	US 12,227,799 B2
	Compositions and methods for nucleic acid sequencing
Eli N. Glezer, Del Mar, CA (US); Mohammad Vatankhah Varnosfaderani, San Marcos, CA (US); Daan Witters, San Diego, CA (US); and Vahid Karimkhani, San Diego, CA (US)
Assigned to Singular Genomics Systems, Inc., San Diego, CA (US)
Filed by Singular Genomics Systems, Inc., La Jolla, CA (US)
Filed on Dec. 3, 2021, as Appl. No. 17/542,343.
Application 17/542,343 is a continuation of application No. 17/030,886, filed on Sep. 24, 2020, granted, now 11,236,387.
Application 17/030,886 is a continuation of application No. PCT/US2020/017060, filed on Feb. 6, 2020.
Claims priority of provisional application 62/802,062, filed on Feb. 6, 2019.
Prior Publication US 2022/0154271 A1, May 19, 2022
This patent is subject to a terminal disclaimer.
Int. Cl. C12Q 1/6869 (2018.01); C12Q 1/6844 (2018.01); C12Q 1/6874 (2018.01); G01N 33/53 (2006.01); G01N 33/544 (2006.01)

CPC C12Q 1/6869 (2013.01) [C12Q 1/6844 (2013.01); C12Q 1/6874 (2013.01); G01N 33/5308 (2013.01); G01N 33/544 (2013.01); C12Q 2533/101 (2013.01); C12Q 2565/601 (2013.01)]

20 Claims

1. A method of sequencing target polynucleotides in a polymer scaffold comprising polynucleotide primers, wherein said polynucleotide primers are covalently attached to the polymer scaffold, the method comprising:

(a) amplifying the target polynucleotides to produce discrete amplicon clusters within the polymer scaffold,

wherein amplifying comprises rolling circle amplification the amplicon clusters are arranged at a plurality of depths; and

(b) sequencing the amplicon clusters, wherein sequencing comprises detecting sequences of signals within the polymer scaffold at a first depth and a second depth.