Methods for attaching adapters to sample nucleic acids

A method of preparing nucleic acids for analysis comprising;

• (a) blunt-ending double-stranded nucleic acids with single-stranded overhangs in a sample by the action of one or more enzymes providing a 5′-3′ polymerase activity and 3′-5′ proof reading activity, and four standard nucleotide types, wherein single-stranded overhangs with 5′ ends serve as templates for extension of a complementary strand by the polymerase activity and single-stranded overhangs with 3′ ends are digested by the proof reading activity producing blunt-ended nucleic acids;
• (b) without separating the blunt-ended nucleic acids from other components of the sample, end-tailing the blunt-ended nucleic acids by action of a polymerase without a 3′-5′ proof reading function, which performs a non-template directed addition of a nucleotide to the 3′ ends of blunt-ended nucleic acids, wherein A is added preferentially to G preferentially to C or T;
• (c) annealing the nucleic acids from step (b) with at least partially double-stranded adapters with a single nucleotide T or C overhangs at a 3′-end;
• and
• (d) ligating the nucleic acids to the adapters.