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US 2018/0105638 A1Published application
Beads for nucleic acid sequencing
AssigneeGenapsys, Inc.
InventorsAldrich N.K. Lau, Paul M. Kenney, Henrik Persson
Life of this patent
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Abstract
Provided herein are methods that comprise providing polymer chains comprising a plurality of reactive groups onto the surface of beads and covalent attachment of functionalized biomolecules, such as primers.
Claims21
-156. (canceled)
A method for preparation of a primer-conjugated bead, comprising:
- (a) coupling a plurality of polymer chains comprising a plurality of first reactive groups to an aminated bead to provide a plurality of second reactive groups coupled to the bead through a respective one of the polymer chains;
- and
- (b) covalently attaching amine- or hydroxy-terminated primers to the plurality of second reactive groups to form the primer-conjugated bead, wherein an individual polymer chain of the plurality of polymer chains comprises a repeat unit comprising a functional group that is different from functional groups of the plurality of first reactive groups and the plurality of second reactive groups, wherein the functional group is hydrophilic, and wherein the individual polymer chain comprises greater than or equal to 40 mole % (mol %) of the functional group.
A reagent comprising beads that are functionalized with polymer chains, wherein a given polymer chain of the polymer chains comprises a carbon backbone of the formula:
- wherein ‘R’ is
- with ‘Q’=NO2 or Fx, with ‘x’=1, 2, 3, 4 or 5;
- wherein ‘T’ is selected from the group consisting of NH2, NH(CH3), N(CH3)2, NH(CH2)tOH, (OCH2CH2)vCH3, (OCH2CH2)vOH, (OCH2CH2)vOCH3 and OCH3 wherein ‘t’ is 1 to 4, ‘v’ is 1 to 100;
- wherein ‘R2’ and ‘R3’ are independently H or CH3;
- wherein ‘y’ and ‘z’ are mole percentages, wherein (‘y’+‘z’)=100 mol %, wherein ‘y’ is less than or equal to 60 mol % and ‘z’ is greater than or equal to 40 mol %;
- and
- wherein ‘W’ is a single bond or
- wherein ‘Di’ is H or CH3, and wherein ‘i’ is 1, 2, 3 or 4, independently.
5 more elements
A method for coupling primers to a bead, comprising:
- (a) providing a bead comprising amine groups linked to a surface of the bead;
- (b) reacting at least a portion of the amine groups with polymer chains comprising a carbon backbone of the formula:
- wherein ‘R’ is
- with ‘Q’=NO2 or Fx, with ‘x’=1, 2, 3, 4 or 5;
- wherein ‘T’ is selected from the group consisting of NH2, NH(CH3), N(CH3)2, NH(CH2)tOH, (OCH2CH2)vCH3, (OCH2CH2)vOH, (OCH2CH2)vOCH3 and OCH3, wherein ‘t’ is 1 to 4, ‘v’ is 1 to 100;
- wherein ‘R2’ and ‘R3’ are independently H or CH3;
- wherein ‘y’ and ‘z’ are mole percentages, wherein (‘y’+‘z’)=100 mol %, wherein ‘y’ is less than or equal to 60 mol % and ‘z’ is greater than or equal to 40 mol %;
- and
- wherein ‘W’ is a single bond or
- wherein ‘Di’ is H or CH3, and wherein ‘i’ is 1, 2, 3 or 4, independently, thereby providing immobilized polymer chains that are coupled to the bead;
- and
- (c) reacting the immobilized polymer chains with a primer comprising an amine group, providing immobilized primers coupled to at least a portion of the immobilized polymer chains.
9 more elements
A reagent comprising beads that are functionalized with polymer chains, wherein a given polymer chain of the polymer chains comprises a carbon backbone of the formula:
- wherein ‘T’ is selected from the group consisting of NH2, NH(CH3), N(CH3)2, NH(CH2)tOH, (OCH2CH2)vCH3, (OCH2CH2)vOH, (OCH2CH2)vOCH3 and OCH3, wherein ‘t’ is 1 to 4, ‘v’ is 1 to 100;
- wherein ‘R2’, ‘R3’, ‘R4’ and ‘R5’ are independently H or CH3;
- and
- wherein ‘y’ and ‘z’ are mole percentages, wherein (‘y’+‘z’)=100 mol %, and wherein ‘y’ is less than or equal to 60 mol % and ‘z’ is greater than or equal to 40 mol %.
1 more element
A method for coupling primers to a bead, comprising:
- (a) providing a bead comprising amine groups linked to a surface of the bead;
- (b) reacting at least a portion of the amine groups with polymer chains comprising a carbon backbone of the formula:
- wherein ‘T’ is selected from the group consisting of NH2, NH(CH3), N(CH3)2, NH(CH2)tOH, (OCH2CH2)vCH3, (OCH2CH2)vOH, (OCH2CH2)vOCH3 and OCH3, wherein ‘t’ is 1 to 4, ‘v’ is 1 to 100;
- wherein ‘R2’, ‘R3’, ‘R4’ and ‘R5’ are independently H or CH3;
- and
- wherein ‘y’ and ‘z’ are mole percentages, wherein (‘y’+‘z’)=100 mol %, wherein ‘y’ is less than or equal to 60 mol %, ‘z’ is greater than or equal to 40 mol %;
- and
- (c) reacting the immobilized polymer chains with a primer comprising an amine group, providing immobilized primers coupled to at least a portion of the immobilized polymer chains.
5 more elements
A primer-coated bead comprising:
- (a) a bead;
- (b) a linker attached to the bead, which linker comprises a polymer chain having a discrete number of repeat units;
- and
- (c) a primer attached to the linker.
1 more element
Dependencies are inferred from the claim text.