Methods of sequencing linked fragments
The invention generally relates to sequencing library preparation methods. In certain embodiments, two template nucleic acids are joined together by a linking molecule, such as a PEG derivative. The linked template nucleic acids is amplified, creating linked amplicons.
1. A method for preparing a sequencing library, the method comprising:
ligating adapters onto a plurality of nucleic acid fragments;
exposing the fragments to joined primers attached to a solid support; and
extending (i) a first primer of the joined primers to make a copy of a sense strand from one fragment of the plurality of fragments and (ii) a second primer of the joined primers to make a copy of a copy of an anti-sense strand of the one fragment, wherein the copy of the sense strand and the copy of the copy of the anti-sense strand are linked copies with an identical sequence;
amplifying the linked copies with the identical sequence to generate amplicons attached to the solid support;
sequencing the amplicons; and
identifying an error introduced during the amplifying step when the sequencing step gives a signal indicating disagreement between two bases at corresponding positions in the amplicons.
2. The method of claim 1 , wherein the extending step is performed within a partition.
3. The method of claim 2 , wherein the partition uses one selected from the group consisting of emulsions, droplets, or wells.
4. The method of claim 2 , wherein the partition comprises a droplet.
5. The method of claim 4 , where the solid support is selected from the group consisting of a microbead, a bead, a channel wall, a microchip, and a surface.
6. The method of claim 4 , wherein solid support is a bead.
7. The method of claim 1 , wherein the solid support is a bead.
8. The method of claim 1 , wherein the adapters comprise universal priming sites.
9. The method of claim 1 , wherein the ligating step further comprises ligating molecular barcodes onto the plurality of nucleic acid fragments.
10. The method of claim 9 , where the molecular barcodes are sense specific.