Reproducibility of next-generation-sequencing-based analysis of a CRISPR/Cas9 genome edited oil seed rape
Next-generation-sequencing (NGS) becomes increasingly important for laboratories tasked with the detection of genetically modified organisms (GMOs) in food, feed and seeds. Its implementation into standardized workflows demands reliable intra- and inter-laboratory reproducibility. Here, we analyze t...
| Main Authors: | , , , , |
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| Format: | Article |
| Language: | English |
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Elsevier
2023-12-01
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| Series: | Food Chemistry: Molecular Sciences |
| Subjects: | |
| Online Access: | http://www.sciencedirect.com/science/article/pii/S2666566223000229 |
| _version_ | 1797389415654359040 |
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| author | Steffen Pallarz Stefan Fiedler Daniela Wahler Jörn Lämke Lutz Grohmann |
| author_facet | Steffen Pallarz Stefan Fiedler Daniela Wahler Jörn Lämke Lutz Grohmann |
| author_sort | Steffen Pallarz |
| collection | DOAJ |
| description | Next-generation-sequencing (NGS) becomes increasingly important for laboratories tasked with the detection of genetically modified organisms (GMOs) in food, feed and seeds. Its implementation into standardized workflows demands reliable intra- and inter-laboratory reproducibility. Here, we analyze the reproducibility of short- and long-read targeted NGS and long-read whole genome sequencing (WGS) data between three independent laboratories. Replicate samples were submitted for sequencing and comparatively analyzed. The targeted-NGS-samples consisted of oil seed rape (OSR) sampled from a commodity shipment spiked with a genome edited (GE) OSR and the WGS-samples consisted of leaf material from the GMOs’ parental line. All laboratories delivered highly reproducible high-quality targeted NGS data with little variation. The detection of GMO-related sequences works well regardless of the facility, while the mapping to the complex genome is superior using long read data. Long read WGS is currently not suitable for routine use in enforcement laboratories, due to a large inter-laboratory variation. |
| first_indexed | 2024-03-08T22:55:31Z |
| format | Article |
| id | doaj.art-72e55a17377a41f3931983cbb1fb9af0 |
| institution | Directory Open Access Journal |
| issn | 2666-5662 |
| language | English |
| last_indexed | 2024-03-08T22:55:31Z |
| publishDate | 2023-12-01 |
| publisher | Elsevier |
| record_format | Article |
| series | Food Chemistry: Molecular Sciences |
| spelling | doaj.art-72e55a17377a41f3931983cbb1fb9af02023-12-16T06:09:43ZengElsevierFood Chemistry: Molecular Sciences2666-56622023-12-017100182Reproducibility of next-generation-sequencing-based analysis of a CRISPR/Cas9 genome edited oil seed rapeSteffen Pallarz0Stefan Fiedler1Daniela Wahler2Jörn Lämke3Lutz Grohmann4Department Genetic Engineering and Other Biotechnological Processes, Federal Office of Consumer Protection and Food Safety (BVL), P.O. Box 110260, 10832 Berlin, Germany; Corresponding author.Department Method Standardisation, Reference Laboratories, Resistance To Antibiotics, Federal Office of Consumer Protection and Food Safety (BVL), P.O. Box 110260, 10832 Berlin, GermanyDepartment Genetic Engineering and Other Biotechnological Processes, Federal Office of Consumer Protection and Food Safety (BVL), P.O. Box 110260, 10832 Berlin, GermanyDepartment Method Standardisation, Reference Laboratories, Resistance To Antibiotics, Federal Office of Consumer Protection and Food Safety (BVL), P.O. Box 110260, 10832 Berlin, GermanyDepartment Genetic Engineering and Other Biotechnological Processes, Federal Office of Consumer Protection and Food Safety (BVL), P.O. Box 110260, 10832 Berlin, GermanyNext-generation-sequencing (NGS) becomes increasingly important for laboratories tasked with the detection of genetically modified organisms (GMOs) in food, feed and seeds. Its implementation into standardized workflows demands reliable intra- and inter-laboratory reproducibility. Here, we analyze the reproducibility of short- and long-read targeted NGS and long-read whole genome sequencing (WGS) data between three independent laboratories. Replicate samples were submitted for sequencing and comparatively analyzed. The targeted-NGS-samples consisted of oil seed rape (OSR) sampled from a commodity shipment spiked with a genome edited (GE) OSR and the WGS-samples consisted of leaf material from the GMOs’ parental line. All laboratories delivered highly reproducible high-quality targeted NGS data with little variation. The detection of GMO-related sequences works well regardless of the facility, while the mapping to the complex genome is superior using long read data. Long read WGS is currently not suitable for routine use in enforcement laboratories, due to a large inter-laboratory variation.http://www.sciencedirect.com/science/article/pii/S2666566223000229NGS reproducibilityNGS data qualityNGT (New Genomic Techniques)GE (Gene Editing)GMO (Genetically Modified Organism)Detection |
| spellingShingle | Steffen Pallarz Stefan Fiedler Daniela Wahler Jörn Lämke Lutz Grohmann Reproducibility of next-generation-sequencing-based analysis of a CRISPR/Cas9 genome edited oil seed rape Food Chemistry: Molecular Sciences NGS reproducibility NGS data quality NGT (New Genomic Techniques) GE (Gene Editing) GMO (Genetically Modified Organism) Detection |
| title | Reproducibility of next-generation-sequencing-based analysis of a CRISPR/Cas9 genome edited oil seed rape |
| title_full | Reproducibility of next-generation-sequencing-based analysis of a CRISPR/Cas9 genome edited oil seed rape |
| title_fullStr | Reproducibility of next-generation-sequencing-based analysis of a CRISPR/Cas9 genome edited oil seed rape |
| title_full_unstemmed | Reproducibility of next-generation-sequencing-based analysis of a CRISPR/Cas9 genome edited oil seed rape |
| title_short | Reproducibility of next-generation-sequencing-based analysis of a CRISPR/Cas9 genome edited oil seed rape |
| title_sort | reproducibility of next generation sequencing based analysis of a crispr cas9 genome edited oil seed rape |
| topic | NGS reproducibility NGS data quality NGT (New Genomic Techniques) GE (Gene Editing) GMO (Genetically Modified Organism) Detection |
| url | http://www.sciencedirect.com/science/article/pii/S2666566223000229 |
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