Lipophilic modification of oligonucleotides

Tetrahedron Letters

Volumn 38, Issue 4, 1997, Pages 691-694

  Tomkins, Justin M ^a   Barnes, Karen J ^a   Blacker, A John ^b   Watkins, William J ^b   Abell, Chris ^a  

^a UNIVERSITY OF CAMBRIDGE   (United Kingdom)

^b ASTRAZENECA   (United Kingdom)

Author keywords

[No Author keywords available]

Indexed keywords

OLIGONUCLEOTIDE;

ARTICLE; CHEMICAL MODIFICATION; CONJUGATION; DRUG SYNTHESIS; LIPOPHILICITY; MASS SPECTROMETRY;

EID: 0031013327     PISSN: 00404039     EISSN: None     Source Type: Journal    
DOI: 10.1016/S0040-4039(96)02394-5     Document Type: Article

References (21)

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6. Stein, C. A.; Pal, R.; DeVico, A. L.; Hoke, G.; Mumbauer, S.; Kinstler, O.; Sarngadharan, M. G.; Letsinger, R. L. Biochemistry 1991, 30, 2439-2444.
7. Krieg, A. M.; Tonkinson, J.; Matson, S.; Zhao, Q.; Saxon, M.; Zhang, L.-M.; Bhanja, U.; Yakubov, L.; Stein, C. A. Proc. Natl. Acad. Sci. USA 1993, 90, 1048-1052.
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13. 1H NMR, infrared and ultraviolet spectroscopy, high resolution mass spectrometry, and melting point.
14. Oligonucleotides 3 and 4 were synthesised by Oswel DNA Service based at the University of Southampton, on an Applied Biosystems ABI 394 DNA synthesiser, using phosphoramidite chemistry.
15. This sequence was chosen to hybridise to one end of the polylinker sequence of pBluescript II SK-, however for this study it should just be considered a typical oligonucleotide.
16. Amino-Modifier C6 dT available from Glen Research.
17. a) The couplings to the N-hydroxysuccinimide esters were performed by heating 20 μg of oligonucleotide 3 or 4 in 20 μ1 HEPES buffer (0.1 M, pH 8.2) with 100 μg ester (dissolved in 20 μ1 acetonitrile) at 50 °C for 16 h.
18. b) The couplings to SPDP were carried out by reacting 40 μg of oligonucleotide 3 or 4 in 20 μ1 HEPES buffer (0.1 M, pH 8.2) with 100 μg of SPDP (dissolved in 20 μ1 acetonitrile) at room temperature for 16 h. The product was purified by spun column chromatography and then diluted with 60 μ1 HEPES buffer (0.1 M, pH 8.2). An aliquot (20 μ1) of this solution was reacted with each thiol (100 μg dissolved in 20 μ1 acetonitrile) at room temperature for 96 h.
19. The samples were loaded onto 1 ml Sephadex G-25 columns with milli Q water as eluant, following the method of Sambrook, J.; Fritsch, E. F.; Maniatis, T. Molecular Cloning A Laboratory Manual; Cold Spring Harbor Laboratory Press, 1989; pp. E.37-E.38.
20. A Tosahaas TSKgel OligoDNA RP column (4.6 mm × 15 cm) was used on a Gilson HPLC system with the following gradient of acetonitrile in 0.1 M ammonium acetate pH 7.0: 0-2 min., 5%; 2-32 min., 5-50%; 32-43 min., 50%; 43-44 min., 50-5%; 44-45 min., 5%. The flow rate was 1 ml/min.
21. ESMS data were acquired on a VG BioQ quadrupole mass spectrometer used in negative ion mode. Samples were run in 100:100:1 water:acetonitrile:880 ammonia.