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Title: Method of quantitating dsDNA

Abstract

A method for quantitating dsDNA in an aqueous sample solution containing an unknown amount of dsDNA. A first aqueous test solution containing a known amount of a fluorescent dye-dsDNA complex and at least one fluorescence-attenutating contaminant is prepared. The fluorescence intensity of the test solution is measured. The first test solution is diluted by a known amount to provide a second test solution having a known concentration of dsDNA. The fluorescence intensity of the second test solution is measured. Additional diluted test solutions are similarly prepared until a sufficiently dilute test solution having a known amount of dsDNA is prepared that has a fluorescence intensity that is not attenuated upon further dilution. The value of the maximum absorbance of this solution between 200-900 nanometers (nm), referred to herein as the threshold absorbance, is measured. A sample solution having an unknown amount of dsDNA and an absorbance identical to that of the sufficiently dilute test solution at the same chosen wavelength is prepared. Dye is then added to the sample solution to form the fluorescent dye-dsDNA-complex, after which the fluorescence intensity of the sample solution is measured and the quantity of dsDNA in the sample solution is determined. Once the thresholdmore » absorbance of a sample solution obtained from a particular environment has been determined, any similarly prepared sample solution taken from a similar environment and having the same value for the threshold absorbance can be quantified for dsDNA by adding a large excess of dye to the sample solution and measuring its fluorescence intensity.« less

Inventors:
 [1];  [1];  [1]
  1. Los Alamos, NM
Issue Date:
Research Org.:
Los Alamos National Lab. (LANL), Los Alamos, NM (United States)
OSTI Identifier:
874251
Patent Number(s):
6350578
Assignee:
The Regents of the University of California (Los Alamos, NM)
Patent Classifications (CPCs):
C - CHEMISTRY C12 - BIOCHEMISTRY C12Q - MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS
Y - NEW / CROSS SECTIONAL TECHNOLOGIES Y10 - TECHNICAL SUBJECTS COVERED BY FORMER USPC Y10T - TECHNICAL SUBJECTS COVERED BY FORMER US CLASSIFICATION
DOE Contract Number:  
W-7405-ENG-36
Resource Type:
Patent
Country of Publication:
United States
Language:
English
Subject:
method; quantitating; dsdna; aqueous; sample; solution; containing; amount; fluorescent; dye-dsdna; complex; fluorescence-attenutating; contaminant; prepared; fluorescence; intensity; measured; diluted; provide; concentration; additional; solutions; similarly; sufficiently; dilute; attenuated; dilution; value; maximum; absorbance; 200-900; nanometers; nm; threshold; identical; chosen; wavelength; dye; added; form; dye-dsdna-complex; quantity; determined; obtained; environment; similar; quantified; adding; excess; measuring; fluorescent dye; /435/436/999/

Citation Formats

Stark, Peter C, Kuske, Cheryl R, and Mullen, Kenneth I. Method of quantitating dsDNA. United States: N. p., 2002. Web.
Stark, Peter C, Kuske, Cheryl R, & Mullen, Kenneth I. Method of quantitating dsDNA. United States.
Stark, Peter C, Kuske, Cheryl R, and Mullen, Kenneth I. Tue . "Method of quantitating dsDNA". United States. https://www.osti.gov/servlets/purl/874251.
@article{osti_874251,
title = {Method of quantitating dsDNA},
author = {Stark, Peter C and Kuske, Cheryl R and Mullen, Kenneth I},
abstractNote = {A method for quantitating dsDNA in an aqueous sample solution containing an unknown amount of dsDNA. A first aqueous test solution containing a known amount of a fluorescent dye-dsDNA complex and at least one fluorescence-attenutating contaminant is prepared. The fluorescence intensity of the test solution is measured. The first test solution is diluted by a known amount to provide a second test solution having a known concentration of dsDNA. The fluorescence intensity of the second test solution is measured. Additional diluted test solutions are similarly prepared until a sufficiently dilute test solution having a known amount of dsDNA is prepared that has a fluorescence intensity that is not attenuated upon further dilution. The value of the maximum absorbance of this solution between 200-900 nanometers (nm), referred to herein as the threshold absorbance, is measured. A sample solution having an unknown amount of dsDNA and an absorbance identical to that of the sufficiently dilute test solution at the same chosen wavelength is prepared. Dye is then added to the sample solution to form the fluorescent dye-dsDNA-complex, after which the fluorescence intensity of the sample solution is measured and the quantity of dsDNA in the sample solution is determined. Once the threshold absorbance of a sample solution obtained from a particular environment has been determined, any similarly prepared sample solution taken from a similar environment and having the same value for the threshold absorbance can be quantified for dsDNA by adding a large excess of dye to the sample solution and measuring its fluorescence intensity.},
doi = {},
journal = {},
number = ,
volume = ,
place = {United States},
year = {2002},
month = {1}
}

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