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. 1987 Jul 10;15(13):5305–5321. doi: 10.1093/nar/15.13.5305

Efficient methods for attachment of thiol specific probes to the 3'-ends of synthetic oligodeoxyribonucleotides.

R Zuckermann, D Corey, P Schultz
PMCID: PMC305963  PMID: 3601673

Abstract

Methodology is described for the synthesis of DNA oligomers containing a free 3'-thiol group which can be selectively crosslinked with a wide variety of probes. This chemistry is compatible with both phosphotriester and phosphoramidite solid phase chemistry. Moreover, the sulphydryl group is introduced into the 3'-nucleoside solid support linkage prior to oligonucleotide synthesis. Consequently, no additional coupling steps are required after oligonucleotide synthesis, and isolation of the 3'-thiol oligonucleotide requires only one additional deprotection step. Cross-linking of the thiol-containing oligonucleotide to a fluorescent probe was carried out with high selectivity, in high yield, and under mild conditions.

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Selected References

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  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. 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]
  3. 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]
  4. 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]
  5. Connolly B. A., Rider P. Chemical synthesis of oligonucleotides containing a free sulphydryl group and subsequent attachment of thiol specific probes. Nucleic Acids Res. 1985 Jun 25;13(12):4485–4502. doi: 10.1093/nar/13.12.4485. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Cosstick R., McLaughlin L. W., Eckstein F. Fluorescent labelling of tRNA and oligodeoxynucleotides using T4 RNA ligase. Nucleic Acids Res. 1984 Feb 24;12(4):1791–1810. doi: 10.1093/nar/12.4.1791. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Grassetti D. R., Murray J. F., Jr Determination of sulfhydryl groups with 2,2'- or 4,4'-dithiodipyridine. Arch Biochem Biophys. 1967 Mar;119(1):41–49. doi: 10.1016/0003-9861(67)90426-2. [DOI] [PubMed] [Google Scholar]
  8. Hampton A., Kappler F., Chawla R. R. Design of species- or isozyme-specific enzyme inhibitors. 1. Effect of thymidine substituents on affinity for the thymidine site of hamster cytoplasmic thymidine kinase. J Med Chem. 1979 Jun;22(6):621–631. doi: 10.1021/jm00192a005. [DOI] [PubMed] [Google Scholar]
  9. Itakura K., Rossi J. J., Wallace R. B. Synthesis and use of synthetic oligonucleotides. Annu Rev Biochem. 1984;53:323–356. doi: 10.1146/annurev.bi.53.070184.001543. [DOI] [PubMed] [Google Scholar]
  10. Kempe T., Sundquist W. I., Chow F., Hu S. L. Chemical and enzymatic biotin-labeling of oligodeoxyribonucleotides. Nucleic Acids Res. 1985 Jan 11;13(1):45–57. doi: 10.1093/nar/13.1.45. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Leary J. J., Brigati D. J., Ward D. C. Rapid and sensitive colorimetric method for visualizing biotin-labeled DNA probes hybridized to DNA or RNA immobilized on nitrocellulose: Bio-blots. Proc Natl Acad Sci U S A. 1983 Jul;80(13):4045–4049. doi: 10.1073/pnas.80.13.4045. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Lin T. S., Mancini W. R. Synthesis and antineoplastic activity of 3'-azido and 3'-amino analogues of pyrimidine deoxyribonucleoside. J Med Chem. 1983 Apr;26(4):544–548. doi: 10.1021/jm00358a016. [DOI] [PubMed] [Google Scholar]
  13. Murasugi A., Wallace R. B. Biotin-labeled oligonucleotides: enzymatic synthesis and use as hybridization probes. DNA. 1984 Jun;3(3):269–277. doi: 10.1089/dna.1.1984.3.269. [DOI] [PubMed] [Google Scholar]
  14. Richardson R. W., Gumport R. I. Biotin and fluorescent labeling of RNA using T4 RNA ligase. Nucleic Acids Res. 1983 Sep 24;11(18):6167–6184. doi: 10.1093/nar/11.18.6167. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Smith L. M., Fung S., Hunkapiller M. W., Hunkapiller T. J., Hood L. E. The synthesis of oligonucleotides containing an aliphatic amino group at the 5' terminus: synthesis of fluorescent DNA primers for use in DNA sequence analysis. Nucleic Acids Res. 1985 Apr 11;13(7):2399–2412. doi: 10.1093/nar/13.7.2399. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Wachter L., Jablonski J. A., Ramachandran K. L. A simple and efficient procedure for the synthesis of 5'-aminoalkyl oligodeoxynucleotides. Nucleic Acids Res. 1986 Oct 24;14(20):7985–7994. doi: 10.1093/nar/14.20.7985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Zoller M. J., Smith M. Oligonucleotide-directed mutagenesis of DNA fragments cloned into M13 vectors. Methods Enzymol. 1983;100:468–500. doi: 10.1016/0076-6879(83)00074-9. [DOI] [PubMed] [Google Scholar]

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