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Multiple displacement amplification of isolated DNA from human gallstones: Molecular identification of Helicobacter DNA by means of 16S rDNA-based pyrosequencing analysis
Linköping University, Department of Medicine and Care. Linköping University, Faculty of Health Sciences.
Linköping University, Department of Biomedicine and Surgery. Linköping University, Faculty of Health Sciences.
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.
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.
2005 (English)In: Helicobacter, ISSN 1083-4389, E-ISSN 1523-5378, Vol. 10, no 6, 592-600 p.Article in journal (Refereed) Published
Abstract [en]

Background. Molecular typing of Helicobacter spp. in clinical biopsy specimens has become increasingly important. By means of nested polymerase chain reaction (PCR) amplification and Southern blot analysis of the PCR amplicons, we have shown that Helicobacter spp. DNA is present in human gallstones. In this study we have investigated the possibility of using multiple displacement amplification (MDA) of isolated gallstone DNA and pyrosequencing analysis for the molecular identification of Helicobacter spp. Materials and Methods. DNA isolated from the nucleus of 33 human gallstones and one control strain were used in a MDA assay. Subsequently, pyrosequencing analysis was performed either directly on MDA-DNA using primers flanking the Helicobacter spp. 16S rDNA variable V3 region or on PCR amplicons derived from broad-range primers flanking the 16S rDNA variable V3, V4, and V9 regions. Results. Pyrosequencing analysis of 16S rDNA derived from MDA-DNA revealed that Helicobacter spp.-like DNA was present in 25 of 33 (approximately 76%) gallstones. Using an H. pylori-specific Southern blot analysis, Helicobacter spp.-like DNA was present in 20 of 33 [approximately 61%] of the gallstones. Using MDA-DNA directly in pyrosequencing analysis, Helicobacter spp.-like DNA was present in 13 of 33 [approximately 39%] gallstones. Conclusions. We conclude that multiple displacement amplification combined with pyrosequencing enables a rapid and accurate molecular typing of Helicobacter spp. from small and precious biopsy specimens. © 2005 The Authors Journal compilation © 2005 Blackwell Publishing Ltd.

Place, publisher, year, edition, pages
2005. Vol. 10, no 6, 592-600 p.
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Medical and Health Sciences
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URN: urn:nbn:se:liu:diva-30808DOI: 10.1111/j.1523-5378.2005.00361.xLocal ID: 16435OAI: oai:DiVA.org:liu-30808DiVA: diva2:251631
Available from: 2009-10-09 Created: 2009-10-09 Last updated: 2017-12-13

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Nilsson, IsabelleShabo, IvanSvanvik, JoarMonstein, Hans-Jurg

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Department of Medicine and CareFaculty of Health SciencesDepartment of Biomedicine and SurgeryDivision of surgeryDepartment of Surgery in ÖstergötlandDivision of cell biologyDepartment of Molecular Biological Techniques
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