Independently removable nucleic acid sequencing system and method

Patent metadata

Field	Value
Publication number	US-12497656-B2
Application number	US-202218050183-A
Country	US
Kind code	B2
Filing date	Oct 27, 2022
Priority date	Jan 26, 2007
Publication date	Dec 16, 2025
Grant date	Dec 16, 2025

Abstract

A technique for sequencing nucleic acids in an automated or semi-automated manner is disclosed. Sample arrays of a multitude of nucleic acid sites are processed in multiple cycles to add nucleotides to the material to be sequenced, detect the nucleotides added to sites, and to de-block the added nucleotides of blocking agents and tags used to identify the last added nucleotide. Multiple parameters of the system are monitored to enable diagnosis and correction of problems as they occur during sequencing of the samples. Quality control routines are run during sequencing to determine quality of samples, and quality of the data collected.

First claim

What is claimed is:

1 . A method, comprising
hybridizing a primer to a first region of a first polynucleotide, extending the primer with a polymerase to incorporate a plurality of sequencing nucleotides wherein each of the plurality of sequencing nucleotides comprises a fluorescent tag and a blocking molecule and extending the primer comprises detecting a fluorescent tag, removing the detected fluorescent tag, and removing a blocking molecule, wherein each of the plurality of sequencing nucleotides is complementary to one of a plurality of sequenced nucleotides of the first polynucleotide adjacent to the first region, and the first polynucleotide comprises a capture molecule covalently bound to a bead and a second region, wherein the bead is attached to a substrate and the second region is complementary to a region of a second polynucleotide, wherein the second polynucleotide comprises a fragment isolated from a biological source, attached to the substrate is a plurality of beads to each of which a plurality of capture molecules are covalently bound, and progressively scanning the substrate to produce image data and processing the image data to identify individual sites and a most recently attached nucleotide at each site.

2 . The method of claim 1 wherein the plurality of beads are randomly distributed on the substrate.

3 . The method of claim 1 wherein the plurality of beads are regularly spaced on the substrate.

4 . The method of claim 2 wherein the plurality of beads are disposed in nanowells of the substrate.

5 . The method of claim 3 wherein the plurality of beads are disposed in nanowells of the substrate.

6 . A method, comprising
hybridizing a primer to a first region of a first polynucleotide, extending the primer with a polymerase to incorporate a plurality of sequencing nucleotides wherein each of the plurality of sequencing nucleotides comprises a fluorescent tag and a blocking molecule and extending the primer comprises detecting a fluorescent tag, removing the detected fluorescent tag, and removing a blocking molecule, wherein each of the plurality of sequencing nucleotides is complementary to one of a plurality of sequenced nucleotides of the first polynucleotide adjacent to the first region, and the first polynucleotide comprises a capture molecule covalently bound to a bead and a second region, wherein the bead is attached to a substrate and the second region is complementary to a region of a second polynucleotide, wherein the second polynucleotide comprises a fragment isolated from a biological source, the substrate comprises a plurality of nanowells and attached to surfaces of the plurality of nanowells is a plurality of beads to each of which a plurality of capture molecules are covalently bound, and progressively scanning the substrate to produce image data and processing the image data to identify individual sites and a most recently attached nucleotide at each site.

7 . The method of claim 6 wherein the plurality of beads are randomly distributed on the substrate.

8 . The method of claim 6 wherein the plurality of beads are regularly spaced on the substrate.

Assignees

• Illumina Inc

Inventors

• Kain Robert C
• Heiner David L
• Zhao Chanfeng
• Gunderson Kevin

Classifications

• C12Q1/6869 Primary

  Methods for sequencing · CPC title

• C12Q1/686

  Polymerase chain reaction [PCR] · CPC title

• C12Q1/6874 Primary

  involving nucleic acid arrays, e.g. sequencing by hybridisation · CPC title