Comparison of methods for accurate quantification of DNA mass concentration with traceability to the international system of units.
Anal Chem
; 82(17): 7185-92, 2010 Sep 01.
Article
em En
| MEDLINE
| ID: mdl-20690645
ABSTRACT
Accurate estimation of total DNA concentration (mass concentration, e.g., ng/muL) that is traceable to the International System of Units (SI) is a crucial starting point for improving reproducible measurements in many applications involving nucleic acid testing and requires a DNA reference material which has been certified for its total DNA concentration. In this study, the concentrations of six different lambda DNA preparations were determined using different measurement platforms UV Absorbance at 260 nm (A(260)) with and without prior sodium hydroxide (NaOH) treatment of the DNA, PicoGreen assay, and digital polymerase chain reaction (dPCR). DNA concentration estimates by A(260) with and without prior NaOH treatment were significantly different for five of the six samples tested. There were no significant differences in concentration estimates based on A(260) with prior NaOH treatment, PicoGreen analysis, and dPCR for two of the three samples tested using dPCR. Since the measurand in dPCR is amount (copy number) concentration (copies/muL), the results suggest that accurate estimation of DNA mass concentration based on copy number concentration is achievable provided the DNA is fully characterized and in the double-stranded form or amplification is designed to be initiated from only one of the two complementary strands.
Texto completo:
1
Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
Espectrofotometria Ultravioleta
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DNA
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Reação em Cadeia da Polimerase
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Fluorometria
Idioma:
En
Revista:
Anal Chem
Ano de publicação:
2010
Tipo de documento:
Article
País de afiliação:
Austrália