Linear double stranded DNA coupled to a single support or a tag and methods for producing said linear double stranded DNA

A method for producing RNA in vitro comprising

• (I) producing a linear double stranded DNA comprising the steps of:
• (a) providing linear double stranded DNA comprising a coding strand and a non-coding strand, said coding strand comprising an RNA polymerase promotor sequence element and a coding sequence element followed by a restriction site element;
• (b) incubating said DNA with (i) a modified deoxynucleotide and (ii) an enzyme capable of adding said modified deoxynucleotide at a 3′ end of a strand in order to provide linear double stranded DNA with a modified deoxynucleotide at the 3′ end of each strand;
• (c) coupling the DNA obtained in step (b) via the modified deoxynucleotide at the 3′ end of each strand to a support;
• (d) incubating the DNA obtained in step (c) with a restriction endonuclease recognizing said restriction element in order to provide linear double stranded DNA, wherein the non-coding strand of said DNA is coupled at its 3′ end to a support and wherein said support is the only support coupled to said DNA
• (II) providing (i) ribonucleoside triphosphates and (ii) a DNA-dependent RNA polymerase;
• and
• (III) incubating the DNA provided in step (I)(d) with (i) and (ii) provided in step (II) under suitable conditions in order to produce RNA.