RNA Oligo Modifications Design and Protocols

RNA Oligo Modifications Design / Protocol

RNA and 2'-O-methyl RNA oligonucleotide synthesis is performed at Gene Link using the b -cyanoethyl chemistry and state of the art synthesizers. These involve proprietary software protocols with long coupling times and specialized cycles to obtain ultra clean oligos.
RNA oligos are susceptible to degradation to the same extent as native RNA extracted from various sources. An attractive alternate to prevent degradation from nucleases is the use of 2'F and 2'-O- methyl RNA bases, when specific 2'OH is not required. The 2'-O- methyl oligonucleotides confer considerable nuclease resistance and are similar in hydrogen bonding properties to RNA/RNA than the lower RNA/DNA binding property. The coupling efficiency of 2'-O- methyl phosphoramidite is also higher than the RNA monomers resulting in higher yield of full length oligos.
Gene Link also offers custom synthesis of RNA and DNA chimeric oligos with investigator specified ribo or deoxy bases or 2'-O-methyl bases. The chimeric oligos can also be synthesized with the regular phoshodiester bonds or substituted with phosphorothioate linkages. The combination of 2'-O- methyl RNA bases with phosphorothioate internucleotide linkages imparts these oligos greater nuclease resistance which is particularly useful for antisense studies (please refer to our technical sheet on Antisense Oligonucleotides for other modifications). Custom phosphorothioate oligonucleotides synthesized by Gene Link can be specified to have all the diester bonds substituted or only some selected diester linkages depending upon the researchers experimental requirement. Substitution of all diester linkage is recommended to provide greater nuclease resistance.

References

REFERENCES:
1. Kawasaki, A.M., et al., Uniformly modified 2'-deoxy-2'-fluoro phosphorothioate oligonucleotides as nuclease resistant antisense compounds with high affinity and specificity for RNA targets, Journal of Medicinal Chemistry (1993), 36: 831-841.
2. Khati, M.; Schuman, M.; Ibrahim, J.; Sattentau, Q.; Gordon, S.; James, W. Neutralization of infectivity of diverse R5 clinical isolates of human immunodeficiency virus type 1 by gp120-binding 2'F-RNA aptamers. J Virol, (2003). 77: 2692-8.
3. Goringer, H.U.; Adler, Annette; Forster, Nicole; Homann, Matthias. Post-SELEX Chemical Optimization of a Trypanosome-Specific RNA Aptamer. Combinatorial Chemistry & High Throughput Screening (2008), 11: 16-23.
4. Layzer, J.M.; McCaffrey, A.P.; Tanner, A.K.; Huang, Z.; Kay, M.A.; Sullenger, B.A. In vivo activity of nuclease-resistant siRNAs. RNA (2004), 10, 766-71.
5. Allerson, C.R.; et al. Fully 2'-modified oligonucleotide duplexes with improved in vitro potency and stability compared to unmodified small interfering RNA. Journal of Medicinal Chemistry (2005), 48: 901-904.