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Clinical Chemistry and Laboratory Medicine (CCLM)

Published in Association with the European Federation of Clinical Chemistry and Laboratory Medicine (EFLM)

Editor-in-Chief: Plebani, Mario

Ed. by Gillery, Philippe / Greaves, Ronda / Lackner, Karl J. / Lippi, Giuseppe / Melichar, Bohuslav / Payne, Deborah A. / Schlattmann, Peter


IMPACT FACTOR 2018: 3.638

CiteScore 2018: 2.44

SCImago Journal Rank (SJR) 2018: 1.191
Source Normalized Impact per Paper (SNIP) 2018: 1.205

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1437-4331
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Volume 52, Issue 11

Issues

Selection of an optimal method for co-isolation of circulating DNA and miRNA from the plasma of pregnant women

Tatiana Sedlackova
  • Corresponding author
  • Institute of Molecular BioMedicine, Faculty of Medicine, Comenius University in Bratislava, Bratislava, Slovakia
  • Email
  • Other articles by this author:
  • De Gruyter OnlineGoogle Scholar
/ Gabriela Repiska
  • Institute of Molecular BioMedicine, Faculty of Medicine, Comenius University in Bratislava, Bratislava, Slovakia
  • Other articles by this author:
  • De Gruyter OnlineGoogle Scholar
/ Gabriel Minarik
  • Institute of Molecular BioMedicine, Faculty of Medicine, Comenius University in Bratislava, Bratislava, Slovakia
  • Department of Molecular Biology, Faculty of Natural Sciences, Comenius University in Bratislava, Bratislava, Slovakia
  • Geneton Ltd., Bratislava, Slovakia
  • Other articles by this author:
  • De Gruyter OnlineGoogle Scholar
Published Online: 2014-05-09 | DOI: https://doi.org/10.1515/cclm-2014-0021

Abstract

Background: Circulating nucleic acids acquired non-invasively have been confirmed as useful biomarkers in cancer and prenatal medicine. The most important molecules in the field of circulating nucleic acids research are circulating DNA and miRNA. In this study, the possibility of co-isolation of total circulating DNA, cell-free fetal DNA and miRNA from the plasma of pregnant women was tested, and the yields of co-isolated circulating nucleic acids using two commercial kits and three protocols were compared.

Methods: Cell-free fetal DNA and miRNA from the plasma of pregnant women carrying male fetuses were co-isolated with the miRCURY™ RNA Isolation Kit according to the original protocol and the QIAamp Circulating Nucleic Acid Kit (CNA kit) according to the manufacturer’s protocol for DNA isolation and miRNA isolation. For comparison of DNA isolation, the AR and DYS14 gene-based assays were used for the detection and quantification of total circulating and cell-free fetal DNA. For miRNA detection and quantification, the miR-16 and miR-451 assays were used.

Results: Two different protocols for isolation using the CNA kit did not significantly differ in the yields of isolated tcDNA and cffDNA; however, the amount of isolated cffDNA using the miRCURY™ RNA Isolation Kit was significantly less (p<0.05, F=4.776). There was a statistically significant difference for miRNA isolation (p<0.0001, F=859 for miR-16 and p<0.0001, F=854.4 for miR-451), with the highest amount of isolated miRNA obtained using the miRCURY™ RNA Isolation Kit.

Conclusions: All three methods used in our study were successful in the co-isolation of tcDNA, cffDNA and miRNA from the same sample. The best combined results were obtained with the miRCURY™ RNA Isolation Kit.

Keywords: cell free fetal DNA; cffDNA; circulating nucleic acids; miRNA; non-invasive testing; nucleic acids co-isolation; tcDNA

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About the article

Corresponding author: Tatiana Sedlackova, Institute of Molecular BioMedicine, Faculty of Medicine, Comenius University in Bratislava, Bratislava, Slovakia, Phone: +421 2 59357371, Fax: +421 2 59357631, E-mail:


Received: 2014-01-06

Accepted: 2014-04-16

Published Online: 2014-05-09

Published in Print: 2014-11-19


Citation Information: Clinical Chemistry and Laboratory Medicine (CCLM), Volume 52, Issue 11, Pages 1543–1548, ISSN (Online) 1437-4331, ISSN (Print) 1434-6621, DOI: https://doi.org/10.1515/cclm-2014-0021.

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