How to Show the Real Microbial Biodiversity? A Comparison of Seven DNA Extraction Methods for Bacterial Population Analyses in Matrices Containing Highly Charged Natural Nanoparticles

Abstract

A DNA extraction that comprises the DNA of all available taxa in an ecosystem is an essential step in population analysis, especially for next generation sequencing applications. Many nanoparticles as well as naturally occurring clay minerals contain charged surfaces or edges that capture negatively charged DNA molecules after cell lysis within DNA extraction. Depending on the methodology of DNA extraction, this phenomenon causes a shift in detection of microbial taxa in ecosystems and a possible misinterpretation of microbial interactions. With the aim to describe microbial interactions and the bio-geo-chemical reactions during a clay alteration experiment, several methods for the detection of a high number of microbial taxa were examined in this study. Altogether, 13 different methods of commercially available DNA extraction kits provided by seven companies as well as the classical phenol-chloroform DNA extraction were compared. The amount and the quality of nucleic acid extracts were determined and compared to the amplifiable amount of DNA. The 16S rRNA gene fragments of several taxa were separated using denaturing gradient gel electrophoresis (DGGE) to determine the number of different species and sequenced to get the information about what kind of species the microbial population consists of. A total number of 13 species was detected in the system. Up to nine taxa could be detected with commercially available DNA extraction kits while phenol-chloroform extraction lead to three detected species. In this paper, we describe how to combine several DNA extraction methods for the investigation of microbial community structures in clay.

Evaluation of V2 18S rDNA barcode marker and assessment of sample collection and DNA extraction methods for metabarcoding of autotrophic euglenids

“…Water DNA Kit (Omega Bio‐tek; WAT(O)). The selected methods are commonly used in metabarcoding surveys (Kaden and Krolla‐Sidenstein, 2015 ; Braun and Szewzyk, 2016 ; Wang et al ., 2018 ; Gołębiewski and Tretyn, 2019 ; Jeunen et al ., 2019 ). All isolation steps were performed according to the manufacturer's instructions.…”

“…presence of amplification inhibitors, such as humic acids), (iv) cost of reagents, (v) preparation complexity and time required to process a single sample. (Kaden and Krolla-Sidenstein, 2015;Braun and Szewzyk, 2016;Wang et al, 2018;Gołębiewski and Tretyn, 2019;Jeunen et al, 2019). All isolation steps were performed according to the manufacturer's instructions.…”

Sustainable farming genes: spatial distribution and influencing factors of Chinese Agricultural Heritage Sites (CAHSs)

“…To understand the impact of soil attributes on the CAHSs distribution visually, we select the components of clay and sand The spatial distribution of 7 CAHS types. (Kaden and Krolla-Sidenstein, 2015), and draw a histogram of CAHSs. Statistics show that 87.22 and 92.67% of CAHSs are distributed in areas with 16-48% clay and 16-64% sand, respectively.…”