Sequence tag directed subassembly of short sequencing reads into long sequencing reads

Abstract

The present disclosure relates to a method for preparing a DNA sequencing library that brings more distal fragmentation breakpoints into close proximity to the non-degenerate sequence tag. In an embodiment, the method comprises circularizing a target fragment library with a plurality of adaptor molecules to produce a population of first circularized double-stranded DNA molecules, wherein the plurality of adaptor molecules comprises a first defined sequence P1 comprising a first restriction enzyme recognition site R1, a degenerate sequence tag, and a second defined sequence P2 comprising a second restriction enzyme recognition site R2, such that at least one of the first circularized double-stranded DNA molecule comprises a non-degenerate sequence tag and a member of the target fragment library. The method is useful because some sequencing platforms perform optimally with template molecules that are relatively short, for example, less than about 500 base pairs in length.