US 12,071,665 B2
Nucleic acid circularization and amplification on a surface
Daan Witters, San Diego, CA (US); Eli N. Glezer, Del Mar, CA (US); Andrew King, San Diego, CA (US); Sabrina Shore, San Diego, CA (US); and Ryan Shultzaberger, San Diego, CA (US)
Assigned to Singular Genomics Systems, Inc., San Diego, CA (US)
Filed by Singular Genomics Systems, Inc., San Diego, CA (US)
Filed on Mar. 1, 2023, as Appl. No. 18/176,616.
Application 18/176,616 is a continuation of application No. PCT/US2021/056349, filed on Oct. 22, 2021.
Claims priority of provisional application 63/104,363, filed on Oct. 22, 2020.
Prior Publication US 2023/0357843 A1, Nov. 9, 2023
Int. Cl. C12Q 1/68 (2018.01); C12Q 1/6874 (2018.01)
CPC C12Q 1/6874 (2013.01) 23 Claims
 
1. A method of amplifying a single-stranded polynucleotide, the method comprising:
contacting a solid support with the single-stranded polynucleotide comprising a first primer binding sequence and a second primer binding sequence, and hybridizing said first primer binding sequence and said second primer binding sequence to a splint oligonucleotide attached to the solid support;
extending the first primer binding sequence and ligating the extended first primer binding sequence to the second primer binding sequence together to form a circular polynucleotide;
extending the splint oligonucleotide with a polymerase to generate an amplification product comprising a complement of the circular polynucleotide; and
hybridizing the amplification product to a second oligonucleotide attached to the solid support comprising a blocking group at a 3′ end that prevents polymerase extension.