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US 2025/0320487 A1Published application
Preparation Of Size-Controlled Nucleic Acid Fragments
AssigneeIllumina, Inc.
InventorsNiall Anthony Gormley, Andrew Slatter, Stephen Gross, Kayla Busby, Allison Yunghans, Morgan Roos, Robert Scott Kuersten
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Abstract
A transposome complex capable of producing size-controlled nucleic acid fragments is described herein. In some embodiments, the transposome complex includes multiple inactive transposomes with active transposomes on both ends of the multiple inactive transposomes. Applications, uses, and variations of the disclosed transposome complex include, but are not limited to, library preparation for a nucleic acid and tuning the length of the transposome complex to produce nucleic acid fragments of predetermined or desired lengths.
Claims57
A transposome complex comprising:
- a plurality of inactive transposomes coupled to one another;
- a first active transposome coupled to a first end of the plurality of inactive transposomes;
- a second active transposome coupled to a second end of the plurality of inactive transposomes such that the plurality of inactive transposomes are positioned between the first active transposome and the second active transposome;
- and
- wherein the first active transposome, the second active transposome, and each inactive transposome of the plurality of inactive transposomes comprise a transposase and an adaptor.
2 more elements
A method of preparing a transposome complex, comprising:
- providing an initiator transposome comprising a first oligonucleotide adaptor and a second oligonucleotide adaptor;
- and
- hybridizing at least one linking transposome to the initiator transposome via a linking adaptor of the at least one linking transposome, wherein the at least one linking transposome is catalytically inactive, and wherein the linking adaptor is complementary to the first oligonucleotide adaptor, the second oligonucleotide adaptor, or both;
- and
- coupling at least one terminal transposome to the at least one linking transposome via a terminal adaptor of the terminal transposome that is complementary to the linking adaptor or a different linking adaptor of the at least one linking transposome, wherein the terminal transposome is catalytically active.
2 more elements
A method of preparing a nucleic acid library, comprising:
- contacting target nucleic acids with a plurality of transposome complexes, wherein each transposome complex of the plurality comprises a first active transposome coupled to a second active transposome via an intervening plurality of inactive transposomes, to permit binding of the plurality of transposome complexes to the target nucleic acids;
- and
- tagmenting the target nucleic acids to generate nucleic acid fragments, wherein a size of the generated nucleic acid fragments is a function of a size of an individual transposome complex of the plurality of transposome complexes.
A surface-linked transposome complex comprising:
- a solid surface;
- and
- a plurality of transposomes coupled to the solid surface, and wherein at least one transposome of the plurality of transposomes is inactive based on a modification of an oligonucleotide adaptor of the at least one transposome.
A method of separating nucleic acids, comprising:
- contacting a plurality of inactive transposome complexes with a mixed nucleic acid sample in solution, the mixed nucleic acid sample comprising double-stranded DNA and RNA such that the double-stranded DNA selectively binds to the plurality of inactive transposome complexes relative to the RNA, wherein each inactive transposome complex of the plurality comprises a plurality of inactive transposomes coupled to a surface to permit binding of the plurality of inactive transposome complexes to the double-stranded DNA;
- and
- separating the double-stranded DNA from RNA by removing the plurality of inactive transposomes complexes with bound double-stranded DNA from the solution, the solution comprising the RNA.
A method of normalizing an amount of nucleic acids for a plurality of samples, comprising:
- contacting a first plurality of double-stranded nucleic acids of a first sample with a first plurality of inactive transposome complexes, wherein each inactive transposome complex of the first plurality of transposome complexes comprises a predetermined amount of inactive transposomes coupled to a bead surface, and wherein the contacting is under conditions such that a portion of the first plurality of double-stranded nucleic acids binds to the first plurality of inactive transposome complexes;
- contacting a second plurality of double-stranded nucleic acids of a second sample with a second plurality of inactive transposome complexes, wherein each inactive transposome complex of the second plurality of inactive transposome complexes comprises the predetermined amount of inactive transposomes coupled to a bead surface, and wherein the contacting is under conditions such that a portion of the second plurality of double-stranded nucleic acids binds to the second plurality of inactive transposome complexes;
- and
- sequencing the bound portion of the first plurality of double-stranded nucleic acids and the bound portion of the second plurality of double-stranded nucleic acids.
1 more element
A method of performing a buffer exchange, comprising:
- contacting a plurality of nucleic acids suspended in a first buffer solution with a plurality of inactive transposome complexes, wherein each inactive transposome complex of the plurality of inactive transposome complexes comprises a plurality of inactive transposomes coupled to a surface;
- producing a pellet comprising the plurality of nucleic acids bound to the plurality of inactive transposome complexes;
- separating the pellet from the first buffer solution;
- and
- suspending the pellet in a second buffer solution.
2 more elements
Dependencies are inferred from the claim text.