Methods, compositions, and kits for determining the location of an analyte in a biological sample

A method for determining a location of a target nucleic acid in a biological sample, the method comprising:

• (a) contacting the biological sample with a plurality of primers, wherein the plurality of primers collectively comprise nucleic acid sequences that hybridize to complementary sequences in the target nucleic acid and a functional domain;
• (b) hybridizing the plurality of primers to the target nucleic acid and extending one or more primers of the plurality of primers using the target nucleic acid as a template to generate one or more extension products;
• (c) incorporating a polynucleotide sequence to the 3′ end of the one or more extension products;
• (d) hybridizing the polynucleotide sequence of the one or more extension products of (c) to a plurality of capture probes on an array, wherein each capture probe in the plurality of capture probes comprises a spatial barcode and a capture domain;
• and
• for an extension product in the one or more extension products, (e) determining (i) the sequence of the spatial barcode of the capture probe to which the extension product is hybridized, or a complement thereof, and (ii) all or a portion of the sequence of the extension product corresponding to the target nucleic acid, or a complement thereof, and using the determined sequences of (i) and (ii) to determine the location of the target nucleic acid in the biological sample.

A kit comprising:

• (a) an array comprising a plurality of capture probes, wherein a capture probe of the plurality of capture probes comprises:
• (i) a spatial barcode;
• and
• (ii) a capture domain that binds a capture sequence, or a complement thereof;
• and
• (b) a plurality of primers, wherein a primer of the plurality of primers comprises a sequence complementary to a target nucleic acid and a functional domain, wherein the functional domain comprises a primer binding sequence or a sequencing specific sequence.

A composition comprising:

• a) a target nucleic acid encoding a B cell receptor or a T cell receptor;
• and
• b) one or more extension products hybridized to the target nucleic acid, wherein the one or more extension products each comprise in a 5′ to 3′ direction:
• i) a primer hybridized to the target nucleic acid in a region encoding a constant region of the B cell receptor or T cell receptor, wherein the primer comprises a functional domain;
• ii) a complementary sequence to the target nucleic acid, preferably wherein the complementary sequence is complementary to a sequence encoding V and J sequences of the B cell receptor or T cell receptor;
• and
• iii) a capture sequence comprising a polynucleotide sequence.

A composition comprising:

• one or more extension products wherein the one or more extension products comprise in a 5′ to 3′ direction comprising:
• i) a primer, wherein the primer comprises a functional domain;
• ii) a complementary sequence, wherein the complementary sequence is complementary to a target nucleic acid that encodes V and J sequences of an immune cell receptor;
• and
• iii) a polynucleotide sequence,
• wherein the polynucleotide sequence is hybridized to a capture domain on an array, wherein the array comprises a plurality of capture probes, wherein a capture probe of the plurality of capture probes comprises a spatial barcode and the capture domain, and wherein the capture domain comprises RNA.

A method for processing a target nucleic acid in a biological sample, the method comprising:

• (a) contacting the biological sample with a primer comprising a nucleic acid sequence that hy bridizes to a complementary sequence in the target nucleic acid and a functional domain;
• (b) hybridizing the primer to the target nucleic acid and extending the primer using the target nucleic acid as a template to generate an extension product;
• (c) incorporating a polynucleotide sequence comprising at least three nucleotides to the 3′ end of the extension product;
• and
• (d) hybridizing the polynucleotide sequence of the extension product to a capture domain on an array, wherein the array comprises a plurality of capture probes, and wherein a capture probe of the plurality of capture probes comprises a spatial barcode and the capture domain.