# US 10,370,709 B2 · Compositions and methods for sequencing nucleic acids

> Agents: this data is available as JSON from the patentgrep API and MCP server — see https://patentgrep.com/llms.txt

- Number: US 10,370,709 B2
- Document ID: US10370709B2
- Status: Granted
- Kind: B2
- Application number: 15/263,203
- Filed: 2016-09-12
- Published: 2016-12-29
- Granted: 2019-08-06
- Earliest claimed: 2010-09-29
- Assignees: Illumina Cambridge Limited
- Applicants: Ilumina Cambridge Limited
- Inventors: Roberto Rigatti; Jonathan Mark Boutell; Jason Betley; Niall Anthony Gormley
- Web page: https://patentgrep.com/patent/US10370709B2

## CPC

- C12Q 1/6874: involving nucleic acid arrays, e.g. sequencing by hybridisation
- C12Q 1/6806: Preparing nucleic acids for analysis, e.g. for polymerase chain reaction [PCR] assay (C12Q1/6804 takes precedence)
- C12Q 2525/101: incorporating non-naturally occurring nucleotides, e.g. inosine
- C12Q 2525/186: incorporating a non-extendable or blocking moiety
- C12Q 2525/191: incorporating an adaptor
- C12Q: MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS (immunoassay G01N33/53); COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES

## Abstract

Disclosed herein are compositions and methods for sequencing nucleic acids.

## Claims

### Claim 1

A method of generating overlapping sequencing reads, said method comprising:
providing a plurality of polynucleotides;
processing said plurality of polynucleotides by extending random primers hybridized to a polynucleotide of said plurality of polynucleotides and/or randomly fragmenting a double-stranded polynucleotide of said plurality of polynucleotides using transposition, so as to produce a set of overlapping nucleotide regions to be sequenced;
providing a substrate having a surface, said surface comprising a site having said set of overlapping nucleotide regions attached thereto; and
sequencing said set of overlapping nucleotide regions, thereby generating a set of overlapping sequencing reads.

### Claim 2 (depends on 1)

The method of claim 1, further comprising the step of assembling sequencing reads from said set of overlapping sequencing reads so as to obtain a contiguous nucleotide sequence of a nucleic acid of interest.

### Claim 3 (depends on 2)

The method of claim 2, wherein at least one of said polynucleotides comprises said contiguous nucleotide sequence of the nucleic acid of interest.

### Claim 4 (depends on 1)

The method of claim 1, wherein said processing step generates an overlapping set of deletion products from said plurality of polynucleotides.

### Claim 5 (depends on 4)

The method of claim 4, wherein said plurality of nucleotides are attached to said site before said processing step.

### Claim 6 (depends on 4)

The method of claim 4, wherein said plurality of nucleotides are attached to said site after said processing step.

### Claim 7 (depends on 4)

The method of claim 4, further comprising amplifying said set of deletion products on the surface to form amplicons corresponding to the set of deletion products.

### Claim 8 (depends on 1)

The method of claim 1, wherein said random primers comprise a random nucleotide sequence suitable for random priming and initiation of polymerase-mediated extension.

### Claim 9 (depends on 8)

The method of claim 8, wherein said random primers further comprise a sequencing primer site, an adaptor sequence, or a combination thereof.

### Claim 10 (depends on 1)

The method of claim 1, wherein said transposition comprises a transposase enzyme fragmenting said double-stranded polynucleotide at random positions and attaching resulting fragments to oligonucleotide sequences comprising transposon ends.

### Claim 11 (depends on 10)

The method of claim 10, wherein said oligonucleotide sequences further comprise a sequencing primer site, an adaptor sequence, or a combination thereof.

### Claim 12 (depends on 11)

The method of claim 11, which generates deletion products corresponding to end fragments of said double-stranded polynucleotide flanked by one transposon end.

### Claim 13 (depends on 11)

The method of claim 11, which generates deletion products corresponding to internal fragments of said double-stranded polynucleotide flanked by two transposon ends.