Skip to main content
PMC home page
Nucleic Acids Research logoLink to Nucleic Acids Research
. 1987 Jul 10;15(13):5353–5372. doi: 10.1093/nar/15.13.5353

Covalent attachment of oligonucleotides to solid supports.

S S Ghosh, G F Musso
PMCID: PMC305966  PMID: 3601674

Abstract

Coupling efficiencies for the covalent attachment of oligonucleotides (17-29 bases in length) to solid supports derivatized with alkyl-amino and -carboxylic functionalities have been determined. Attachment efficiencies of 60-80% were obtained for coated long-chain alkylamino controlled pore glass (CPG) supports. Similar efficiencies of immobilization were observed for carboxyl-bearing supports, which additionally exhibited lower levels of non-covalent binding. The extent of terminally linked oligonucleotide was determined to be 50-55% of the overall attachment in the carbodiimide-mediated coupling reaction of a 5'-aminohexyl phosphoramidate derivative of a 29-mer to Sephacryl carboxyl support. While lower overall efficiencies of attachment were obtained in the reaction with Sephacryl N-hydroxysuccinimide-activated carboxyl support, greater than 80% of this coupling results in end-attached oligonucleotides.

Full text

PDF
5356

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Agrawal S., Christodoulou C., Gait M. J. Efficient methods for attaching non-radioactive labels to the 5' ends of synthetic oligodeoxyribonucleotides. Nucleic Acids Res. 1986 Aug 11;14(15):6227–6245. doi: 10.1093/nar/14.15.6227. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Arndt-Jovin D. J., Jovin T. M., Bähr W., Frischauf A. M., Marquardt M. Covalent attachment of DNA to agarose. Improved synthesis and use in affinity chromatography. Eur J Biochem. 1975 Jun;54(2):411–418. doi: 10.1111/j.1432-1033.1975.tb04151.x. [DOI] [PubMed] [Google Scholar]
  3. Bayer E., Breitmaier E., Jung G., Parr W. Synthese des C-terminalen Hexapeptidamids von Secretin an einem neuen festen Träger. Hoppe Seylers Z Physiol Chem. 1971 May;352(5):759–760. [PubMed] [Google Scholar]
  4. Benton W. D., Davis R. W. Screening lambdagt recombinant clones by hybridization to single plaques in situ. Science. 1977 Apr 8;196(4286):180–182. doi: 10.1126/science.322279. [DOI] [PubMed] [Google Scholar]
  5. Bridson P. K., Orgel L. E. Catalysis of accurate poly(C)-directed synthesis of 3'-5'-linked oligoguanylates by Zn2+. J Mol Biol. 1980 Dec 25;144(4):567–577. doi: 10.1016/0022-2836(80)90337-x. [DOI] [PubMed] [Google Scholar]
  6. Bünemann H., Westhoff P., Herrmann R. G. Immobilization of denatured DNA to macroporous supports: I. Efficiency of different coupling procedures. Nucleic Acids Res. 1982 Nov 25;10(22):7163–7180. doi: 10.1093/nar/10.22.7163. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Chollet A., Kawashima E. H. Biotin-labeled synthetic oligodeoxyribonucleotides: chemical synthesis and uses as hybridization probes. Nucleic Acids Res. 1985 Mar 11;13(5):1529–1541. doi: 10.1093/nar/13.5.1529. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Chu B. C., Kramer F. R., Orgel L. E. Synthesis of an amplifiable reporter RNA for bioassays. Nucleic Acids Res. 1986 Jul 25;14(14):5591–5603. doi: 10.1093/nar/14.14.5591. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Chu B. C., Orgel L. E. Detection of specific DNA sequences with short biotin-labeled probes. DNA. 1985 Aug;4(4):327–331. doi: 10.1089/dna.1985.4.327. [DOI] [PubMed] [Google Scholar]
  10. Chu B. C., Wahl G. M., Orgel L. E. Derivatization of unprotected polynucleotides. Nucleic Acids Res. 1983 Sep 24;11(18):6513–6529. doi: 10.1093/nar/11.18.6513. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Connolly B. A. The synthesis of oligonucleotides containing a primary amino group at the 5'-terminus. Nucleic Acids Res. 1987 Apr 10;15(7):3131–3139. doi: 10.1093/nar/15.7.3131. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Gisin B. F. The monitoring of reactions in solid-phase peptide synthesis with picric acid. Anal Chim Acta. 1972 Jan;58(1):248–249. doi: 10.1016/S0003-2670(00)86882-8. [DOI] [PubMed] [Google Scholar]
  13. Grunstein M., Hogness D. S. Colony hybridization: a method for the isolation of cloned DNAs that contain a specific gene. Proc Natl Acad Sci U S A. 1975 Oct;72(10):3961–3965. doi: 10.1073/pnas.72.10.3961. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Kafatos F. C., Jones C. W., Efstratiadis A. Determination of nucleic acid sequence homologies and relative concentrations by a dot hybridization procedure. Nucleic Acids Res. 1979 Nov 24;7(6):1541–1552. doi: 10.1093/nar/7.6.1541. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Kaiser E., Colescott R. L., Bossinger C. D., Cook P. I. Color test for detection of free terminal amino groups in the solid-phase synthesis of peptides. Anal Biochem. 1970 Apr;34(2):595–598. doi: 10.1016/0003-2697(70)90146-6. [DOI] [PubMed] [Google Scholar]
  16. Langdale J. A., Malcolm A. D. A rapid method of gene detection using DNA bound to Sephacryl. Gene. 1985;36(3):201–210. doi: 10.1016/0378-1119(85)90175-1. [DOI] [PubMed] [Google Scholar]
  17. Meinkoth J., Wahl G. Hybridization of nucleic acids immobilized on solid supports. Anal Biochem. 1984 May 1;138(2):267–284. doi: 10.1016/0003-2697(84)90808-x. [DOI] [PubMed] [Google Scholar]
  18. Noyes B. E., Stark G. R. Nucleic acid hybridization using DNA covalently coupled to cellulose. Cell. 1975 Jul;5(3):301–310. doi: 10.1016/0092-8674(75)90105-1. [DOI] [PubMed] [Google Scholar]
  19. Panet A., Khorana H. G. Studies on polynucleotides. The linkage of deoxyribopolynucleotide templates to cellulose and its use in their replication. J Biol Chem. 1974 Aug 25;249(16):5213–5221. [PubMed] [Google Scholar]
  20. Siddell S. G. RNA hybridization to DNA coupled with cyanogen-bromide-activated sephadex. The purification of polyoma messenger RNA. Eur J Biochem. 1978 Dec;92(2):621–629. doi: 10.1111/j.1432-1033.1978.tb12785.x. [DOI] [PubMed] [Google Scholar]
  21. Southern E. M. Detection of specific sequences among DNA fragments separated by gel electrophoresis. J Mol Biol. 1975 Nov 5;98(3):503–517. doi: 10.1016/s0022-2836(75)80083-0. [DOI] [PubMed] [Google Scholar]

Articles from Nucleic Acids Research are provided here courtesy of Oxford University Press

RESOURCES