Ghantous, A and Saffery, R and Cros, M-P and Ponsonby, A-L and Hirschfeld, S and Kasten, C and Dwyer, T and Herceg, Z and Hernandez-Vargas, H, Optimized DNA extraction from neonatal dried blood spots: application in methylome profiling, BMC Biotechnology, 14 Article 60. ISSN 1472-6750 (2014) [Refereed Article]
Licensed under Creative Commons Attribution 4.0 International (CC BY 4.0) http://creativecommons.org/licenses/by/4.0/
Results: Using combinations of in-house-derived and modified commercial extraction kits, we developed a robust and efficient protocol, compatible with methylome studies, many of which require stringent bisulfite conversion steps. Several parameters were tested in a step-wise manner, including blood extraction, cell lysis, protein digestion, and DNA precipitation, purification and elution. DNA quality was assessed based on spectrophotometric measurements, DNA detectability by PCR, and DNA integrity by gel electrophoresis and bioanalyzer analyses. Genome scale Infinium HumanMethylation450 and locus-specific pyrosequencing data generated using the refined DBS extraction protocol were of high quality, reproducible and consistent.
Conclusions: This study may prove useful to meet the increased demand for research on prenatal, particularly epigenetic, origins of human diseases and for newborn screening programs, all of which are often based on DNA extracted from DBS.
|Item Type:||Refereed Article|
|Keywords:||blood spot, DNA extraction, epigenetics, methylome, HM450, pyrosequencing, whole bisulfitome amplification, QIAamp, GenSolve, NucleoSpin|
|Research Division:||Medical and Health Sciences|
|Research Group:||Paediatrics and Reproductive Medicine|
|Objective Group:||Clinical Health (Organs, Diseases and Abnormal Conditions)|
|Objective Field:||Clinical Health (Organs, Diseases and Abnormal Conditions) not elsewhere classified|
|UTAS Author:||Ponsonby, A-L (Professor Anne Ponsonby)|
|UTAS Author:||Dwyer, T (Professor Terry Dwyer)|
|Web of Science® Times Cited:||18|
|Deposited By:||Menzies Institute for Medical Research|
|Downloads:||239 View Download Statistics|
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