# US 2014/0193813 A1 · Isolation of nucleic acids

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- Number: US 2014/0193813 A1
- Document ID: US20140193813A1
- Status: Published application
- Kind: A1
- Application number: 14/145,056
- Filed: 2013-12-31
- Published: 2014-07-10
- Earliest claimed: 2011-05-11
- Assignees: Exact Sciences Corporation
- Applicants: Exact Sciences Corporation
- Inventors: Janelle J. Bruinsma; Hemanth D. Shenoi; Michael J. Domanico; James P. Light, II; Keith Kopitzke; John Zeis
- Web page: https://patentgrep.com/patent/US20140193813A1

## CPC

- C12Q 1/6806: Preparing nucleic acids for analysis, e.g. for polymerase chain reaction [PCR] assay (C12Q1/6804 takes precedence)
- B01D 33/15: with rotary plane filtering surfaces
- B01D 33/155: {the filtering surface being parallel to the rotation axis}
- B01L 3/00: Containers or dishes for laboratory use, e.g. laboratory glassware; Droppers
- B01L 3/5021: {Test tubes specially adapted for centrifugation purposes}
- B03C 1/30: Combinations with other devices, not otherwise provided for
- B04B 3/00: Centrifuges with rotary bowls in which solid particles or bodies become separated by centrifugal force and simultaneous sifting or filtering
- C12N 15/1006: {by means of a solid support carrier, e.g. particles, polymers}
- C12N 15/1013: {by using magnetic beads}
- C12N 15/1017: {by filtration, e.g. using filters, frits, membranes}
- C12Q 1/6886: for cancer (immunoassay for cancer G01N33/575)
- C12Q 2600/158: Expression markers
- C12Q 2600/16: Primer sets for multiplex assays
- Y10T 436/143333: Saccharide [e.g., DNA, etc.]
- 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
- B01D: SEPARATION (separating solids from solids by wet methods B03B, B03D; by pneumatic jigs or tables B03B; by other dry methods B07; magnetic or electrostatic separation of solid materials from solid materials or fluids, separation by high-voltage electric fields B03C; centrifuges, vortex apparatus B04; presses per se for squeezing-out liquid from liquid-containing material B30B9/02; treatment of water C02F, e.g. softening by ion-exchange C02F1/42; {arrangements of air intake cleaners in gas turbine plants F02C7/05} ; arrangements or mounting of filters in air-conditioning, air-humidification or ventilation F24F13/28)
- B01L: CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
- B03C: MAGNETIC OR ELECTROSTATIC SEPARATION OF SOLID MATERIALS FROM SOLID MATERIALS OR FLUIDS; SEPARATION BY HIGH-VOLTAGE ELECTRIC FIELDS (separating isotopes B01D59/00; combinations of magnetic or electrostatic separation with separation of solids by other means B03B, B07B)
- B04B: CENTRIFUGES (high-speed drum mills B02C19/11; domestic spin driers D06F; analysing, measuring or monitoring physical or chemical properties of samples during centrifuging, see the relevant subclasses for these procedures, e.g. G01N)
- C12N: MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA (microbiological testing media C12Q1/00)
- Y10T: TECHNICAL SUBJECTS COVERED BY FORMER US CLASSIFICATION

## Abstract

Provided herein is technology relating to isolating nucleic acids. In particular, the technology relates to methods and kits for extracting nucleic acids from problematic samples such as stool.

## Claims

### Claim 1

A method for removing an assay inhibitor from a crude sample preparation comprising a nucleic acid, the method comprising:
a) adding insoluble polyvinylpyrrolidone to said crude sample preparation prior to isolating the nucleic acid under conditions wherein said assay inhibitor binds to said polyvinylpyrrolidone to produce a complex in a sample to be filtered;
b) separating the complex from the crude sample preparation to produce a clarified sample preparation comprising said nucleic acid
wherein said separating comprises placing said sample to be filtered into a spin filter, said spin filter comprising
i) a hollow body;
ii) a bottom end; and
iii) an open top end opposite the bottom end,
wherein the hollow body is made from a porous filtering material, preferably a polyethylene porous filtering material, and wherein the bottom end is optionally made from said porous filtering material, and
centrifuging said spin filter, wherein during said centrifuging, a fraction of said sample to be filtered passes through a porous filtering material of said spin filter to produce a filtrate.

