# US 2013/0252829 A1 · Parallel Methods For Insertional Mutagenesis

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- Number: US 2013/0252829 A1
- Document ID: US20130252829A1
- Status: Published application
- Kind: A1
- Application number: 13/908,191
- Filed: 2013-06-03
- Published: 2013-09-26
- Earliest claimed: 2007-08-09
- Inventors: Michael Paul Strathmann
- Web page: https://patentgrep.com/patent/US20130252829A1

## CPC

- C12N 15/1082: {Preparation or screening gene libraries by chromosomal integration of polynucleotide sequences, HR-, site-specific-recombination, transposons, viral vectors}
- C12Q 1/6869: Methods for sequencing
- C12Q 1/6876: Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes
- C12N: MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA (microbiological testing media C12Q1/00)
- 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

The present invention provides parallel methods for identifying the locations of insertion elements that are distributed at different sites in the genome among a collection of cells.

## Claims

### Claim 1

A parallel method for locating insertion elements in a collection of cells, comprising;
a) integrating insertion elements into host cells to produce a collection of the cells comprising the insertion elements at a plurality of locations in the genomes of the cells wherein the insertion elements are flanked by junctions comprising host DNA;
b) preparing a pool of polynucleotides comprising the junctions from a plurality of the locations;
c) identifying sequences from the host DNA at the junctions by sequencing the pool of polynucleotides in parallel to locate the insertion elements.

### Claim 2 (depends on 1)

The method of claim 1, further comprising isolating a cell from the collection of cells such that the cell comprises at least one of the insertion elements located in step c).

### Claim 3 (depends on 1)

The method of claim 1, wherein the host cells are eukaryotic cells.

### Claim 4 (depends on 1)

The method of claim 1, wherein the insertion elements are derived from transposable elements.

### Claim 5 (depends on 1)

The method of claim 1, wherein the insertion elements are derived from viruses.

### Claim 6 (depends on 1)

The method of claim 1, wherein the pool comprises the junctions from at least 100 distinct locations.

### Claim 7 (depends on 1)

The method of claim 1, wherein the pool comprises the junctions from at least 1000 distinct locations.

### Claim 8 (depends on 1)

The method of claim 1, wherein the polynucleotides comprise sample tags.

### Claim 9 (depends on 8)

The method of claim 8, wherein sequencing the pool of polynucleotides identifies the sequences from the junctions and sequences from the sample tags.

### Claim 10 (depends on 9)

The method of claim 9, wherein the insertion elements comprise the sample tags.

### Claim 11 (depends on 9)

The method of claim 9, wherein the sample tags are genomic tags.

### Claim 12 (depends on 9)

The method of claim 9, wherein the collection of cells comprises a plurality of separate, clonal populations of cells, and further comprising identifying at least one clonal population that contains at least one of the located insertion elements by associating at least one of the sample tags with the clonal population.

### Claim 13 (depends on 12)

The method of claim 12, wherein the step of associating comprises amplifying the sample tags to produce tagged amplicons, and hybridizing the amplicons to an array comprising tag complements.