US 12,215,379 B2
In situ analysis of chromatin interaction
Patrik Ståhl, Stockholm (SE); and Pelin Sahlén, Uppsala (SE)
Assigned to 10X GENOMICS, INC., Pleasanton, CA (US)
Filed by 10x Genomics, Inc., Pleasanton, CA (US)
Filed on Feb. 17, 2021, as Appl. No. 17/177,501.
Claims priority of provisional application 62/977,512, filed on Feb. 17, 2020.
Prior Publication US 2021/0254140 A1, Aug. 19, 2021
Int. Cl. C12Q 1/6818 (2018.01); C12Q 1/6837 (2018.01)
CPC C12Q 1/6818 (2013.01) [C12Q 1/6837 (2013.01)] 20 Claims
 
1. A method, comprising:
(a) contacting a sample with a first chromatin accessing probe and a second chromatin accessing probe simultaneously or sequentially in either order, wherein:
the sample comprises a first chromatin region and a second chromatin region, wherein the first chromatin region and the second chromatin region are separated by a genomic distance of at least 0.5 kilobases (kb), and wherein the first and second chromatin regions interact with each other in a cell and are in the same chromosome in the cell, and
the first chromatin accessing probe hybridizes to the complementary strand of a first nucleic acid strand in the first chromatin region and the second chromatin accessing probe hybridizes to the complementary strand of a second nucleic acid strand in the second chromatin region,
thereby allowing access for binding to the first and second nucleic acid strands;
(b) contacting the sample with a first detection probe and a second detection probe simultaneously or sequentially in either order, wherein:
the first detection probe hybridizes to the first nucleic acid strand and the second detection probe hybridizes to the second nucleic acid strand;
(c) contacting the sample with a first bridging probe that bridges the 3′ end of the first detection probe and the 5′ end of the second detection probe, and a second bridging probe that bridges the 3′ end of the second detection probe and the 5′ end of the first detection probe;
(d) circularizing the first and second bridging probes using the first and second detection probes as templates to form a circular probe, wherein the circular probe comprises a sequence of the first nucleic acid strand and a sequence of the second nucleic acid strand;
(e) generating a rolling circle amplification (RCA) product of the circular probe in situ in the sample; and
(f) detecting the RCA product.