5' Adenylation Oligonucleotide Modification
Bio-Synthesis offers 5' adenylation of DNA or RNA synthesis services. 5'-App modified oligonucleotide can be ligated to the 3'-OH of a different sequence. 5' adenylated App oligonuceltoides has been use in applications such as:
- 5'-end labeling
- activated 5'-pyrimidine-rinch RNA
- miRNA library construction/next-generation sequencing
- as activated nucleic acid substrates for in vitro-selected ribo/deoxyribozyme
Bio-Synthesis will custom adenylate an oligonucleotide for use with RNA-Ligase. This method allow us to customize your sequence and introduce various type of modified bases. Our chemical approach also allows for cost effetive scale of your adenylated oligonucleotide that are critical for those developing commercial assays with set linker sequences.
5' Adenylated oligonucleotide Synthesis:
- Synthesis and 5' adenylation
- dual HPLC purification
- 3' blocking group C3 spacer (included)
- Minimum starting scale: 1 umole; Yield ~ 1 nmole
Caution!
5' adenylated oligs require additional purification steps and this modification is not compatible other with heat-sensitive modifications. To prevent a 5' adenylated oligonucleotide from self-ligation, it is
recommended to incorporate a blocking group is required on the 3'-end with a C3 propyl spacer. Unlike ddNuc modifications, the propyl spacer cannot be removed by any 3′ exonuclease activity. Furthermore, the propyl spacer is much less expensive than a ddNuc, while giving a higher yield and a better quality product.
Contact Bio-Synthesis for
Adenylated Oligonucleotide Synthesis Service.
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Product Information
5' Adenylation Oligonucleotide Modification
linker, ligation chemistry
409.02 (100.0%), 410.02 (13.0%), 411.02 (2.1%)
C, 29.35; H, 3.20; N, 17.12; O, 35.19; P, 15.14
Extinction Coefficient, ε(λ):
15400
-20°C To -70°C
Oligonucleotides are stable in solution at 4°C for up to 2 weeks. Properly reconstituted material stored at -20°C should be stable for at least 6 months. Dried DNA (when kept at -20°C) in a nuclease-free environment should be stable for years.
References/Citations:
- Ohtsuka, E., Nishikawa, S., Sugiura, M., Ikehara, M. “Joining of ribooligonucleotides with T4 RNA ligase and identification of the oligonucleotide-adenylate intermediate”, Nucleic Acids Res. (1976), 3: 1613-1623
- Wang, Y., Silverman, S.K. “Efficient RNA 5'-adenylation by T4 DNA ligase to facilitate practical applications” RNA (2006), 12: 1142-1146.
- Pak, J., Fire A. “Distinct Populations of Primary and Secondary Effectors During RNAi in C. elegans”, Science (2007), 315: 241-244.
- Vigneault, F., Sismour, A.M., Church, G.M. “Efficient microRNA capture and bar-coding via enzymatic oligonucleotide adenylation”, Nature Methods (2008), 5: 777-779.
- Flynn-Charlebois, A., Wang, Y., Prior, T.K., Rashid, I., Hoadley, K.A., Coppins, R.L., Wolf, A.C., Silverman, S.K. “Deoxyribozymes with 2'-5' RNA Ligase Activity” J. Am. Chem. Soc. (2003), 125: 2444-2454.
- Milligan, J.F., Groebe, D.R., Witherell, G.W., Uhlenbeck, O.C. (1987) “Oligoribonucleotide synthesis using T7 RNA polymerase and synthetic DNA templates”, Nucl. Acids. Res. (1987), 15: 8783-8798.
- Fukunaga, J., Gouda, M., Umeda, K., Ohno, S., Yokogawa, T., Nishikawa, K. “Use of RNase P for efficient preparation of yeast tRNATyr transcript and its mutants”, J Biochem (Tokyo) (2006), 139: 123-127.
- Proudnikov, D., Mirzabekov, A. “Chemical methods of DNA and RNA fluorescent labeling”, Nucl. Acids Res. (1996), 24: 4535-4542. - Adenylation-5' (rApp)
- Lehman, I.R. “DNA ligase: Structure, mechanism, and function”, Science (1974), 186: 790-797.
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