Granted Patent
B2
US 12,416,002 · App. 16/456,763 · Granted Sep 16, 2025
Analysis system for orthogonal access to and tagging of biomolecules in cellular compartments
Inventors:
Ramesh Ramji (San Diego, CA); Frank J. Steemers (San Diego, CA); Lena Christiansen (San Diego, CA); Dmitry K. Pokholok (San Diego, CA); Fan Zhang (San Diego, CA)
Assignee:
Illumina, Inc.
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Abstract
The invention relates to a system and methods for enhancing access to nuclear informational molecules, such as DNA, RNA, and proteins, by analytical biomolecules, such as transposome complexes, by treating nuclei with a nuclear permeability enhancer, and to methods of using nuclear membrane, cell membrane, and external compartmentalization approaches as contiguity preserving elements.
Claims (5)
1. A method of reacting a nuclear target nucleic acid with a transposome complex, comprising: (a) contacting a cell nucleus comprising the nuclear target nucleic acid with an aliphatic alcohol chosen from trans-1,2-cyclohexanediol, n-hexane-1,2-diol, and 1,6-hexane-diol and reacting a transposome complex with the nuclear target nucleic acid, wherein reacting comprises fragmenting the nuclear target nucleic acid into double-stranded nucleic acid fragments, each having a 3′ end and a 5′ end, and (b) tagging the double-stranded nucleic acid fragments with the first adaptor sequence at the 5′ end of the first strand of the double-stranded nucleic acid fragments and the second adaptor sequence at the 5′ end of second strand of the double-stranded nucleic acid fragments to form tagged double-stranded nuclear nucleic acid fragments, wherein the transposome complex comprises: (i) a transposase enzyme, (ii) a first transposon end composition comprising a first transposon end sequence comprising a first adaptor sequence, and (iii) a second transposon end composition comprising a second transposon end sequence comprising a second adaptor sequence, and wherein the transposase enzyme is non-covalently bound to the first transposon end composition and the second transposon end composition.
2. The method of claim 1 , wherein the transposome complex further comprises a fluorescein label, thereby delivering a fluorescein-labeled transposome complex into the cell nucleus.
3. The method of claim 2 , wherein the fluorescein label is a fluorescein amidite (FAM) label.
4. The method of claim 3 , further comprising isolating the cell nucleus treated with FAM-labeled transposome complex in the presence of the aliphatic alcohol chosen from trans-1,2-cyclohexanediol, n-hexane-1,2-diol, and 1,6-hexane-diol.
5. The method of claim 3 , further comprising visualizing the cell nucleus treated with FAM-labeled transposome complexes in the presence of the aliphatic alcohol chosen from trans-1,2-cyclohexanediol, n-hexane-1,2-diol, and 1,6-hexane-diol.
Assignments (1)
ASSIGNMENT OF ASSIGNOR'S INTEREST
Recorded Jul 30, 2020
From: RAMJI, RAMESH; STEEMERS, FRANK J; CHRISTIANSEN, LENA; POKHOLOK, DMITRY K; ZHANG, FAN
To: ILLUMINA, INC.
Reel/Frame 053359/0627 →
Continuity (3)
Continuation
PCTUS2017068672
· Dec 28, 2017
Provisional Application
62440089
· Dec 29, 2016
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