rPhosphoramidites and dPhosphoramidites

Structural studies on large DNA and RNA molecules by nuclear magnetic resonance (NMR) require labeling of these molecules with stable isotopes. Silantes offers a wide range of stable isotope-labeled ribo- and deoxyribo-phosphoramidites for the solid phase synthesis of oligonucleotides.

Available in many Uniform and Site-Specific Labeling Patterns

The phosphoramidites are either uniformly, or site-specifically labeled with ²H, ¹³C and/or ¹⁵N at individual positions within the nucleobase or ribose. The individual labeling patterns can be seen in the product list.

Selected products in this field (complete product range at bottom of page):

Uniformly stable isotope labeled phosphoramidites

Our uniformly stable isotope-labeled phosphoramidites are synthesised using a combination of biotechnological and chemical synthesis steps. In the first step, we obtain nucleosides from ²H, ¹³C and/or ¹⁵N-labeled bacterial biomass. In a second step, these nucleosides are chemically protected to obtain the phosphoramidites.

In vivo enrichment technology of biomass with stable isotopes results in:

• efficient incorporation of the stable isotopes and thus cost-effectively labeled amidites.
• high and homogeneous isotope labeling (> 98 atom %) of the nucleotides by using a closed system.

Site-specific labeled phosphoramidites

We also offer site-specific labeled phosphoramidites. The site-specific labeling patterns are chosen to create independent spin systems either intramolecularly within the nucleotide, or intramolecularly to other molecules, providing high quality NMR information.

High quality at reasonable prices

The isotope enrichment of the amidites is >98 % and the chemical purity >95 %. This is confirmed by mass spectrometry analysis and ³¹P-NMR. The figure below shows that phosphonate (at 150ppm), a well-known contaminant in the synthesis of deoxy-phosphoramidites, is absent.

References

Use cases of the Silantes NTPs in scientific publications:

• Mieczkowski, M., Steinmetzger, C., Bessi, I., Lenz, A., Schmiedel, A., Holzapfel, M., Lambert, C., Pena, V., & Höbartner, C. (2021). Large Stokes shift fluorescence activation in an RNA aptamer by intermolecular proton transfer to guanine. Nature Communications, 12(1). https://doi.org/10.1038/s41467-021-23932-0
• Musheev, M. U., Schomacher, L., Basu, A., Han, D., Krebs, L., Scholz, C., & Niehrs, C. (2022). Mammalian N1-adenosine PARylation is a reversible DNA modification. Nature Communications, 13(1). https://doi.org/10.1038/s41467-022-33731-w
• Xu, Y., McSally, J., Andricioaei, I., & Al-Hashimi, H. M. (2018). Modulation of Hoogsteen dynamics on DNA recognition. Nature Communications, 9(1). https://doi.org/10.1038/s41467-018-03516-1
• Li, M., Wang, Y., Wei, X., Cai, W., Wu, J., Zhu, M., Wang, Y., Liu, Y., Xiong, J., Qu, Q., Chen, Y., Tian, X., Yao, L., Xie, R., Li, X., Chen, S., Huang, X., Zhang, C., Xie, C., . . . Lin, S. (2024). AMPK targets PDZD8 to trigger carbon source shift from glucose to glutamine. Cell Research. https://doi.org/10.1038/s41422-024-00985-6
• Cromsigt, J., Schleucher, J., Gustafsson, T., Kihlberg, J., & Wijmenga, S. (2002). Preparation of partially 2H/13C-labelled RNA for NMR studies. Stereo-specific deuteration of the H5’’ in nucleotides. Nucleic Acids Research, 30(7), 1639–1645. https://doi.org/10.1093/nar/30.7.1639
• Rangadurai, A., Szymanski, E. S., Kimsey, I., Shi, H., & Al-Hashimi, H. M. (2020). Probing conformational transitions towards mutagenic Watson–Crick-like G·T mismatches using off-resonance sugar carbon R1ρ relaxation dispersion. Journal of Biomolecular NMR, 74(8–9), 457–471. https://doi.org/10.1007/s10858-020-00337-7
• Noeske, J., Richter, C., Grundl, M. A., Nasiri, H. R., Schwalbe, H., & Wöhnert, J. (2005). An intermolecular base triple as the basis of ligand specificity and affinity in the guanine- and adenine-sensing riboswitch RNAs. Proceedings of the National Academy of Sciences, 102(5), 1372–1377. https://doi.org/10.1073/pnas.0406347102
• Ohira, T., Minowa, K., Sugiyama, K., Yamashita, S., Sakaguchi, Y., Miyauchi, K., Noguchi, R., Kaneko, A., Orita, I., Fukui, T., Tomita, K., & Suzuki, T. (2022). Reversible RNA phosphorylation stabilizes tRNA for cellular thermotolerance. Nature, 605(7909), 372–379. https://doi.org/10.1038/s41586-022-04677-2
• Vögele, J., Duchardt-Ferner, E., Bains, J. K., Knezic, B., Wacker, A., Sich, C., Weigand, J. E., Šponer, J., Schwalbe, H., Krepl, M., & Wöhnert, J. (2024). Structure of an internal loop motif with three consecutive U•U mismatches from stem–loop 1 in the 3′-UTR of the SARS-CoV-2 genomic RNA. Nucleic Acids Research, 52(11), 6687–6706. https://doi.org/10.1093/nar/gkae349
• Broft, P., Rosenkranz, R. R., Schleiff, E., Hengesbach, M., & Schwalbe, H. (2022). Structural analysis of temperature-dependent alternative splicing of HsfA2 pre-mRNA from tomato plants. RNA Biology, 19(1), 266–278. https://doi.org/10.1080/15476286.2021.2024034

