evaluation of whole genome amplified dna to decrease material expenditure and increase quality

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ID: 258588
2017
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Abstract
Aim: The overall aim of this study is to evaluate whole genome amplification of DNA extracted from dried blood spot samples. We wish to explore ways of optimizing the amplification process, while decreasing the amount of input material and inherently the cost. Our primary focus of optimization is on the amount of input material, the amplification reaction volume, the number of replicates and amplification time and temperature. Increasing the quality of the amplified DNA and the subsequent results of array genotyping is a secondary aim of this project. Methods: This study is based on DNA extracted from dried blood spot samples. The extracted DNA was subsequently whole genome amplified using the REPLIg kit and genotyped on the PsychArray BeadChip (assessing > 570,000 SNPs genome wide). We used Genome Studio to evaluate the quality of the genotype data by call rates and log R ratios. Results: The whole genome amplification process is robust and does not vary between replicates. Altering amplification time, temperature or number of replicates did not affect our results. We found that spot size i.e. amount of input material could be reduced without compromising the quality of the array genotyping data. We also showed that whole genome amplification reaction volumes can be reduced by a factor of 4, without compromising the DNA quality. Discussion: Whole genome amplified DNA samples from dried blood spots is well suited for array genotyping and produces robust and reliable genotype data. However, the amplification process introduces additional noise to the data, making detection of structural variants such as copy number variants difficult. With this study, we explore ways of optimizing the amplification protocol in order to reduce noise and increase data quality. We found, that the amplification process was very robust, and that changes in amplification time or temperature did not alter the genotyping calls or quality of the array data. Adding additional replicates of each sample also lead to insignificant changes in the array data. Thus, the amount of noise introduced by the amplification process was consistent regardless of changes made to the amplification protocol. We also explored ways of decreasing material expenditure by reducing the spot size or the amplification reaction volume. The reduction did not affect the quality of the genotyping data.
Reference Key
bkvad-hansen2017molecularevaluation Use this key to autocite in the manuscript while using SciMatic Manuscript Manager or Thesis Manager
Authors ;Marie Bækvad-Hansen;Jonas Bybjerg-Grauholm;Jesper B. Poulsen;Christine S. Hansen;David M. Hougaard;Mads V. Hollegaard
Journal advances in skin & wound care
Year 2017
DOI
10.1016/j.ymgmr.2017.04.002
URL
http://www.sciencedirect.com/science/article/pii/S2214426916301367
Keywords
dried blood spot samplesmedicine (general)biology (general)

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