Vibration as a pitfall in pyrosequencing analyses

Since methylation analysis has become an important tool in forensic genetics, the reliability and credibility of the method must be ensured. After a successful validation and establishment of several pyrosequencing assays using a PyroMark® Q48 Autoprep instrument (Qiagen, Hilden, Germany), we decide...

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Autores principales: Konrad, Helen, Schäfer, Laura, Sturm, Hannah, Hördt, Lena, Bajanowski, Thomas, Poetsch, Micaela
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Springer Berlin Heidelberg 2021
Materias:
Method Paper
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8813862/
https://www.ncbi.nlm.nih.gov/pubmed/34637025
http://dx.doi.org/10.1007/s00414-021-02716-7
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author Konrad, Helen
Schäfer, Laura
Sturm, Hannah
Hördt, Lena
Bajanowski, Thomas
Poetsch, Micaela
author_facet Konrad, Helen
Schäfer, Laura
Sturm, Hannah
Hördt, Lena
Bajanowski, Thomas
Poetsch, Micaela
author_sort Konrad, Helen
collection PubMed
description Since methylation analysis has become an important tool in forensic genetics, the reliability and credibility of the method must be ensured. After a successful validation and establishment of several pyrosequencing assays using a PyroMark® Q48 Autoprep instrument (Qiagen, Hilden, Germany), we decided to expand the method further purchasing a second instrument. But after initializing this second instrument side by side with the first, the majority of analyses failed (97 samples of 133 samples (73%)). The number of error messages increased rapidly and the average RFU values decreased. After purchasing two anti-vibration weighing tables for the PyroMark® instruments and repeating the analyses under the same conditions and with identical samples the results improved considerably, 115 samples of 130 samples (88%) showed successful and reproducible results. These findings demonstrate the impact of vibrations and percussions on PyroMark® Q48 Autoprep performance and the reliability of methylation analyses. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1007/s00414-021-02716-7.
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spelling pubmed-88138622022-02-09 Vibration as a pitfall in pyrosequencing analyses Konrad, Helen Schäfer, Laura Sturm, Hannah Hördt, Lena Bajanowski, Thomas Poetsch, Micaela Int J Legal Med Method Paper Since methylation analysis has become an important tool in forensic genetics, the reliability and credibility of the method must be ensured. After a successful validation and establishment of several pyrosequencing assays using a PyroMark® Q48 Autoprep instrument (Qiagen, Hilden, Germany), we decided to expand the method further purchasing a second instrument. But after initializing this second instrument side by side with the first, the majority of analyses failed (97 samples of 133 samples (73%)). The number of error messages increased rapidly and the average RFU values decreased. After purchasing two anti-vibration weighing tables for the PyroMark® instruments and repeating the analyses under the same conditions and with identical samples the results improved considerably, 115 samples of 130 samples (88%) showed successful and reproducible results. These findings demonstrate the impact of vibrations and percussions on PyroMark® Q48 Autoprep performance and the reliability of methylation analyses. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1007/s00414-021-02716-7. Springer Berlin Heidelberg 2021-10-12 2022 /pmc/articles/PMC8813862/ /pubmed/34637025 http://dx.doi.org/10.1007/s00414-021-02716-7 Text en © The Author(s) 2021 https://creativecommons.org/licenses/by/4.0/Open AccessThis article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/ (https://creativecommons.org/licenses/by/4.0/) .
spellingShingle Method Paper
Konrad, Helen
Schäfer, Laura
Sturm, Hannah
Hördt, Lena
Bajanowski, Thomas
Poetsch, Micaela
Vibration as a pitfall in pyrosequencing analyses
title Vibration as a pitfall in pyrosequencing analyses
title_full Vibration as a pitfall in pyrosequencing analyses
title_fullStr Vibration as a pitfall in pyrosequencing analyses
title_full_unstemmed Vibration as a pitfall in pyrosequencing analyses
title_short Vibration as a pitfall in pyrosequencing analyses
title_sort vibration as a pitfall in pyrosequencing analyses
topic Method Paper
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8813862/
https://www.ncbi.nlm.nih.gov/pubmed/34637025
http://dx.doi.org/10.1007/s00414-021-02716-7
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