title: How Severely Is DNA Quantification Hampered by RNA Co-extraction?
abstract: The optional RNase digest that is part of many DNA extraction protocols is often omitted, either because RNase is not provided in the kit or because users do not want to risk contaminating their laboratory. Consequently, co-eluting RNA can become a "contaminant" of unknown magnitude in a DNA extraction. We extracted DNA from liver, lung, kidney, and heart tissues and established that 28-52% of the "DNA" as assessed by spectrophotometry is actually RNA (depending on tissue type). Including an RNase digest in the extraction protocol reduced 260:280 purity ratios. Co-eluting RNA drives an overestimation of DNA yield when quantification is carried out using OD 260 nm spectrophotometry, or becomes an unquantified contaminant when spectrofluorometry is used for DNA quantification. This situation is potentially incompatible with the best practice guidelines for biobanks issued by organizations such as the International Society for Biological and Environmental Repositories, which state that biospecimens should be accurately characterized in terms of their identity, purity, concentration, and integrity. Consequently, we conclude that an RNase digest must be included in DNA extractions if pure DNA is required. We also discuss the implications of unquantified RNA contamination in DNA samples in the context of laboratory accreditation schemes.
authors.affiliation: 1 Integrated Biobank of Luxembourg , Luxembourg .
authors.forename: Ignacio
authors.initials: I
authors.lastname: Sanchez
authors.affiliation: 1 Integrated Biobank of Luxembourg , Luxembourg .|2 Departement Genie Biologique, University of Lorraine , Yutz, France .
authors.forename: Matthieu
authors.initials: M
authors.lastname: Remm
authors.affiliation: 1 Integrated Biobank of Luxembourg , Luxembourg .
authors.forename: Sonia
authors.initials: S
authors.lastname: Frasquilho
authors.affiliation: 1 Integrated Biobank of Luxembourg , Luxembourg .
authors.forename: Fay
authors.initials: F
authors.lastname: Betsou
authors.affiliation: 1 Integrated Biobank of Luxembourg , Luxembourg .
authors.forename: William
authors.initials: W
authors.lastname: Mathieson
mesh_terms: D000818:Animals; D004247:DNA / Q000032:analysis* / Q000737:chemistry*; D016591:Histocytological Preparation Techniques / Q000379:methods / Q000592:standards; D008099:Liver / Q000737:chemistry; D008168:Lung / Q000737:chemistry; D008297:Male; D012313:RNA / Q000032:analysis / Q000737:chemistry / Q000302:isolation & purification*; D051381:Rats; D017207:Rats, Sprague-Dawley; D013053:Spectrophotometry
chemical_list: D012313:RNA; D004247:DNA