites. Model substrates for DNA polymerases and apurinic/apyrimidinic endonucleases. J. Biol. Chem. (1987), 262: 10171-10179.
(7) Ordoukhanian, P., Taylor, J-S. Design and synthesis of a versatile photocleavable DNA building block. Application to phototriggered hybridization. J. Am. Chem. Soc. (1995), 117: 9570-9571.