Exonuclease treatment. The ligated sample was snap-cooled as described above. The exonuclease digestion was carried out in 25 L reaction volume as described below with final oligonucleotide concentrations of 1.5 M for self-annealed ligation, 0.75 M for cohesive end ligation, and 1.5 M for self-annealed ligation. For exclusive Exo I or III digestion, the reaction was carried out with the appropriate exonuclease enzyme (2.4 units μL -1 for Exo I or 12 units L -1 for Exo III) in Exo I buffer (67 mM Glycine-KOH pH 9.5, 6.7 mM MgCl2, 10 mM βmercaptoethanol) or Exo III buffer (10 mM Bis-Tris-Propane-HCl pH 7, 10 mM MgCl2, 1 mM DTT). For double digestion, both Exo I (2.4 units μL -1 ) and III (12 units L -1 ) were used in the stated buffer-salt condition as in Table . When required, the Mg 2+ was supplemented by adding MgSO4 as necessary (NEB # B1003S). All the reactions were supplemented with DTT (1 mM) to ensure complete digestion. The reagents were added while keeping the vials on ice. The solutions were incubated at 37C for 4 hours, followed by enzyme inactivation at 85C for 20 minutes, and then stepwise annealing from 90C to 4C to generate the desired secondary structure.