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US 2025/0320486 A1Published application
Pre-library target enrichment for nucleic acid sequencing
AssigneesIllumina, Inc., University of Utah Research Foundation
InventorsAlexander A. Gall, Robert Schlaberg
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Abstract
The disclosed technology relates to selectively binding complementary (SBC) oligodeoxynucleotide (ODN) derivatives and their use in enriching targeted dsDNA for sequencing-based detection and genomic profiling.
Claims9
A selectively binding complementary oligodeoxynucleotide derivative having the structure:
- 5′-(Affinity tag)-(SBC sequence 2)-(Spacer)-(SBC sequence 1)-(Blocking group or OH)-3′
- wherein
- the 3′-blocking group is no blocking (3′-OH), or any alkene diol (e.g., OCH2CH2CH2OH), or another biotin with a linker, or stabilizing group (MGB, or intercalators such as phenazinium, acridine, pyrene);
- SBC sequences 1 and 2, each is 5-100 bases long with strands complementary to a selected region of larger targeted dsDNA, such that the targeted DNA preserves a partial double stranded structure after invasion of SBC probe;
- and
- 5′-(Affinity tag) is an affinity tag connected to 5′-end of the probe by a linker.
3 more elements
A method for enriching targeted dsDNA in a DNA sample, comprising:
- (a) mixing a selectively binding complementary (SBC) probe with dsDNA extracted from sample or cDNA produced from corresponding RNA, targeted and non-targeted, wherein the SBC probe is composed of two complementary weakly bound strands tethered with a linker and containing an affinity tag;
- (b) hybridizing each strand of SBC probe with the targeted region of dsDNA and forming two strong double stand areas (double D-loop) within the targeted dsDNA or cDNA;
- (c) treating the mixture with affinity partner-labeled beads (or other solid phase or substrate) and binding the hybridized affinity tagged probe to the beads;
- (d) separating the beads containing targeted DNA bound to the beads from non-targeted DNA in the solution and washing the beads;
- and
- (e) melting off the probe while protecting the rest of the targeted DNA in a double stranded form to provide an enriched dsDNA pool.
3 more elements
(canceled)
Dependencies are inferred from the claim text.