A. Annealing of the oligo d(T) stretch of the Affymetrix T7 primer (blue) to the poly A tail of a mRNA molecule (black). Indicated are the location of the core T7 RNA polymerase binding site, the 5′ and 3′ spacer sequences flanking it and the start of reverse transcription (red) by the RT enzyme that produces the first stand cDNA incorporating the complete T7 primer sequence. B. Transcription start (blue arrow) directly 3′ from the RNA polymerase binding domain generates antisense RNA copies that have the 3′ spacer sequence of the T7 primer as a leader sequence at their 5′ side. Between this leader sequence and the gene sequences the copies have a stretch of uridine nucleotides that originate from the poly A tail of the mRNA.