Two-step synthesis of 5′-deoxy-5′-thioguanosine-5′-monophosphorothioate and its incorporation efficiency into 5′-terminus of RNA for preparation of thiol-functionalized RNA

Tetrahedron Letters

Volumn 51, Issue 26, 2010, Pages 3446-3448

  Kim, Il Hyun ^a   Shin, Seonmi ^a   Jeong, Yong Joo ^b   Hah, Sang Soo ^a  

^a Kyung Hee University   (South Korea)

^b Kookmin University   (South Korea)

Author keywords

Alkaline phosphatase; GSMP; Thiol functionalized RNA

Indexed keywords

5' DEOXY 5' THIOGUANOSINE 5' MONOPHOSPHOROTHIOATE; GUANOSINE DERIVATIVE; PHOSPHOROTHIOIC ACID DERIVATIVE; RNA; RNA POLYMERASE; UNCLASSIFIED DRUG;

ARTICLE; CHEMICAL MODIFICATION; CHEMICAL REACTION; RNA ANALYSIS; RNA STRUCTURE; SYNTHESIS;

EID: 77953119689     PISSN: 00404039     EISSN: None     Source Type: Journal    
DOI: 10.1016/j.tetlet.2010.04.124     Document Type: Article

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26. 2O): δ 7.82 (s, 1H), 5.63 (d, J = 5.9 Hz, 1H), 4.28 (dd, J = 3.9 Hz, 1H), 4.08 (ddd, 2H), 2.83 (m, 2H).
27. 2, and 40 mM Tris buffer (pH 7.9) at 37 °C in a total of 0.2 mL solution. The 71-nt 5′-GSMP-RNA was synthesized by runoff transcription in the presence of GSMP 3 with a ratio of GSMP:GTP:ATP:CTP:UTP = 0:1:1:1:1, 4:1:1:1:1, 8:1:1:1:1, 10:1:1:1:1, 20:1:1:1:1, and 50:1:1:1:1 mM, respectively. Each of the GSMP-labeled RNAs was purified by denaturing 7.5 M urea/8% polyacrylamide gel electrophoresis, and the resulting 5′-GSMP-RNA was dephosphorylated by incubation with 10 units of alkaline phosphatase in buffer 3 (New England Biolab, MA) at 37 °C for 2 h and stopped by incubation with 10 μl of 200 mM EGTA for 10 min at 65 °C. The RNA was recovered by ethanol precipitation, and tested for quantitation of the thiol group using thiol and sulfide quantitation kit (Molecular Probes, OR) according to the manufacturer's manual.
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