Methods and compositions for generating asymmetrically-tagged nucleic acid fragments

A method of producing an asymmetrically-tagged nucleic acid, comprising:

• providing a double-stranded nucleic acid fragment, wherein the 5′ ends of the nucleic acid fragment are dephosphorylated;
• ligating a hairpin adapter to the double-stranded nucleic acid fragment to produce a first ligation product, wherein the first ligation product comprises nicks at the dephosphorylated 5′ ends of the nucleic acid fragment;
• contacting the first ligation product to a polymerase, wherein the polymerase binds to the nicks in the first ligation product;
• incubating the polymerase-contacted first ligation product under nucleic acid synthesis conditions, wherein the polymerase extends the adapter in a template-dependent nucleic acid synthesis reaction to produce a nucleic acid synthesis product;
• ligating a second adapter to the nucleic acid synthesis product, wherein the second adapter is ligated to the end opposite the hairpin adapter on the nucleic acid synthesis product, and wherein the second adapter has at least one difference as compared to the hairpin adapter, thereby producing an asymmetrically-tagged nucleic acid.