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Multiple strand displacement amplification of DNA isolated from human archival plasma/serum: Identification of cytokine polymorphism by pyrosequencing analysis
Linköping University, Faculty of Health Sciences. Linköping University, Department of Biomedicine and Surgery, Division of surgery.
Linköping University, Faculty of Health Sciences. Linköping University, Department of Biomedicine and Surgery, Division of cell biology. Östergötlands Läns Landsting, Centre for Laboratory Medicine, Department of Molecular Biological Techniques.
Linköping University, Faculty of Health Sciences. Linköping University, Department of Biomedicine and Surgery, Division of surgery. Östergötlands Läns Landsting, Centre of Surgery and Oncology, Department of Surgery in Östergötland.
2007 (English)In: Clinica Chimica Acta, ISSN 0009-8981, Vol. 377, no 1-2, 108-113 p.Article in journal (Refereed) Published
Abstract [en]

Background: DNA isolation from formalin-fixed paraffin-embedded tissue appears to be problematic due to degradation caused by fixative. Our aim was to investigate if the isolated genomic DNA from archival plasma/serum, combined with multiple strand displacement amplification (MDA) can be used for genotyping. Methods: Nine archival plasma/serum samples and freshly frozen gastric biopsies from the same nine H. pylori-infected subjects were used for DNA isolation. Subsequently, MDA-DNA derived from the plasma/serum samples and DNA isolated from the antrum biopsies were analyzed by PCR amplification and pyrosequencing for the presence of interleukin-1beta gene (IL-1B) single nucleotide polymorphism (SNP). In addition, Southern blot and pyrosequencing analysis of H. pylori-specific PCR amplicons were performed. Results: IL-1B SNP profiles obtained from the plasma/serum MDA-DNA and antrum biopsy DNA were identical. A C/C genotype was observed in 7 of 9 samples, and 2 of 9 revealed a C/T genotype for IL-1B - 511. Similarly, 7 of 9 had a T/T, and 2 of 9 had a C/T genotype for IL-1B - 31, 4 of 9 had a C/C, 4 of 9 had a C/T, and 1 of 9 had a T/T genotype, respectively, for IL-1B + 3954. Moreover, pyrosequencing analysis revealed the presence of H. pylori 26695 and J99-like 16S rDNA variable V3 region sequence motifs in the antrum biopsies but not in the plasma/serum samples. Conclusions: We conclude that MDA combined with pyrosequencing enables a rapid and accurate molecular typing of cytokine single nucleotide polymorphisms from archival plasma/serum samples. © 2006 Elsevier B.V. All rights reserved.

Place, publisher, year, edition, pages
2007. Vol. 377, no 1-2, 108-113 p.
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Medical and Health Sciences
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URN: urn:nbn:se:liu:diva-39211DOI: 10.1016/j.cca.2006.09.003Local ID: 47286OAI: oai:DiVA.org:liu-39211DiVA: diva2:260060
Available from: 2009-10-10 Created: 2009-10-10 Last updated: 2011-01-11

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Sun, Yi-QianMonstein, Hans-JurgBorch, Kurt

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Faculty of Health SciencesDivision of surgeryDivision of cell biologyDepartment of Molecular Biological TechniquesDepartment of Surgery in Östergötland
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Clinica Chimica Acta
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