Compositions and methods for synthesizing 5′-capped RNAs
Provided herein are methods and compositions for synthesizing 5′Capped RNAs wherein the initiating capped oligonucleotide primers have the general form m7 Gppp[N 2′Ome ] n [N] m wherein m7 G is N7-methylated guanosine or any guanosine analog, N is any natural, modified or unnatural nucleoside, “n” can be any integer from 0 to 4 and “m” can be an integer from 1 to 9.
1. An initiating capped oligonucleotide primer comprising the following structure:
wherein:
each of B 1 through B 10 is independently a natural, modified or unnatural nucleoside base;
M is 0 or 1;
L is 0 or 1;
q 1 is 1 and each of q 2 through q 9 is independently 0 or 1;
each R 1 is independently H or methyl;
R 2 and R 3 are independently H, OH, alkyl, O-alkyl, halogen, a linker or a detectable marker;
each of X 1 through X 13 is independently O or S;
each of Y 1 through Y 13 is independently OH, SH, BH 3 , aryl, alkyl, O-alkyl or O-aryl;
each of Z 0 through Z 22 is independently O, S, NH, CH 2 , C(halogen) 2 or CH(halogen); and
each of R 4 through R 12 are independently H, OH, OMe or a detectable marker,
wherein a nucleic acid backbone of the initiating capped oligonucleotide primer comprises at least one locked nucleic acid unit.
2. The initiating capped oligonucleotide primer of claim 1 , wherein at least one of R 2 or R 4 through R 12 is a linker.
3. The initiating capped oligonucleotide primer of claim 2 , wherein the linker comprises an O-hydrocarbyl moiety.
4. The initiating capped oligonucleotide primer of claim 2 , wherein the linker is a component of a locked nucleic acid unit.
5. The initiating capped oligonucleotide primer of claim 2 , wherein R 2 is a linker.
6. The initiating capped oligonucleotide primer of claim 5 , wherein the linker is a component of a locked nucleic acid unit.