Optimized DNA Extraction from Gut Samples

DNA was isolated from cecal contents and fecal pellets using the QIAamp DNA stool mini kit (Qiagen) as previously described [56 (link)]. For enhancing the pathogen’s DNA extraction we made an improvement in the original protocol: a vigorous homogenization of the samples with 300 mg of 1.0 mm zirconia beads (BioSpec, Bartlesville, OK, USA) using a MiniBeadBeater (BioSpec, Bartlesville, OK, USA). After extraction, DNA was eluted in 200μl Elution Buffer and stored at −20°C.

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Giallourou N., Medlock G.L., Bolick D.T., Medeiros P.H., Ledwaba S.E., Kolling G.L., Tung K., Guerry P., Swann J.R, & Guerrant R.L. (2018). A novel mouse model of Campylobacter jejuni enteropathy and diarrhea. PLoS Pathogens, 14(3), e1007083.

Publication 2018

QIAamp DNA stool mini kit 1.0 mm zirconia beads MiniBeadBeater

Buffer Cecal Pathogen Pellets Stool Zirconia

Corresponding Organization : University of Virginia

Other organizations : Imperial College London, Universidade Federal do Ceará, University of Venda, Naval Medical Research Command

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Variable analysis

independent variables

Homogenization of the samples with 300 mg of 1.0 mm zirconia beads using a MiniBeadBeater

dependent variables

DNA extraction efficiency

control variables

Use of QIAamp DNA stool mini kit (Qiagen) for DNA isolation from cecal contents and fecal pellets as previously described
Elution of DNA in 200μl Elution Buffer and storage at −20°C

controls

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

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