Stool DNA Extraction and 16S Sequencing

Total DNA was isolated from frozen stool samples using MagnaPure LC DNA Isolation Kit III (Bacteria, Fungi) (Roche, Mannheim, Germany) according to manufacturer’s instructions including mechanic and enzymatic lysis [22 (link)]. Hypervariable regions V1-V2 were amplified in a target specific PCR using the primers 27F and R357 (27F-AGAGTTTGATCCTGGCTCAG; R357-CTGCTGCCTYCCGTA) and sequenced with the Illumina MiSeq technique (Illumina, Eindhoven, The Netherlands) [22 (link)]. Sequencing was done in cooperation with the Core Facility for Molecular Biology at the Center for Medical Research in Graz.

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Stadlbauer V., Engertsberger L., Komarova I., Feldbacher N., Leber B., Pichler G., Fink N., Scarpatetti M., Schippinger W., Schmidt R, & Horvath A. (2020). Dysbiosis, gut barrier dysfunction and inflammation in dementia: a pilot study. BMC Geriatrics, 20, 248.

Publication 2020

MiSeq technique

Bacteria Enzymatic Frozen Fungi Isolation Mechanic Primers Stool

Corresponding Organization : Medical University of Graz

Other organizations : Albert Schweitzer Klinik, Center For Biomarker Research In Medicine

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Variable analysis

independent variables

Isolation of total DNA from frozen stool samples using MagnaPure LC DNA Isolation Kit III (Bacteria, Fungi) according to manufacturer's instructions including mechanic and enzymatic lysis

dependent variables

Sequencing of hypervariable regions V1-V2 using the primers 27F and R357 and the Illumina MiSeq technique

control variables

Frozen stool samples
MagnaPure LC DNA Isolation Kit III (Bacteria, Fungi)
Primers 27F and R357
Illumina MiSeq technique

controls

No positive or negative controls were explicitly mentioned in the provided information.

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