Droplet tagging contiguity preserved tagmented DNA
Methods and compositions for maintaining DNA contiguity for sequencing is provided. For example, a plurality of partitions is provided comprising a bead, a forward primer oligonucleotide cleaved from the bead, the forward primer oligonucleotide having a bead-specific barcode and a 3′ end specific for and complementary to a first or second adaptor; a reverse primer oligonucleotide having a 3′ end complementary to the first or second adaptor, wherein the forward primer 3′ end and the reverse primer 3′ end are complementary to different adaptors selected from the first adaptor and the second adaptor; and fragments of genomic DNA reacted with an adapter-loaded tagmentase such that the DNA fragments comprise breakpoints in the fragments and an inserted adaptor at the break points, wherein the tagmentase binds the breakpoints to form linked DNA segments in the form of DNA segment-first adaptor tagmentase second adaptor-(DNA segment-first adaptor tagmentase second adaptor)n-DNA segment, where n is any integer and “-” indicates a covalent linkage.
1. A plurality of partitions, the partitions comprising:
a bead,
a forward primer oligonucleotide cleaved from the bead, the forward primer oligonucleotide having a bead-specific barcode and a 3′ end specific for and complementary to a first or second adaptor;
a reverse primer oligonucleotide having a 3′ end complementary to the first or second adaptor, wherein the forward primer 3′ end and the reverse primer 3′ end are complementary to different adaptors selected from the first adaptor and the second adaptor; and
fragments of genomic DNA reacted with an adapter-loaded tagmentase such that the DNA fragments comprise breakpoints in the fragments and an inserted adaptor at the break points, wherein the tagmentase binds the breakpoints to form linked DNA segments in the form of DNA segment-first adaptor tagmentase second adaptor-(DNA segment-first adaptor tagmentase second adaptor)n-DNA segment, where n is any integer and “-” indicates a covalent linkage.
2. The plurality of partitions of claim 1 , wherein the partitions are droplets in an emulsion.
3. The plurality of partitions of claim 1 , wherein the partitions comprise on average 0.02-3 beads.
4. The plurality of partitions of claim 1 , wherein the genomic DNA is from a single cell.
5. The plurality of partitions of claim 1 , wherein the genomic DNA is from a mammal or plant.
6. The plurality of partitions of claim 1 , wherein the first adaptor and the second adaptor have different sequences.
7. The plurality of partitions of claim 6 , wherein the first adaptor and the second adaptor are less than 50% identical.
8. The plurality of partitions of claim 1 , wherein the first adaptor and the second adaptor have identical sequences.
9. The plurality of partitions of claim 1 , wherein the first adaptor and the second adaptor are linked by both the tagmentase and a linking sequence.
10. The plurality of partitions of claim 9 , wherein the linking sequence comprises a restriction recognition sequence.
11. The plurality of partitions of claim 9 , wherein the linking sequence comprises one or more uracils.
12. The plurality of partitions of claim 9 , wherein the linking sequence comprises one or more ribonucleotide.
13. The plurality of partitions of claim 1 , wherein n is an integer selected from 1-10,000.
14. The plurality of partitions of claim 1 , comprising at least 10,000 different partitions.
15. The plurality of partitions of claim 10 , wherein the partitions further comprise a restriction enzyme that recognizes the restriction recognition sequence.
16. The plurality of partitions of claim 11 , wherein the partitions further comprise a uracil-DNA N-glycosylase.