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Internal Validation of the Investigator 26Plex QS Amplification Kit: a high-throughput multiplex assay for reference and low copy number DNA samples

2024-09Other Restricted access
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dc.contributor.authorCardoso, Paula
dc.contributor.authorSerra, Armando
dc.contributor.authorBogas, Vanessa
dc.contributor.authorBalsa, Filipa
dc.contributor.authorLopes, Virginia
dc.contributor.authorDario, Rita
dc.contributor.authorPorto, Maria JoĆ£o
dc.contributor.authorAmorim, AntĆ³nio
dc.contributor.authorCorte Real, F.
dc.contributor.authorBrito, Pedro
dc.date.accessioned2024-09-25T15:34:48Z
dc.date.available2024-09-25T15:34:48Z
dc.date.issued2024-09
dc.descriptionPoster apresentado no 30th Congress of the International Society for Forensic(2024), Santiago de Compostela (Espanha)pt_PT
dc.description.abstractThe InvestigatorĀ® 26plex QS Amplification Kit, from QIAGEN, provides reliable and rapid DNA profiles while enabling the multiplex amplification of 24 STRs, 2 Quality Sensors and a gender-determining marker, Amelogenin. The Quality Sensors incorporated in this kit provide insight into the sample quality and the PCR's success while alerting to the presence of inhibitors. Through an extensive internal validation study, this research sought to implement the InvestigatorĀ® 26plex QS in the laboratory routine of the Forensic Genetic and Biology Service, Central branch (SGBF-C) of the Portuguese National Institute of Legal Medicine and Forensic Sciences. Here we detail the procedures and parameters applied which followed the SWGDAM guidelines, as well as report the results obtained throughout. Firstly, it was crucial to establish unique analytical criteria that would be the baseline for the interpretation of the results obtained. To accomplish such, analytical, stochastic, heterozygous balance and stutter thresholds were defined. Thereupon, this study intended to gauge the kitā€™s performance, by evaluating its concordance with normalized methodologies while assessing its sensitivity, specificity, robustness, and precision, besides its efficiency in the presence of degraded and/or inhibited samples. Lastly, in favour of optimizing the laboratory workflow, an assay was carried out to test half volume reactions and direct amplification on reference samples. In this study a wide range of sample types were analysed, which made it possible to acquire robust data pertaining to the performance of the assay. The laboratory methodology applied comprehended: DNA extraction using Prep-n-Goā„¢ Buffer and PrepFiler Expressā„¢ Forensic DNA Extraction Kit, quantification with the Quantifilerā„¢ Trio Quantification Kit, followed by the amplification with the InvestigatorĀ® 26plex QS. Capillary electrophoresis was performed in the Applied Biosystemsā„¢ 3500 Genetic Analyzer, and the electropherogramsā€™ were analysed using the GeneMapperā„¢ ID-X Software v1.6. Through this work, it was possible to characterize the main advantages of the InvestigatorĀ® 26plex QS kit, as well as its limitations. The validation data demonstrated that this system produces reliable profiles in the presence of minute DNA quantities and identifies the minor contributor in a mixture up to a 1:50 ratio. The results inferred a high human specificity, robustness, and sensitivity, while producing concordant and reproducible results with optimized protocols for both reference and low copy number DNA samples. Through concordance studies it was further shown that there is a high consistency between this novel amplification kit and those currently implemented in the SGBF-C, thus proving its suitability for the analysis of forensic samples.pt_PT
dc.description.versionN/Apt_PT
dc.identifier.urihttp://hdl.handle.net/10400.26/52201
dc.language.isoengpt_PT
dc.peerreviewedyespt_PT
dc.subjectInvestigatorĀ® 26plex QSpt_PT
dc.subjectForensic Geneticspt_PT
dc.subjectPCRpt_PT
dc.subjectInternal Validationpt_PT
dc.subjectAnalysis Thresholdspt_PT
dc.subjectForensic Samplespt_PT
dc.subjectSTRspt_PT
dc.titleInternal Validation of the Investigator 26Plex QS Amplification Kit: a high-throughput multiplex assay for reference and low copy number DNA samplespt_PT
dc.typeother
dspace.entity.typePublication
oaire.citation.conferencePlaceSantiago de Compostela, Espanhapt_PT
oaire.citation.title30th Congress of the International Society for Forensic Geneticspt_PT
person.familyNameNeves Cardoso
person.familyNameSerra
person.familyNameBogas
person.familyNameLopes
person.familyNameOliveira
person.familyNameAmorim
person.familyNameCorte Real GonƧalves
person.familyNameBrito
person.givenNamePaula Liliana
person.givenNameArmando
person.givenNameVanessa
person.givenNameVirginia
person.givenNameAna Rita
person.givenNameAntĆ³nio
person.givenNameFrancisco
person.givenNamePedro
person.identifier.ciencia-id4814-A310-A09E
person.identifier.ciencia-idD71C-9C88-4E0E
person.identifier.ciencia-idF317-39DF-EC9B
person.identifier.ciencia-id0B18-918A-80ED
person.identifier.orcid0000-0003-0361-9768
person.identifier.orcid0000-0003-0162-9305
person.identifier.orcid0000-0001-7929-5814
person.identifier.orcid0000-0002-7915-826X
person.identifier.orcid0000-0003-1202-7748
person.identifier.orcid0000-0002-7506-4426
person.identifier.orcid0000-0003-1495-9362
person.identifier.orcid0000-0002-9622-8303
person.identifier.ridE-5476-2013
person.identifier.ridA-4414-2013
person.identifier.scopus-author-id35076952400
person.identifier.scopus-author-id54785934400
person.identifier.scopus-author-id35253426400
person.identifier.scopus-author-id6603582554
rcaap.rightsrestrictedAccesspt_PT
rcaap.typeotherpt_PT
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