A systematic approach to improve downstream single‐cell analysis for the DEPArray™ technology
Article Swipe
Janine Schulte
,
Michael A. Marciano
,
Eva Scheurer
,
Iris Schulz
·
YOU?
·
· 2023
· Open Access
·
· DOI: https://doi.org/10.1111/1556-4029.15344
YOU?
·
· 2023
· Open Access
·
· DOI: https://doi.org/10.1111/1556-4029.15344
Most commercially available STR amplification kits have never been fully validated for low template DNA analysis, highlighting the need for testing different PCR kits and conditions for improving single‐cell profiling. Here, current strategies rely mainly on adjusting PCR cycle number and analytical threshold settings, with a strong preference for using 30 amplification cycles and thresholds at 30–150 RFU for allele detection. This study aimed to (1) determine appropriate conditions for obtaining informative profiles utilizing a dilution series, and (2) test the outcome on single cells using the DEPArray™ technology. Four routinely applied forensic STR kits were compared by using three different amplification volumes and DNA dilutions down to 3.0 pg, while two well‐performing kits were used for single/pooled leucocyte and sperm cell genotyping. Besides reduced costs, the results demonstrate that a 50%–75% PCR volume reduction was beneficial for peak height evaluation. However, this was counteracted by an increased artifact generation in diluted DNA volumes. Regarding profile completeness, the advantage of volume reduction was only prominent in samples processed with Fusion 6C. For single and pooled cells, ESIFast and NGMDetect provided a solid basis for consensus profiling regarding locus failure, although locus dropouts were generally observed as stochastic events. Amplification volume of 12.5 μL was confirmed as appropriate in terms of peak heights and stutter frequencies, with increased stutter peaks being the main artifact in single‐cell profiles. Limitations associated with these analyses are discussed, providing a solid foundation for further studies on low template DNA.
Related Topics
Concepts
Genotyping
DNA profiling
Computational biology
Polymerase chain reaction
Microsatellite
Biology
Computer science
Molecular biology
DNA
Allele
Genetics
Genotype
Gene
Metadata
- Type
- article
- Language
- en
- Landing Page
- https://doi.org/10.1111/1556-4029.15344
- https://onlinelibrary.wiley.com/doi/pdfdirect/10.1111/1556-4029.15344
- OA Status
- hybrid
- Cited By
- 10
- References
- 55
- Related Works
- 10
- OpenAlex ID
- https://openalex.org/W4385295432
All OpenAlex metadata
Raw OpenAlex JSON
- OpenAlex ID
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https://openalex.org/W4385295432Canonical identifier for this work in OpenAlex
- DOI
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https://doi.org/10.1111/1556-4029.15344Digital Object Identifier
- Title
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A systematic approach to improve downstream single‐cell analysis for the DEPArray
™ technologyWork title - Type
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articleOpenAlex work type
- Language
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enPrimary language
- Publication year
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2023Year of publication
- Publication date
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2023-07-27Full publication date if available
- Authors
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Janine Schulte, Michael A. Marciano, Eva Scheurer, Iris SchulzList of authors in order
- Landing page
-
https://doi.org/10.1111/1556-4029.15344Publisher landing page
- PDF URL
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https://onlinelibrary.wiley.com/doi/pdfdirect/10.1111/1556-4029.15344Direct link to full text PDF
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YesWhether a free full text is available
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hybridOpen access status per OpenAlex
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https://onlinelibrary.wiley.com/doi/pdfdirect/10.1111/1556-4029.15344Direct OA link when available
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Genotyping, DNA profiling, Computational biology, Polymerase chain reaction, Microsatellite, Biology, Computer science, Molecular biology, DNA, Allele, Genetics, Genotype, GeneTop concepts (fields/topics) attached by OpenAlex
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10Total citation count in OpenAlex
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2025: 5, 2024: 3, 2023: 2Per-year citation counts (last 5 years)
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10Other works algorithmically related by OpenAlex
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