Title: Preparation of DNA-containing extract for PCR amplification

Abstract

Environmental samples typically include impurities that interfere with PCR amplification and DNA quantitation. Samples of soil, river water, and aerosol were taken from the environment and added to an aqueous buffer (with or without detergent). Cells from the sample are lysed, releasing their DNA into the buffer. After removing insoluble cell components, the remaining soluble DNA-containing extract is treated with N-phenacylthiazolium bromide, which causes rapid precipitation of impurities. Centrifugation provides a supernatant that can be used or diluted for PCR amplification of DNA, or further purified. The method may provide a DNA-containing extract sufficiently pure for PCR amplification within 5–10 minutes.

Inventors:

Dunbar, John M.; Kuske, Cheryl R.

Issue Date:

2006-07-11

Research Org.:

Los Alamos National Laboratory (LANL), Los Alamos, NM (United States)

Sponsoring Org.:

USDOE

OSTI Identifier:

1175827

Patent Number(s):

7074565

Application Number:

10/439,507

Assignee:

The Regents of the University of California (Los Alamos, NM)

Patent Classifications (CPCs):

C - CHEMISTRY C07 - ORGANIC CHEMISTRY C07H - SUGARS

C - CHEMISTRY C12 - BIOCHEMISTRY C12N - MICROORGANISMS OR ENZYMES

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C - CHEMISTRY C07 - ORGANIC CHEMISTRY C07H - SUGARS

C07H21/04 - with deoxyribosyl as saccharide radical

C - CHEMISTRY C12 - BIOCHEMISTRY C12N - MICROORGANISMS OR ENZYMES

C12N15/1003 - {Extracting or separating nucleic acids from biological samples, e.g. pure separation or isolation methods

Y - NEW / CROSS SECTIONAL TECHNOLOGIES Y10 - TECHNICAL SUBJECTS COVERED BY FORMER USPC Y10S - TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS

Y10S435/912 - Absidia

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DOE Contract Number:  

W-7405-ENG-36

Resource Type:

Patent

Country of Publication:

United States

Language:

English

Subject:

59 BASIC BIOLOGICAL SCIENCES