6-Thio-dG, 4-Thio-dT and 4-Thio-dU are protected as the S-cyanoethyl ether which is stable during synthesis and readily removed by ammonium hydroxide. It is critical to add 50mM sodium hydrosulfide (NaSH) to the ammonium hydroxide used for deprotection. Especially if room temperature deprotection is carried out, this technique radically reduces the level of ammonolysis which would lead to undesired aminated bases. Moreover, it is also desirable to remove the cyanoethyl protecting group (1M DBU in acetonitrile, 2-5h/RT) prior to the ammonium hydroxide cleavage and deprotection.
Frequently Asked Technical Question
QUESTION: Can 4-thiodU be used for crosslinking and how is it prepared?
RESPONSE:4-thioU is efficiently activated for crosslinking by exposure to long-wavelength UV light for up to 10 minutes. Crosslinks are formed with RNA and proteins.
Oligonucleotides containing 4-thiodU (10-1051) and 4-thioT (10-1033) can be produced from the triazole modified phosphoramidites using the convertible nucleoside strategy (1,2). However, we now offer the nuclesides 4-thio-dT (10-1034), 4-thio-dU (10-1052), 2-thio-dT (10-1036), and 4-thio-U (10-3052) for direct incorporation into oligonucleotides and the convertible monomers have been discontinued. A further enhancement of this strategy used 4-thioxU as an intermediated in the preparation of thiocarbonyl crosslinkers (3).
FERENCE(S):
(1) Y.Z. Xu, Q. Zheng, and P.F. Swann, J. Org. Chem., 1992, 57, 3841.
(2) The Glen Report, 1993, 6.1, 1, and references cited therein.
(3) R.S. Coleman and J.M. Siedlecki, Journal of the American Chemical Society, 1992, 114, 9229-9230.
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