### Claim 2 (depends on 1)

The method of claim 1 wherein the crude sample preparation is a supernatant prepared from a stool sample.

### Claim 3 (depends on 2)

The method of claim 2 wherein the stool sample has a mass of at least 4 grams.

### Claim 4 (depends on 2)

The method of claim 2 wherein the stool sample has a mass of at least 8 grams.

### Claim 5 (depends on 1)

The method of claim 1 wherein the polyvinylpyrrolidone is a polyvinylpolypyrrolidone.

### Claim 6 (depends on 1)

The method of claim 1, wherein said separating comprises centrifuging.

### Claim 7 (depends on 1)

The method of claim 1 wherein the polyvinylpyrrolidone comprises particles having a diameter averaging from approximately 100 to approximately 130 micrometers.

### Claim 8

A system for removing an assay inhibitor from a crude sample preparation comprising a nucleic acid, the system comprising:
a) insoluble polyvinylpyrrolidone for binding the assay inhibitor and producing a complex;
b) functionality for separating the complex from the crude sample preparation to produce a clarified sample preparation comprising said nucleic acid
wherein said functionality for separating the complex from the crude sample preparation comprises a spin filter comprising
i) a hollow body;
ii) a bottom end; and
iii) an open top end opposite the bottom end,
wherein the hollow body is made from a porous filtering material; and
c) functionality for retaining a clarified sample preparation comprising the nucleic acid.

### Claim 9 (depends on 8)

The system of claim 8 wherein said polyvinylpyrrolidone is a polyvinylpolypyrrolidone.

### Claim 10 (depends on 8)

The system of claim 8, wherein said polyvinylpyrrolidone is in a premeasured form.

### Claim 11 (depends on 8)

The system of claim 8, wherein said polyvinylpyrrolidone is provided as a tablet.

### Claim 12 (depends on 8)

The system of claim 8, wherein the bottom end of said spin filter is made from a porous filtering material.

### Claim 13 (depends on 8)

The system of claim 8, wherein the porous filtering material is polyethylene.

### Claim 14 (depends on 8)

The system of claim 8, wherein the porous filtering material has a nominal pore size of 20 micrometers.

### Claim 15 (depends on 8)

The system of claim 8, wherein the bottom end of said spin filter has a shape selected from the group consisting of a hemisphere, a disc, a cone, or a portion of an ellipsoid.

### Claim 16 (depends on 8)

The system of claim 8, wherein said functionality for retaining a clarified sample preparation comprising the nucleic acid is a collection vessel adapted to receive the spin filter.

### Claim 17

A kit for removing an assay inhibitor from a crude sample preparation comprising a nucleic acid, the kit comprising:
a) insoluble polyvinylpyrrolidone; and
b) a spin column comprising a spin filter comprising:
i) a hollow body;
ii) a bottom end; and
iii) an open top end opposite the bottom end,
wherein the hollow body is made from a porous filtering material.

### Claim 18 (depends on 17)

The kit of claim 17, wherein said polyvinylpyrrolidone is a polyvinylpolypyrrolidone.

### Claim 19 (depends on 17)

The kit of claim 17, wherein said polyvinylpyrrolidone comprises particles having a diameter averaging from approximately 100 to approximately 130 micrometers.

### Claim 20

A system for removing an assay inhibitor from a crude sample preparation comprising a nucleic acid, the system comprising:
a) insoluble polyvinylpyrrolidone; and
b) a spin column comprising a spin filter comprising
i) a hollow body;
ii) a bottom end; and
iii) an open top end opposite the bottom end,
wherein the hollow body is made from a porous filtering material.

### Claim 21 (depends on 20)

The system of claim 20, wherein said polyvinylpyrrolidone is a polyvinylpolypyrrolidone.

### Claim 22 (depends on 20)

The system of claim 20, wherein said polyvinylpyrrolidone comprises particles having a diameter averaging from approximately 100 to approximately 130 micrometers.