US 12,264,316 B2
Methods and systems for droplet-based single cell barcoding
Zahra Kamila Belhocine, Fremont, CA (US); Rajiv Bharadwaj, Pleasanton, CA (US); Christopher Hindson, Pleasanton, CA (US); Michael Schnall-Levin, San Francisco, CA (US); Bill Kengli Lin, Pleasanton, CA (US); Anthony Makarewicz, Livermore, CA (US); Pranav Patel, Fremont, CA (US); Katherine Pfeiffer, Berkeley, CA (US); Andrew D. Price, Hayward, CA (US); Mohammad Rahimi Lenji, Pleasanton, CA (US); Tobias Daniel Wheeler, Alameda, CA (US); and Yifeng Yin, Elk Grove, CA (US)
Assigned to 10X GENOMICS, INC., Pleasanton, CA (US)
Filed by 10X Genomics, Inc., Pleasanton, CA (US)
Filed on May 22, 2019, as Appl. No. 16/419,428.
Application 16/419,428 is a continuation of application No. 15/887,947, filed on Feb. 2, 2018, granted, now 10,428,326.
Application 15/887,947 is a continuation of application No. PCT/US2018/016019, filed on Jan. 30, 2018.
Claims priority of provisional application 62/570,783, filed on Oct. 11, 2017.
Claims priority of provisional application 62/500,943, filed on May 3, 2017.
Claims priority of provisional application 62/452,261, filed on Jan. 30, 2017.
Prior Publication US 2020/0024596 A1, Jan. 23, 2020
This patent is subject to a terminal disclaimer.
Int. Cl. C12N 15/10 (2006.01); C12Q 1/6806 (2018.01); C12Q 1/6869 (2018.01)
CPC C12N 15/1065 (2013.01) [C12Q 1/6806 (2013.01); C12Q 1/6869 (2013.01)] 19 Claims
 
1. A method for nucleic acid sequencing, comprising:
(a) providing a plurality of cell beads, wherein a cell bead of said plurality of cell beads comprises a cell, wherein said cell comprises a nucleic acid molecule;
(b) in bulk, processing said plurality of cell beads to denature said nucleic acid molecule to generate a denatured nucleic acid molecule in said cell bead;
(c) within said cell bead, processing said denatured nucleic acid molecule with a reagent to generate a modified nucleic acid molecule in said cell bead, which modified nucleic acid molecule comprises one or more modified nucleic acid bases that are modified with respect to said nucleic acid molecule;
(d) providing a nucleic acid barcode molecule comprising a barcode sequence configured to identify said cell bead;
(e) using said nucleic acid barcode molecule and said modified nucleic acid molecule to generate a barcoded nucleic acid molecule comprising (i) a sequence of said modified nucleic acid molecule or reverse complement thereof and (ii) said barcode sequence or reverse complement thereof; and
(f) sequencing said barcoded nucleic acid molecule or a derivative thereof to identify a sequence of said nucleic acid molecule.