| 主权项 |
1. A method for analyzing at least two nucleic acid sequences in a single cell contained within a population of at least 10,000 cells, comprising:
isolating each of a plurality of single cells from a population of at least 10,000 cells in an emulsion microdroplet or a reaction container, introducing a unique barcode sequence comprising at least six nucleotides into each of the plurality of single cells, wherein each barcode sequence is selected from a pool of barcode sequences with greater than 1000-fold diversity in sequence; for each of the plurality of single cells,
providing at least one set of nucleic acid probes, the set comprising a first probe comprising a sequence that is complementary to a nucleic acid sequence that is located at the 5′ end of the barcode sequence, a second probe comprising a sequence that is complementary to a nucleic acid sequence that is located at the 3′ end of the barcode sequence and a second region of sequence that is complementary to a non-human, exogenous sequence, a third probe comprising a sequence that comprises the non-human, exogenous sequence and a sequence that is complementary to a first subsequence of a second target nucleic acid sequence, and a fourth probe comprising a sequence that is complementary to a second subsequence of the second target nucleic acid sequence;amplifying the first and second nucleic acid sequences independently, wherein the first target nucleic acid sequence is amplified using the first probe and the second probe, and wherein the second target nucleic acid sequence is amplified using the third probe and the fourth probe;hybridizing the exogenous sequence to its complement;amplifying the first target nucleic acid sequence, the second target nucleic acid sequence, and the exogenous sequence using the first and fourth probes; performing bulk sequencing of the fused complexes; and identifying a single cell for each of the fused complexes based on the barcode sequence. |
