Patent ID: 8389244

Claim:
An amplification-based method for producing at least one mammalian promoter-containing expression cassette, comprising: (i) adding a double stranded nucleic acid comprising a mammalian promoter to an amplification reaction mixture, wherein the double stranded nucleic acid has a sense strand and an antisense strand and wherein each of the sense strand and antisense strand has a 5′ end and a 3′ end, wherein the mammalian promoter is capable of transcribing an RNA molecule in mammalian cells; (ii) adding a first oligonucleotide primer to the reaction mixture, wherein the first oligonucleotide primer is complementary to the 3′ end of the antisense strand of the double stranded nucleic acid; (iii) adding a second oligonucleotide primer to the reaction mixture, wherein the second oligonucleotide primer is complementary to the 3′ end of the sense strand of the double stranded nucleic acid and wherein the second oligonucleotide primer comprises a nucleotide sequence that is complementary to a nucleotide sequence that encodes (1) a sense sequence of a double stranded siRNA molecule and (2) a terminator sequence; and (iv) amplifying the double stranded nucleic acid in a polymerase chain reaction amplification comprising (a) annealing the primers to the complementary strands of the double stranded nucleic acid, (b) extending the annealed primers to produce extension products, (c) denaturing the extension products and (d) repeating the polymerase chain reaction amplification steps a sufficient number of times to produce a first amplified product comprising a first mammalian promoter-containing expression cassette, wherein the first mammalian promoter-containing expression cassette comprises (1) the mammalian promoter, (2) the sense strand of the double stranded siRNA molecule and (3) the terminator sequence.