Fig. 9.5.

Conversion of an RNA sequence into a DNA sequence suitable for cloning into the pRAV12 expression vector. (A) Sequence of the RNA used to crystallize the SAM-I riboswitch in complex with S-adenosylmethionine. (B) DNA sequence encoding this RNA (third block of letters) with the addition at the 5′-end of an EcoRI restriction site (bold, block 1) and T7 RNA polymerase promoter (block 2) and the H∂V ribozyme/NcoI restriction sequence at the 3′-end (block 4). (C) Output from the GeneDesign website, in which the sequence of (B) was input with target primer length of 60 nucleotides, 18 base pair overlap, and 56°C overlap melting temperature. (D) The four DNA oligonucleotides that would be synthesized for use in a PCR reaction to create the full length DNA insert.