×

Single cell bar-coding for antibody discovery

  • US 9,816,088 B2
  • Filed: 03/14/2014
  • Issued: 11/14/2017
  • Est. Priority Date: 03/15/2013
  • Status: Active Grant
First Claim
Patent Images

1. A method comprising:

  • (a) forming a plurality of first vessels each comprising;

    (i) a single cell, and(ii) a single solid support;

    (b) copying onto the single solid support;

    (i) a first copy of a first cell polynucleotide from the single cell, and(ii) a second copy of a second cell polynucleotide from the single cell;

    (c) forming a plurality of second vessels each comprising(i) a single solid support from the plurality of first vessels, and(ii) a barcoded polynucleotide; and

    (d) amplifying(i) the first copy and the second copy with a first primer set, wherein the first primer set comprises a first primer complementary to a 3′

    portion of the first cell polynucleotide and a 3′

    portion of the second cell polynucleotide, and a second primer complementary to a 5′

    portion of the first cell polynucleotide and a 5′

    portion of the second cell polynucleotide, and(ii) the barcoded polynucleotide with a second primer set, wherein a primer of the second primer set comprises a third primer complementary to a forward primer binding sequence of the barcoded polynucleotide, and a fourth primer complementary to a reverse primer binding sequence of the barcoded polynucleotide,wherein the first primer and the fourth primer are complementary, the second primer and third primer are complementary, the first primer and the third primer are complementary, or the second primer and the fourth primer are complementary; and

    wherein the amplifying comprises extending the first copy when hybridized to the barcoded polynucleotide or an amplicon thereof, and extending the second copy when hybridized to the barcoded polynucleotide or an amplicon thereof;

    thereby forming first and second single cell barcoded sequences.

View all claims
  • 3 Assignments
Timeline View
Assignment View
    ×
    ×