Reverse transcriptase generates a short hairpin that functions as a primer for second-strand synthesis. The hairpin was cleaved by S1 nuclease, the ends were prepared for the terminal transferase reaction by λ-exonuclease treatment, and oligo-dT was added to the ends. The plasmid PM9 vector was cleaved by EcoRI and subjected to oligo-dA addition. The cDNA was annealed to the plasmid DNA through dA:dT base pairing and the hybrid DNA molecule introduced into E. coli bacterium. Reproduced with permission from Cell (9).