Capturing and amplifying polynucleotides using dendritic molecules

Patent metadata

Field	Value
Publication number	US-2025101505-A1
Application number	US-202418894517-A
Country	US
Kind code	A1
Filing date	Sep 24, 2024
Priority date	Sep 26, 2023
Publication date	Mar 27, 2025
Grant date	—

Abstract

In some examples, a dendritic molecule may include a dendritic core; a seeding primer coupled to the dendritic core; and a plurality of dendrons, each of the dendrons comprising an inert, elongated polymer comprising a first end coupled to the dendritic core and a second end coupled to a first functional group, wherein the first functional group is to react with a second functional group to form a covalent bond. In other examples, a dendritic molecule may include a dendritic core; a single-stranded polynucleotide covalently bonded to the dendritic core; and a plurality of dendrons, each of the dendrons comprising an inert, elongated polymer comprising a first end coupled to the dendritic core and a second end.

First claim

1 . A dendritic molecule for use in capturing a polynucleotide, the dendritic molecule comprising:
a dendritic core; a seeding primer coupled to the dendritic core; and a plurality of dendrons, each of the dendrons comprising an inert, elongated polymer comprising a first end coupled to the dendritic core and a second end coupled to a first functional group, wherein the first functional group is to react with a second functional group to form a covalent bond.

2 . The dendritic molecule of claim 1 , further comprising a polynucleotide comprising a seeding adapter, wherein the seeding adapter is hybridized to the seeding primer.

3 . The dendritic molecule of claim 2 , wherein the polynucleotide is double-stranded, and the seeding adapter is single-stranded.

4 . The dendritic molecule of claim 1 , wherein the dendritic core comprises a branched polypeptide, polyester, ester, amide, ethylene glycol, or oligonucleotide.

5 . The dendritic molecule of claim 1 , wherein the inert, elongated polymer comprises a polypeptide, polyester, polyamide, poly(ethylene glycol), or oligonucleotide.

6 . The dendritic molecule of claim 1 , wherein the first functional end group comprises an alkyne, amine, carboxylic acid, acetylene, N-hydroxy succinimide, biotin, or thiol.

7 . A device, comprising:
a flowcell comprising a plurality of wells, each well comprising a plurality of first functional groups; and a plurality of dendritic molecules, each dendritic molecule comprising:
a dendritic core;
a seeding primer coupled to the dendritic core; and
a plurality of dendrons, each of the dendrons comprising an inert, elongated polymer comprising a first end coupled to the dendritic core and a second end coupled to a second functional group, wherein the second functional group is to react with one of the first functional groups to form a covalent bond,
wherein at least some of the wells contain a single one of the dendritic molecules.

8 . The device of claim 7 , wherein the first functional groups form covalent bonds with respective ones of the second functional groups, wherein the covalent bonds immobilize the dendritic molecule within a respective well.

9 . (canceled)

10 . The device of claim 7 , each well further comprising a hydrogel to which the plurality of first functional groups is coupled.

11 . The device of claim 7 , further comprising a polynucleotide comprising a seeding adapter hybridized to the seeding primer.

12 . The device of claim 11 , wherein the polynucleotide is double-stranded, and wherein the seeding adapter is single-stranded.

13 . The device of claim 7 , each well further comprising a plurality of amplification primers, wherein the amplification primers have different sequences than the seeding primer.

14 . (canceled)

15 . The device of claim 7 , further comprising, within wells in which a respective one of the dendritic molecules is disposed and for which the seeding adapter of one of the polynucleotides is hybridized to the seeding primer of that dendritic molecule, a substantially monoclonal cluster of amplicons.

16 . The device of claim 7 , wherein each of the dendritic molecules contained within a well has a hydrodynamic diameter which is about 60% to about 100% of a diameter of that well.

17 . A method of capturing a polynucleotide in a flowcell, the method comprising:
flowing a plurality of dendritic molecules into a flowcell comprising a plurality of wells, each well comprising a plurality of first functional groups, wherein each dendritic molecule of the plurality comprises:
a dendritic core;
a seeding primer coupled to the dendritic core; and
a plurality of dendrons, each of the dendrons comprising an elongated polymer comprising a first end coupled to the dendritic core and a second end comprising a second functional group;
within at least some of the wells, respectively disposing a respective one of the dendritic molecules within that well; for each well in which a respective one of the dendritic molecules is disposed, forming covalent bonds between the first functional groups of that well and the second functional groups of that dendritic molecule; flowing a plurality of polynucleotides into the flowcell, the polynucleotides respectively comprising seeding adapters; and for each well in which a respective one of the dendritic molecules is disposed, hybridizing a seeding adapter of one of the polynucleotides to the seeding primer of that dendritic molecule.

18 . The method of claim 17 , wherein the covalent bonds immobilize the dendritic molecule within that well.

19 . The method of claim 17 , each well further comprising a hydrogel to which the plurality of first functional groups is coupled.

20 . The method of claim 17 , each well further comprising a plurality of amplification primers, wherein the amplification primers have different sequences than the seeding primer.

21 . (canceled)

22 . The method of claim 17 , further comprising, for each well in which a respective one of the dendritic molecules is disposed and for which the seeding adapter of one of the polynucleotides is hybridized to the seeding primer of that dendritic molecule, using the amplification primers to generate a substantially monoclonal cluster of amplicons of the polynucleotide hybridized to the seeding primer of that dendritic molecule.

23 . The method of claim 17 , wherein each of the dendritic molecules may have a hydrodynamic diameter which is about 60% to about 100% of a diameter of the wells.

24 - 37 . (canceled)

Assignees

• Illumina Inc

Inventors

• Nguyen Nam Trong
• Von Hatten Xavier
• Yang Xiangyuan
• Brustad Eric
• Tovey Will
• Worsdale Matthew
• Artioli Gianluca
• Gal Teodora

Classifications

• C12Q1/6853 Primary

  using modified primers or templates · CPC title

• C12Q1/6806 Primary

  Preparing nucleic acids for analysis, e.g. for polymerase chain reaction [PCR] assay (C12Q1/6804 takes precedence) · CPC title