title: Evaluation of DNA extraction kits and phylogenetic diversity of the porcine gastrointestinal tract based on Illumina sequencing of two hypervariable regions.
abstract: A robust DNA extraction method is important to identify the majority of microorganisms present in environmental microbial communities and to enable a consistent comparison between different studies. Here, 15 manual and four automated commercial DNA extraction kits were evaluated for their efficiency to extract DNA from porcine feces and ileal digesta samples. DNA yield, integrity, and purity varied among the different methods. Terminal restriction fragment length polymorphism (T-RFLP) and Illumina amplicon sequencing were used to characterize the diversity and composition of the microbial communities. We also compared phylogenetic profiles of two regions of the 16S rRNA gene, one of the most used region (V1-2) and the V5-6 region. A high correlation between community structures obtained by analyzing both regions was observed at genus and family level for ileum digesta and feces. Based on our findings, we want to recommend the FastDNA(™) SPIN Kit for Soil (MP Biomedical) as a suitable kit for the analyses of porcine gastrointestinal tract samples.
authors.affiliation: Institute of Animal Science, University of Hohenheim, Stuttgart, Germany.
authors.forename: Katharina
authors.initials: K
authors.lastname: Burbach
authors.affiliation: Institute of Animal Science, University of Hohenheim, Stuttgart, Germany.
authors.forename: Jana
authors.initials: J
authors.lastname: Seifert
authors.affiliation: Microbial Interactions and Processes Research Group, Helmholtz Centre for Infection Research, Braunschweig, Germany.
authors.forename: Dietmar H
authors.initials: DH
authors.lastname: Pieper
authors.affiliation: Institute of Animal Science, University of Hohenheim, Stuttgart, Germany.
authors.forename: Amélia
authors.initials: A
authors.lastname: Camarinha-Silva
mesh_terms: D000818:Animals; D001483:Base Sequence; D004269:DNA, Bacterial / Q000032:analysis / Q000235:genetics*; D005243:Feces / Q000382:microbiology*; D007082:Ileum / Q000382:microbiology*; D016133:Polymerase Chain Reaction / Q000379:methods*; D012150:Polymorphism, Restriction Fragment Length; D012336:RNA, Ribosomal, 16S / Q000235:genetics; D017422:Sequence Analysis, DNA / Q000379:methods*; D013552:Swine
chemical_list: D004269:DNA, Bacterial; D012336:RNA, Ribosomal, 16S
keywords: DNA extraction; Illumina amplicon sequencing; T-RFLP; feces; ileum digesta; porcine