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  4. Chemical Phosphorylation

Chemical Phosphorylation

Oligonucleotides containing a 5'-phosphate group have various applications, and are often used as a means of ligating one oligo to another, e.g. as linkers and adapters, in cloning, gene construction, and ligation in general.

3'-Phosphorylations are used to block enzyme activity. For example, this is an efficient and commonly used PCR blocking technique.

5’-Phosphate Modifications

Phosphorylation of the 5'-terminus on oligonucleotides is routinely achieved, with higher yields than using kinase, using a Chemical Phosphorylation Reagent (also known as CPR or “Phosphate-ON”)1.

Chemical phosphorylation reagents for 5'-terminus modification

Product Description and Advantages
Chemical Phosphorylating Reagent (CPR)
NACE1-008
  • 5' or 3' phosphorylation of an oligonucleotide 
  • DMT group can be used to monitor phosphorylation efficiency 
  • DMT group cannot be used as purification handle because it gets eliminated during the ammonium hydroxide deprotection
  • Product is viscous glass
5'-Phosphorylating Amidite II
NACE1-006
  • 5' phosphorylation of an oligonucleotide
  • DMT group can be used to monitor phosphorylation efficiency and as purification handle
  • Product is viscous glass
Chemical Phosphorylating Reagent (CPR II)2
NACE1-007
  • 5' phosphorylation of an oligonucleotide
  • DMT group can be used to monitor phosphorylation efficiency and as purification handle
  • Product is viscous glass
Solid Chemical Phosphorylating Reagent (solid CPR)3
NACE1-011
  • 5' phosphorylation of an oligonucleotide
  • Available as solid powder, permitting easy weighing, handling and dissolution
  • DMT group can be used to monitor phosphorylation efficiency and as purification handle
  • Suitable for use with Fmoc or levulinyl protected branching monomers

3' Phosphate Modification of an Oligonucleotide

3'-Phosphate CPGs allow direct preparation of oligonucleotides with a 3'-phosphate group. 

Chemical phosphorylation reagents for 3'-terminus modification

Product Pore Size Availability
3'-Phosphate CPG
NACE1-002		 500 Å, 1000 Å, 3000 Å
3'-Phosphate CPG Column
NACE1-003		 1000 Å, 3000 Å (available in different column formats)
DMT-Phosphate-Suc-CPG 
NACE1-009		 500 Å, 1000 Å
DMT-Phosphate-Suc-CPG Column Suc-CPG)
NACE1-010​​​​​​​		 1000 Å, 1400 Å (available in different column formats)

Our 3’-Phosphate CPGs are available in different pore sizes:

CPG Pore Size Application
500 Å Suitable for high yield applications such as therapeutic oligos (≤ 30mers)
1000 Å Suitable for highly modified oligonucleotides (> 20mers)
3000 Å Performs well for very long sequences (> 120mers)

References

  1. A chemical 5'-phosphorylation of oligodeoxyribonucleotides that can be monitored by trityl cation release, T. Horn and M. Urdea, Tetrahedron Lett., 27, 4705-4708, 1986.
  2. A new approach for chemical phosphorylation of oligonucleotides at the 5'-terminus, A. Guzaev, H. Salo, A. Azhayev and H. Lonnberg, Tetrahedron, 51, 9375-9384, 1995.
  3. Chemical phosphorylation of oligonucleotides and reactants therefor, A. Guzaev, A. Azhayev and H. Lonnberg, US Patent No. 5959090, 1999.