Description: The simultaneous extraction of intracellular DNA (iDNA) and extracellular DNA (eDNA) can help to separate the living in situ community (represented by iDNA) from background DNA that originated both from past communities and from allochthonous sources. As iDNA and eDNA extraction protocols require separating cells from the sample matrix, their DNA yields are generally lower than direct methods that lyse the cells within the sample matrix. We, therefore, tested different buffers with and without adding a detergent mix (DM) in the extraction protocol to improve the recovery of iDNA from surface and subsurface samples that covered a variety of terrestrial environments. The combination of a highly concentrated sodium phosphate buffer plus DM significantly improved iDNA recovery for almost all tested samples. Additionally, the combination of sodium phosphate and EDTA improved iDNA recovery in most of the samples and even allowed the successful extraction of iDNA from extremely low-biomass iron-bearing rock samples taken from the deep biosphere. Based on our results, we recommend using a protocol with sodium phosphate in combination with either a DM (NaP 300 mM + DM) or EDTA (NaP + EDTA 300 mM). Furthermore, for studies that rely on the eDNA pool, we recommend using buffers solely based on sodium phosphate because the addition of EDTA or a DM resulted in a decrease in eDNA for most of the tested samples. These improvements can help reduce community bias in environmental studies and contribute to better characterizations of both modern and past ecosystems.

Description: This data publication presents quantitative DNA data obtained through fluorometric detection of genomic DNA and the estimation of 16S rRNA gene copies using quantitative Polymerase Chain Reaction (qPCR). The data encompasses various soil and rock samples collected across a climate gradient. The DNA was extracted using a protocol enabling the separate analysis of intracellular DNA (iDNA) and extracellular DNA (eDNA) from the same sample. The primary objective of this study was to enhance a previously established method developed by Alawi et al. (2014) for analyzing terrestrial samples by introducing modifications to the extraction buffer. Phosphate buffers at two different concentrations (120 mM and 300 mM), EDTA (300 mM), and a high-concentration phosphate buffer in combination with EDTA (300 mM each) were tested in conjunction with a detergent mix (detailed in Medina et al., 2023; submitted). Thorough tests, including spiked DNA experiments and cell counts, were conducted on one low biomass sample to validate the extraction setups. The two most effective extraction protocols were then applied to all samples from the four designated sites and compared with the phosphate buffer described by Alawi et al. (2014), resulting in the calculation of improvement factors. The resulting dataset provides valuable quantitative DNA information and estimates of 16S rRNA gene copies across diverse soil and rock samples along a climate gradient. The modifications made to the extraction buffer demonstrated improved efficiency in extracting especially iDNA compared to the original method. These findings contribute to the refinement and optimization of DNA extraction protocols for terrestrial samples, enabling more accurate and comprehensive analyses of microbial communities in different environments.