Methods and compositions for nucleic acid analysis
The present invention is directed to methods, compositions and systems for analyzing sequence information while retaining structural and molecular context of that sequence information.
1. A method of analyzing nucleic acids obtained from a formalin-fixed paraffin embedded (FFPE) tissue sample while retaining spatial context, comprising:
a) partitioning the nucleic acids obtained from the FFPE tissue sample into a plurality of wells; wherein nucleic acids in spatial proximity to each other in the FFPE tissue sample are introduced into the same well;
b) barcoding partitioned nucleic acids to form a plurality of barcoded nucleic acids, wherein barcoded nucleic acids within a given discrete well each comprise a common partition-specific barcode sequence, such that the barcode sequences identify nucleic acids from a given well;
c) obtaining sequence information from the plurality of barcoded nucleic acids, wherein the sequence information from the plurality of barcoded nucleic acids comprises sequence information of the partition-specific barcode sequence; and
d) attributing the plurality of barcoded nucleic acids to a region of spatial proximity, wherein barcoded nucleic acids derived from a region of spatial proximity in the FFPE tissue sample comprise the same partition-specific barcode sequence.
2. The method of claim 1 , wherein the barcoding comprises amplifying with a primer comprising a barcode sequence.
3. The method of claim 1 , wherein at least two of the nucleic acids partitioned into the same well in the partitioning a) step comprise different sequences.
4. The method of claim 1 , further comprising, prior to (a), a prior step of imaging the FFPE tissue sample.
5. The method of claim 1 , wherein the FFPE tissue sample is a cancer tissue sample.
6. The method of claim 1 , wherein the barcoded nucleic acids in different wells are pooled prior to obtaining sequence information.
7. The method of claim 1 , wherein the sequence information further comprises information relating to a nucleic acid obtained from the FFPE tissue sample.
8. The method of claim 1 , further comprising, prior to a), a prior step of releasing the nucleic acids from the FFPE tissue sample.
9. The method of claim 1 , wherein the nucleic acids obtained from the FFPE tissue sample comprise nucleic acid tags previously applied to the sample.
10. The method of claim 1 , wherein the step of obtaining sequence information comprises high throughput sequencing of the plurality of barcoded nucleic acids.
11. The method of claim 1 , wherein the partition-specific barcode sequences comprise two or more separate subsequences.
12. The method of claim 4 , wherein said imaging comprises imaging tags with optical properties, such that particular tags are associated with particular regions of the imaged sample.
13. The method of claim 12 , further comprising correlating barcoded nucleic acids derived from a region of spatial proximity in the FFPE tissue sample with the imaged tags.