Quality assessment of biobanked nucleic acid extracts for downstream molecular analysis.
Sample quality is of major importance when conducting molecular analysis of nucleic acids, and factors such as degradation, presence of impurities, and enzymatic inhibitors may have a significant impact on the quality of data. Issues of quality assessment become more important as the increased use of biobanking means that whole blood samples are being stored for longer periods. A range of commercially available kits/methods have been specifically developed for extraction of high quality nucleic acids from a variety of clinical samples, including blood, but there is limited information on how best to quality assess the extracts to determine their fitness for purpose in specific downstream applications. In this study, we have performed nucleic acid extractions from stored blood using a number of different methods. The resulting extracts were analyzed by a panel of quantity and quality metrics including UV spectrophotometry, PicoGreen fluorescence, electrophoresis, and a PCR approach. To evaluate the relevance of different metrics, DNA samples were subsequently assessed for their performance in real time PCR and microarray experiments. Our findings demonstrate that the most suitable extraction technique and quality assessment approach depends on the required downstream analytical method.
LGC , Teddington, Middlesex, United Kingdom .
Karin
K
Wahlberg
Jim
J
Huggett
Rebecca
R
Sanders
Alexandra S
AS
Whale
Claire
C
Bushell
Ramnath
R
Elaswarapu
Daniel J
DJ
Scott
Carole A
CA
Foy
D001771:Blood Banks / Q000458:organization & administration; D004586:Electrophoresis; D006801:Humans; D008297:Male; D009696:Nucleic Acids / Q000097:blood* / Q000302:isolation & purification*; D011786:Quality Control; D012313:RNA / Q000302:isolation & purification; D060888:Real-Time Polymerase Chain Reaction; D013056:Spectrophotometry, Ultraviolet
D009696:Nucleic Acids; D012313:RNA