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ASSAY FOR SIMULTANEOUS GENOMIC AND PROTEOMIC ANALYSIS

  • US 20180208975A1
  • Filed: 01/17/2018
  • Published: 07/26/2018
  • Est. Priority Date: 01/20/2017
  • Status: Abandoned Application
First Claim
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1. A method for detecting a plurality of target proteins and mRNA in a cell or cell lysate comprising:

  • a. contacting a cell or cell lysate with1) a plurality of protein target probes, wherein each target probe in the plurality comprises;

    i. a protein-binding molecule;

    ii. a target nucleotide sequence comprising a primer for a polymerase chain reaction (PCR), a protein identification sequence that identifies the protein-binding molecule, a linker sequence that hybridizes to a sequence in the detection probe;

    iii. a linker between the protein-binding molecule and the target nucleotide sequence; and

    2) a plurality of microparticles comprising a plurality of detection probes comprising;

    i. a protein detection nucleotide sequence comprising a primer for a polymerase chain reaction (PCR), a cell identification sequence that identifies the cell, a unique molecular identifier (UMI), and a complimentary linker sequence that hybridizes to the linker sequence in the target probe;

    ii. an mRNA detection nucleotide sequence comprising a primer for a polymerase chain reaction (PCR), an cell identification sequence that identifies the cell, a unique molecular identifier, and polydT;

    b. allowing the target nucleotide sequence to hybridize to the protein detection nucleotide sequence and the mRNA to hybridize to the mRNA detection nucleotide sequence in the microparticle;

    c. conducting reverse transcription and/or polymerase extension to generatei. a first analyte sequence comprising the primer, cell identification sequence, unique molecular identifier, and complimentary linker sequence in the detection probe; and

    protein identification sequence and the primer from the protein target probe;

    ii. a second analyte sequence comprising the primer, cell identification sequence, unique molecular identifier, and polydT in the detection probe; and

    cDNA of the mRNA;

    d. enriching the analyte sequences from unbound target and detection nucleotide sequences in the sample;

    e. detecting signals from the analyte sequences based on PCR amplification and sequencing, wherein the signals are distinguishable for each protein and mRNA.

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