Patent ID: 8536318

Claim:
A method of synthesis of a single-stranded RNA, characterized in that it comprises the following steps: a) binding, to a solid substrate, of a monomer of the following formula II in which: X 1 is a dimethoxytrityl group, X 6 is H or a group OAc or OX 3 in which X 3 is a group of the following formula A: in which X is O or S, R′ is H or CH 3 , and R is selected from a linear or branched C 1 to C 4 alkyl group and a group R 1 —O—R 2 in which R 1 is a C 1 to C 2 alkyl group and R 2 is a CH 3 or CH 2 CH 2 —O—CH 3 or aryl group, Bp is a natural or modified thymine nucleic acid base when X 6 is H or a natural or modified uracil nucleic acid base when X 6 is OAc or OX 3 or a protected natural or modified adenine nucleic acid base or a protected natural or modified cytosine nucleic acid base or a protected natural or modified guanine nucleic acid base regardless of X 6 , b) assembly, with the monomer of formula II bound to its substrate obtained in step a), of at least one monomer of the following formula III: in which X 1 , Bp, X 3 are as defined for formula II and X 5 is a 2-cyanoethyl-N,N-diisopropylphosphoroamidite group, by which a protected single-stranded RNA bound to a substrate of following formula I is obtained in which: X 1 is H or a hydroxyl-protecting group selected from a dimethoxytrityl group, a monomethoxytrityl group and a pixyl group, preferably a dimethoxytrityl group, X 2 is H or a group protecting the β-removable phosphate, preferably a cyanoethyl group, X 3 is a base-labile group protecting the hydroxyls in position 2′ of the ribose of the following formula A: in which X is O or S, R′ is H or CH 3 , and R is selected from a linear or branched C 1 to C 4 alkyl group and a group R 1 —O—R 2 in which R 1 is a C 1 to C 2 alkyl group and R 2 is a CH 3 or CH 2 CH 2 —O—CH 3 or aryl group, X 4 represents the linker-substrate assembly, X 6 is H or a group OX 3 or OAc, Bp is a natural or modified thymine nucleic acid base when X 6 is H or natural or modified uracil when X 6 is OX 3 or OAc, or protected natural or modified adenine, or protected natural or modified cytosine, or protected natural or modified guanine regardless of X 6 , and n is an integer greater than or equal to 0, c) release of the protected single-stranded RNA bound to a substrate obtained in step b), by a method of released comprising a step a1) of treatment of the protected single-stranded RNA bound to a substrate of formula I with a base selected from piperidine, 1,8-diazabicyclo5.4.0 undec-7-ene (DBU), triethylamine at room temperature, to release the phosphate of the 3′-5′ internucleoside linkers, when X 2 is different from H, followed by a step b1) of treatment of the partially released RNA obtained in step a1), with a base selected from concentrated ammonia, methylamine, potassium carbonate, at room temperature.