	US 12,480,090 B2
	Contiguity particle formation and methods of use
Frank J. Steemers, Encinitas, CA (US); Ramesh Ramji, San Diego, CA (US); Steven Norberg, La Mesa, CA (US); Lena Christiansen, San Diego, CA (US); Dmitry K. Pokholok, San Diego, CA (US); and Fan Zhang, San Diego, CA (US)
Assigned to ILLUMINA, INC., San Diego, CA (US)
Filed by ILLUMINA, INC., San Diego, CA (US)
Filed on May 16, 2022, as Appl. No. 17/663,485.
Application 17/663,485 is a division of application No. 16/384,741, filed on Apr. 15, 2019, granted, now 11,359,226.
Claims priority of provisional application 62/660,452, filed on Apr. 20, 2018.
Prior Publication US 2022/0333157 A1, Oct. 20, 2022
Int. Cl. C12N 5/00 (2006.01); A61K 47/58 (2017.01); A61K 47/69 (2017.01); C12M 1/12 (2006.01); C12N 5/16 (2006.01); C12Q 1/6806 (2018.01)

CPC C12N 5/0012 (2013.01) [A61K 47/58 (2017.08); A61K 47/6937 (2017.08); C12M 25/16 (2013.01); C12N 5/16 (2013.01); C12Q 1/6806 (2013.01); C12N 2533/40 (2013.01)]

20 Claims

1. A method of encapsulating a single cell within a hollow bead, comprising:

combining a single cell with a polymer in a spacer oil to form a mixture (a);

mixing the mixture (a) with a crosslinking oil to form a porous polymer shell encapsulating the single cell; and

modulating the pore size of the polymer shell, thereby allowing diffusion of reagents through the polymer shell while retaining the single cell,

wherein the mixing comprises inputting the single cell, the polymer, the spacer oil, and the crosslinking oil into a droplet generator, and

wherein the reagents comprise lysing agents, random hexamers, polymerases, transposases, ligases, catalyzing enzymes, deoxynucleotide triphosphates, buffers, divalent cations, primers, or a combination thereof.