Materials and methods for large-scale spatial transcriptomics
AssigneeThe University of Chicago
InventorNICOLAS CHEVRIER
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Abstract
The present disclosure relates to materials and methods for large-scale spatial transcriptomics. In particular, the disclosure provides methods for producing systems for spatial transcriptomics, along with materials and methods for determining the spatial location of a desired nucleic acid, such as RNA, within a tissue sample.
Claims39
A method of producing a system for spatial detection of nucleic acid in a tissue sample, the method comprising:
- a) providing a support comprising an array of surface probes, each surface probe comprising a first anchor sequence, a spatial barcode, and a second anchor sequence;
- b) hybridizing a first complementary oligonucleotide to the first anchor sequence of each surface probe;
- c) hybridizing a second complementary oligonucleotide to the second anchor sequence of each surface probe, wherein the second oligonucleotide comprises a nucleic acid capture region and a unique molecular identifier;
- and
- d) performing an extension-ligation reaction on the support, wherein the extension-ligation reaction comprises extending the first complementary nucleotide with a sequence complementary to the spatial barcode of the surface probe, and ligating the extended first complementary nucleotide and the second complementary nucleotide together to form a contiguous capture oligonucleotide, the capture oligonucleotide comprising a nucleic acid capture region, a unique molecular identifier, a first anchor sequence, a spatial barcode, and a second anchor sequence.
The method of any one of the preceding claims, wherein performing the extension ligation reaction on the support comprises adding a DNA polymerase and a DNA ligase to the support under conditions such that the reverse complement of the spatial barcode sequence is synthesized and ligated to the first complementary nucleotide and to the second complementary nucleotide, thereby forming the continuous capture probe.
A system for spatial detection of nucleic acid in a tissue sample, the system comprising a plurality of spots immobilized on a support, wherein:
- a) each spot comprises a plurality of capture oligonucleotides,
- b) each capture oligonucleotide comprises a nucleic acid capture region, a unique molecular identifier, a first anchor sequence, and a spatial barcode,
- c) each capture oligonucleotide in a single spot comprises the same spatial barcode, and
- d) the spatial barcode for each distinct spot is unique,
- wherein the support comprises a working surface area of at least 2 cm2.
Dependencies are inferred from the claim text.