Evaluation of DNA extraction kits and phylogenetic diversity of the porcine gastrointestinal tract based on Illumina sequencing of two hypervariable regions

Publication Type

Journal contribution (peer reviewed)

Authors

Burbach K, Seifert J, Pieper DH, Camarinha-Silva A

Year of publication

2016

Published in

MicrobiologyOpen

Band/Volume

5/1

DOI

10.1002/mbo3.312

Page (from - to)

70-82

Keywords

digesta, DNA Extraktion, feces, ileum, Illumina sequencing, Schwein, T-RFLP

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