Patent ID: 8628940

Claim:
A method of performing paired-end sequencing on a single template molecule, the method comprising: a) providing a double-stranded nucleic acid molecule comprising a first terminal portion, an intermediate portion, and a second terminal portion; b) ligating a first linker to the first terminal portion of the nucleic acid molecule, wherein the first linker connects the 3′ terminus at the first terminal portion of the sense strand with the 5′ terminus at the first terminal portion of the antisense strand; c) ligating a second linker to the second terminal portion of the nucleic acid molecule, wherein the second linker connects the 3′ terminus at the second terminal portion of the antisense strand with the 5′ terminus at the second terminal portion of the sense strand, and further wherein the nucleic acid molecule having both the first linker and the second linker ligated thereto forms a single-stranded circular template molecule that contains (i) the first linker between the 3′ terminus of the sense strand and the 5′ terminus of the antisense strand, and (ii) the second linker between the 3′ terminus of the antisense strand and the 5′ terminus of the sense strand; and d) subjecting the template molecule to a sequencing process in which sequence reads are generated, for the first terminal portion and the second terminal portion, but sequence reads are not generated for the intermediate portion, thereby performing paired-end sequencing on the template molecule.