Use cases of the Silantes phosphoramidites in scientific publications:

• Becette, O., Olenginski, L. T., & Dayie, T. K. (2019). Solid-Phase chemical synthesis of stable Isotope-Labeled RNA to aid structure and dynamics studies by NMR spectroscopy. Molecules, 24(19), 3476. https://doi.org/10.3390/molecules24193476
• Štih, V., Amenitsch, H., Plavec, J., & Podbevšek, P. (2023). Spatial arrangement of functional domains in OxyS stress response sRNA. RNA, 29(10), 1520–1534. https://doi.org/10.1261/rna.079618.123

Use cases of the Silantes oligonucleotide synthesis service in scientific publications:

• Belfetmi, A., Zargarian, L., Tisné, C., Sleiman, D., Morellet, N., Lescop, E., Maskri, O., René, B., Mély, Y., Fosse, P., & Mauffret, O. (2016). Insights into the mechanisms of RNA secondary structure destabilization by the HIV-1 nucleocapsid protein. RNA, 22(4), 506–517. https://doi.org/10.1261/rna.054445.115
• Borggräfe, J., Victor, J., Rosenbach, H., Viegas, A., Gertzen, C. G. W., Wuebben, C., … Etzkorn, M. (2021). Time-resolved structural analysis of an RNA-cleaving DNA catalyst. Nature, 601(7891), 144–149. https://doi.org/10.1038/s41586-021-04225-4
• Chernatynskaya, A. V., Deleeuw, L., Trent, J. O., Brown, T., & Lane, A. N. (2009). Structural analysis of the DNA target site and its interaction with Mbp1. Organic & Biomolecular Chemistry, 7(23), 4981. https://doi.org/10.1039/b912309a
• Van Melckebeke, H., Devany, M., Di Primo, C., Beaurain, F., Toulmé, J., Bryce, D. L., & Boisbouvier, J. (2008). Liquid-crystal NMR structure of HIV TAR RNA bound to its SELEX RNA aptamer reveals the origins of the high stability of the complex. Proceedings of the National Academy of Sciences, 105(27), 9210–9215. https://doi.org/10.1073/pnas.0712121105

Use cases of the Silantes 14-mer RNA Standard in scientific publications:

• Duchardt, E., & Schwalbe, H. (2005). Residue Specific Ribose and Nucleobase Dynamics of the cUUCGg RNA Tetraloop Motif by MNMR 13C Relaxation. Journal of Biomolecular NMR, 32(4), 295–308. https://doi.org/10.1007/s10858-005-0659-x
• Hartlmüller, C., Günther, J. C., Wolter, A. C., Wöhnert, J., Sattler, M., & Madl, T. (2017). RNA structure refinement using NMR solvent accessibility data. Scientific Reports, 7(1). https://doi.org/10.1038/s41598-017-05821-z
• Nozinovic, S., Fürtig, B., Jonker, H. R. A., Richter, C., & Schwalbe, H. (2009). High-resolution NMR structure of an RNA model system: the 14-mer cUUCGg tetraloop hairpin RNA. Nucleic Acids Research, 38(2), 683–694. https://doi.org/10.1093/nar/gkp956
• Richter, C., Kovacs, H., Buck, J., Wacker, A., Fürtig, B., Bermel, W., & Schwalbe, H. (2010). 13C-direct detected NMR experiments for the sequential J-based resonance assignment of RNA oligonucleotides. Journal of Biomolecular NMR, 47(4), 259–269. https://doi.org/10.1007/s10858-010-9429-5
• Ferner, J., Villa, A., Duchardt, E., Widjajakusuma, E., Wöhnert, J., Stock, G., & Schwalbe, H. (2008). NMR and MD studies of the temperature-dependent dynamics of RNA YNMG-tetraloops. Nucleic Acids Research, 36(6), 1928–1940. https://doi.org/10.1093/nar/gkm1183

Relevant blog articles:

• What Are Stable-Isotope Labeled Nucleic Acids?
• Synthesizing Stable Isotope-Labeled Nucleic Acids
• The Advantages of Using Stable Isotope-Labeled Nucleic Acids
• Applications of Stable Isotope-Labeled Molecules: Exploring the Power of Isotopic Tracers
• Custom RNA & DNA Synthesis Services : Tailored Solutions for Your Nucleic Acid Needs

Relevant webinars:

• Easy 13C, 15N-Labeling of DNA through isothermal amplification: Applications to G-quadruplex, aptamer, and DNAzyme