# US 5,707,812 A · Purification of plasmid DNA during column chromatography

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

- Number: US 5,707,812 A
- Document ID: US5707812A
- Status: Granted
- Kind: A
- Application number: 08/692,590
- Filed: 1996-08-06
- Granted: 1998-01-13
- Earliest claimed: 1996-08-06
- Assignees: Vical Incorporated
- Inventors: Nancy Horn; Greg Budahazi; Magda Marquet
- Web page: https://patentgrep.com/patent/US5707812A

## CPC

- C12N 15/101: {by chromatography, e.g. electrophoresis, ion-exchange, reverse phase}
- C12Q 1/6806: Preparing nucleic acids for analysis, e.g. for polymerase chain reaction [PCR] assay (C12Q1/6804 takes precedence)
- 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

A method for purifying plasmid DNA during column chromatography is provided. A short chain polymeric alcohol, preferably polyethylene glycol, or another DNA condensation agent, is added to a DNA sample prior to column chromatography. The short chain polymeric alcohol or condensation agent promotes improved isolation of plasmid DNA and may be used for large scale purification, particularly for manufacturing plasmid DNA as a biopharmaceutical.

## Claims

### Claim 1

A method of purifying plasmid DNA from lysate contaminants comprising:
(a) preparing a lysate;
(b) clarifying said lysate to produce a clarified lysate;
(c) adding a short chain polymeric alcohol to said clarified lysate;
(d) binding plasmid DNA present in said clarified lysate to a chromatographic matrix by placing said lysate in contact with said chromatographic matrix; and
(e) eluting said plasmid DNA by placing an eluent in contact with said chromatographic matrix.

### Claim 2

A method of purifying plasmid DNA from lysate contaminants comprising:
(a) preparing a lysate;
(b) clarifying said lysate to produce a clarified lysate;
(c) adding a short chain polymeric alcohol to said clarified lysate;
(d) binding plasmid DNA present in said clarified lysate to an ion exchange chromatographic matrix by placing said lysate in contact with said chromatographic matrix; and
(e) eluting said plasmid DNA by placing an eluent in contact with said chromatographic matrix.

### Claim 3 (depends on 2)

The method of claim 2, wherein said ion exchange chromatographic matrix is an anion exchange chromatographic matrix.

### Claim 4 (depends on 1)

The method of any one of claims 1, or 2, wherein the yield of plasmid DNA is greater than or equal to 90%.

### Claim 5 (depends on 1)

The method of any one of claims 1, or 2, wherein said preparing step is lysis with dilute base and detergent.

### Claim 6 (depends on 1)

The method of any one of claims 1, or 2 wherein said clarifying step is filtration with diatomaceous earth.

### Claim 7 (depends on 1)

The method of any one of claims 1, or 2, wherein said short chain polymeric alcohol is polyethylene glycol.

### Claim 8 (depends on 7)

The method of claim 7, wherein said polyethylene glycol amounted in an amount between about 0.1% and about 4% (w/v).

### Claim 9 (depends on 8)

The method of claim 8, wherein said polyethylene glycol is added in an amount of about 1% (w/v).

### Claim 10 (depends on 7)

The method of claim 7, wherein said polyethylene glycol has an average molecular weight of about 7,000 to about 9,000.

### Claim 11 (depends on 10)

The method of claim 10, wherein said polyethylene glycol is PEG-8000.