Specific quantitation of genomic DNA in the femtogram range by amplification of repetitive sequences.
Anal Biochem
; 242(2): 240-7, 1996 Nov 15.
Article
en En
| MEDLINE
| ID: mdl-8937568
An assay was developed to specifically quantitate concentrations as low as 50 fg/ml genomic DNA based on the amplification of repetitive sequences. Reliable results were obtained by using internal standard molecules which were coextracted, coamplified, and coanalyzed with the nucleic acids of interest. Amplification was performed by the polymerase chain reaction in the presence of fluorescent dye-labeled primers followed by quantitation of fluorescence derived from the reaction products after separation by PAGE. Based on the known amount of added internal standard molecules and the intensities of the fluorescence of the reaction products, the primary results of the assay were obtained as copies per milliliter of sample. These were converted into mass units of DNA by applying an experimentally determined conversion factor. Chicken DNA has been used as an example for genomic DNA, and the sequences amplified were CR1 repetitive elements. This type of assay may be applied in cases where a sensitive and precise quantitation of genomic DNA is required, such as in the quality control of biological products.
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Colección:
01-internacional
Base de datos:
MEDLINE
Asunto principal:
ADN
/
Técnicas de Química Analítica
/
Reacción en Cadena de la Polimerasa
/
Genoma
Tipo de estudio:
Diagnostic_studies
Límite:
Animals
Idioma:
En
Revista:
Anal Biochem
Año:
1996
Tipo del documento:
Article
País de afiliación:
Austria
Pais de publicación:
Estados Unidos