Patent ID: 7291487

Claim:
A method for synthesizing a nucleic acid molecule comprising at least one non-canonical nucleotide, comprising the steps of: a) incubating a template nucleic acid in a reaction mixture under nucleic acid synthesis conditions containing (i) a mutant T7 type RNA polymerase expressed from a gene having a mutation at the nucleotide position that corresponds to the Y639 position in T7 RNA polymerase, wherein the polymerase is T3 and the mutation is at amino acid position 640, wherein said polymerase has a reduced discrimination between canonical and non-canonical nucleoside triphosphates, and (ii) at least one non-canonical nucleoside triphosphate, wherein said non-canonical nucleoside triphosphate is incorporated into the synthesized nucleic acid in place of only one canonical nucleoside triphosphate, and b) obtaining the synthesis of a nucleic acid molecule comprising at least one non-canonical nucleotide.