pone.0161778.pdf 1,91MB
1000 Titel
  • Evaluation of Existing Methods for Human Blood mRNA Isolation and Analysis for Large Studies
1000 Autor/in
  1. Meyer, Anke |
  2. Paroni, Federico |
  3. Günther, Kathrin |
  4. Dharmadhikari, Gitanjali |
  5. Kelm, Sørge |
  6. Maedler, Kathrin |
  7. Ahrens, Wolfgang |
1000 Erscheinungsjahr 2016
1000 Art der Datei
1000 Publikationstyp
  1. Artikel |
1000 Online veröffentlicht
  • 2016-08-30
1000 Erschienen in
1000 Quellenangabe
  • 11(8): e0161778
1000 FRL-Sammlung
1000 Copyrightjahr
  • 2016
1000 Lizenz
1000 Verlagsversion
  • |
  • |
1000 Publikationsstatus
1000 Begutachtungsstatus
1000 Sprache der Publikation
1000 Abstract/Summary
  • AIMS: Prior to implementing gene expression analyses from blood to a larger cohort study, an evaluation to set up a reliable and reproducible method is mandatory but challenging due to the specific characteristics of the samples as well as their collection methods. In this pilot study we optimized a combination of blood sampling and RNA isolation methods and present reproducible gene expression results from human blood samples. METHODS: The established PAXgeneTM blood collection method (Qiagen) was compared with the more recent TempusTM collection and storing system. RNA from blood samples collected by both systems was extracted on columns with the corresponding Norgen and PAX RNA extraction Kits. RNA quantity and quality was compared photometrically, with Ribogreen and by Real-Time PCR analyses of various reference genes (PPIA, β-ACTIN and TUBULIN) and exemplary of SIGLEC-7. RESULTS: Combining different sampling methods and extraction kits caused strong variations in gene expression. The use of PAXgeneTM and TempusTM collection systems resulted in RNA of good quality and quantity for the respective RNA isolation system. No large inter-donor variations could be detected for both systems. However, it was not possible to extract sufficient RNA of good quality with the PAXgeneTM RNA extraction system from samples collected by TempusTM collection tubes. Comparing only the Norgen RNA extraction methods, RNA from blood collected either by the TempusTM or PAXgeneTM collection system delivered sufficient amount and quality of RNA, but the TempusTM collection delivered higher RNA concentration compared to the PAXTM collection system. The established Pre-analytix PAXgeneTM RNA extraction system together with the PAXgeneTM blood collection system showed lowest CT-values, i.e. highest RNA concentration of good quality. Expression levels of all tested genes were stable and reproducible. CONCLUSIONS: This study confirms that it is not possible to mix or change sampling or extraction strategies during the same study because of large variations of RNA yield and expression levels.
1000 Sacherschließung
lokal Polymerase chain reaction
lokal RNA extraction
lokal Centrifugation
lokal Reverse transcriptase-polymerase chain reaction
lokal Deoxyribonucleases
lokal Gene expression
lokal Blood
lokal RNA isolation
1000 Fachgruppe
  1. Biologie |
  2. Medizin |
1000 Fächerklassifikation (DDC)
1000 Liste der Beteiligten
1000 Förderer
  1. University of Bremen |
  2. European Research Council (ERC) |
  3. Federal Ministry of Education and Research (BMBF), Germany |
1000 Fördernummer
  1. -
  2. 260336-SIADIA
  3. -
1000 Förderprogramm
  1. ZF-funds
  2. -
  3. German Diabetes Center grant
1000 Dateien
1000 Förderung
  1. 1000 joinedFunding-child
    1000 Förderer University of Bremen |
    1000 Förderprogramm ZF-funds
    1000 Fördernummer -
  2. 1000 joinedFunding-child
    1000 Förderer European Research Council (ERC) |
    1000 Förderprogramm -
    1000 Fördernummer 260336-SIADIA
  3. 1000 joinedFunding-child
    1000 Förderer Federal Ministry of Education and Research (BMBF), Germany |
    1000 Förderprogramm German Diabetes Center grant
    1000 Fördernummer -
1000 Objektart article
1000 Beschrieben durch
1000 @id frl:6406231.rdf
1000 Erstellt am 2018-01-08T17:40:33.188+0100
1000 Erstellt von 218
1000 beschreibt frl:6406231
1000 Bearbeitet von 218
1000 Zuletzt bearbeitet Mon May 14 14:36:17 CEST 2018
1000 Objekt bearb. Mon May 14 14:36:17 CEST 2018
1000 Vgl. frl:6406231
1000 Oai Id
  1. |
1000 Sichtbarkeit Metadaten public
1000 Sichtbarkeit Daten public
1000 Gegenstand von

View source