Xseq
Provided herein are methods for identification of an expression profile, a transcriptional profile, and/or an epigenetic profile from a cell-containing sample. Also provided are compositions for use in the disclosed methods.
1. A method for biochemical analysis of a cell-containing sample, comprising:
a) contacting a cell-containing sample with a plurality of antigen-binding molecules under conditions that promote specific binding of the antigen-binding molecules to target antigens of the cells in the sample; wherein each antigen-binding molecule in the plurality binds to a unique target antigen in one or more cells of the sample, wherein each antigen-binding molecule is conjugated to an oligonucleotide, and wherein the oligonucleotide comprises (i) a unique molecular identifier sequence, (ii) an antigen-binding molecule identifier sequence corresponding to the unique target antigen, and (iii) an assay molecular identifier sequence;
b) tagmenting genomic DNA in the cells to produce a plurality of double-stranded genomic DNA fragments comprising an oligonucleotide adaptor sequence at the 5′ end of each genomic DNA fragment;
c) lysing the cells in the sample;
d) isolating mRNA transcripts from the cell lysate to produce a first fraction comprising the mRNA transcripts and a second fraction comprising tagmented genomic DNA fragments and the antigen-binding molecule-bound antigens from the cell lysate;
e) generating a cDNA library by reverse transcribing the mRNA transcripts in the first fraction;
f) amplifying (i) the unique molecular identifier sequences; (ii) the antigen-binding molecule identifier sequences; (iii) the assay molecular identifier sequences, and the (iv) tagmented genomic DNA fragments in the second fraction;
g) analyzing the amplified unique molecular identifier sequences, the assay molecular identifier sequences, and the antigen-binding molecule identifier sequences to identify at least one target antigen in the second fraction to identify the expression profile of the sample;
h) analyzing the tagmented genomic DNA fragments in the second fraction to identify the epigenetic profile of the sample; and
i) analyzing the cDNA library generated from the first fraction to identify the transcriptional profile of the sample.
2. The method of claim 1 , further comprising quantifying the amplified unique molecular identifier sequences, antigen-binding molecule identifier sequences and the genomic DNA fragments in the second fraction.
3. The method of claim 1 , wherein analyzing in step g) and step h) comprises sequencing the amplified unique molecular identifier sequences, the antigen-binding molecule identifier sequences, the assay molecular identifier sequences, and the tagmented genomic fragments in the second fraction.
4. The method of claim 1 , wherein the antigen is a protein.
5. The method of claim 4 , wherein the expression of two or more proteins are identified.
6. The method of claim 1 , wherein the method further comprises tagmenting the cDNA library to produce a plurality of double-stranded cDNA fragments comprising an oligonucleotide adaptor sequence at the 5′ end of each cDNA prior to analyzing.
7. The method of claim 1 , wherein the cell-containing sample is a population of cells.
8. The method of claim 7 , wherein single cells of the population are separated into individual compartments prior to lysing the single cell in each compartment.
9. The method of claim 8 , wherein an expression profile, an epigenetic profile and a transcriptional profile are identified for the single cell.
10. The method of claim 8 , wherein the single cells separated into individual compartments are single cells from a subpopulation of the population of cells contacted with the plurality of antigen-binding molecules.
11. The method of claim 10 , wherein the subpopulation of cells is isolated from the population of cells contacted with the plurality of antigen-binding molecules prior to separating.
12. The method of claim 1 , wherein the antigen-binding molecule is an antibody.
13. A method for biochemical analysis of a cell-containing sample, comprising:
a) contacting a cell-containing sample with a plurality of antigen-binding molecules under conditions that promote specific binding of the antigen-binding molecules to target antigens of the cells in the sample; wherein each antigen-binding molecule in the plurality binds to a unique target antigen in one or more cells of the sample, wherein each antigen-binding molecule is conjugated to an oligonucleotide, and wherein the oligonucleotide comprises (i) a unique molecular identifier sequence, (ii) an antigen-binding molecule identifier sequence corresponding to the unique target antigen, and (iii) an assay molecular identifier sequence;
b) tagmenting genomic DNA in the cell lysate to produce a plurality of double-stranded genomic DNA fragments comprising an oligonucleotide adaptor sequence at the 5′ end of each genomic DNA fragment;
c) lysing the cells in the sample;
d) amplifying (i) the unique molecular identifier sequences, (ii) the antigen-binding molecule identifier sequences, and (iii) the assay molecular identifier sequence in the cell lysate;
e) analyzing the amplified unique molecular identifier sequences and antigen-binding molecule identifier sequences to identify at least one target antigen to identify the expression profile in the sample; and
f) analyzing the genomic DNA fragments to identify the epigenetic profile of the sample.
14. The method of claim 13 , further comprising quantifying the amplified unique molecular identifier sequences and antigen-binding molecule identifier sequences.
15. The method of claim 13 , wherein analyzing comprises sequencing the amplified unique molecular identifier sequences, the antigen-binding molecule identifier sequences, the assay molecular identifier sequences, and the tagmented genomic fragments.
16. The method of claim 13 , wherein the antigen is a protein.
17. The method of claim 16 , wherein the expression of two or more proteins are identified.
18. The method of claim 13 , wherein the cell-containing sample is a population of cells.
19. The method of claim 18 , wherein single cells of the population are separated into individual compartments prior to lysing the single cell in each compartment.
20. The method of claim 19 , wherein an expression profile and an epigenetic profile are identified for the single cell.
21. The method claim 19 , wherein the single cells separated into individual compartments are single cells from a subpopulation of the population of cells contacted with the plurality of antigen-binding molecules.
22. The method of claim 13 , wherein the antigen-binding molecule is an antibody.