US 12,258,615 B2
Compositions and methods of labeling mitochondrial nucleic acids and sequencing and analysis thereof
Mo Li, Thuwal (SA); and Chongwei Bi, Thuwal (SA)
Assigned to KING ABDULLAH UNIVERSITY OF SCIENCE AND TECHNOLOGY, Thuwal (SA)
Filed by King Abdullah University of Science and Technology, Thuwal (SA)
Filed on Aug. 23, 2021, as Appl. No. 17/409,731.
Application 17/409,731 is a continuation in part of application No. PCT/IB2020/051894, filed on Mar. 4, 2020.
Claims priority of provisional application 63/073,865, filed on Sep. 2, 2020.
Claims priority of provisional application 62/899,432, filed on Sep. 12, 2019.
Claims priority of provisional application 62/899,142, filed on Sep. 11, 2019.
Claims priority of provisional application 62/813,605, filed on Mar. 4, 2019.
Prior Publication US 2022/0056502 A1, Feb. 24, 2022
This patent is subject to a terminal disclaimer.
Int. Cl. C12Q 1/6806 (2018.01); C12Q 1/6869 (2018.01); C40B 40/08 (2006.01); G16B 30/10 (2019.01)
CPC C12Q 1/6806 (2013.01) [C12Q 1/6869 (2013.01); G16B 30/10 (2019.02); C40B 40/08 (2013.01)] 20 Claims
 
1. A method of analyzing full-length genomic mitochondrial DNA comprising
(a) preparing labeled-target mitochondrial DNA comprising polymerase-based extension of one or more unique molecular identifier (UMI) primers along a target mitochondrial DNA template, each UMI primer comprising a universal primer sequence, a unique molecular identifier (UMI) sequence, and a mitochondrial DNA binding sequence,
wherein the source of the target mitochondrial DNA template is any integer between 1 and 100 cells inclusive, or any range formed of two integers there between;
(b) amplifying the labeled-target mitochondrial DNA by one or more rounds of polymerase chain reaction (PCR) comprising a universal primer that binds to the universal primer sequence and one or more mitochondrial DNA primers that bind to a mitochondrial DNA sequence of the labeled-target mitochondrial DNA;
(c) sequencing the amplified labeled-target mitochondrial DNA using long-read sequencing technology;
(d) grouping sequences having the same UMI into one of more groups; and
(e) determining the full-length sequence of each labeled-target mitochondrial DNA by determining a consensus sequence of each UMI group.