Rapid and Efficient Method for Environmental Dna Extraction and Purification From Soil

Abstract

Large proportion of microbial population in the world is unculturable. Extraction of total DNA from soil is usually a crucial step considering to the difficulties of study the uncultivable microorganisms. Humic acid is considered as the main inhibitory agent in the environmental DNA studies. Here, we introduced a rapid and efficient method for DNA extraction and purification from soil. Yield of DNA extraction by the presented method was 130 ng/µl. Three conventional methods of DNA extraction including liquid nitrogen incursion, bead beating and sonication were performed as control methods. Yield of DNA extraction by these methods were 110, 90 and 50 ng/µl, respectively. A rapid and efficient one step DNA purification method was introduced instead of hazardous conventional phenol-chloroform methods. Humic acid removal percentage by the introduced method was 95.8 % that is comparable with 97 % gained by the conventional gel extraction method and yield of DNA after purification was 84 % and 73 %, respectively. This study could be useful in molecular ecology and metagenomics study as a fast and reliable method.

Evaluation of Molecular-based Methods for the Detection and Quantification ofCryptosporidiumspp. in Wastewater

“…Incorporating a bead-beating step prior to the freeze-thaw cycles did result in improved DNA yields (p < 0.05). This is in alignment with other studies in which these two pretreatments demonstrated more satisfactory results for DNA extraction from Cryptosporidium oocysts and other protozoa compared to solely using freeze-thaw pretreatment (Babaei et al, 2011; Hamedi et al, 2016; Mthethwa et al, 2022). However, the DNA yields from combined pretreatments were still lower than those from bead-beating alone, indicating significant DNA loss due to degradation during the freeze-thaw process.…”

A low‐cost pipeline for soil microbiome profiling

“…Custom gDNA extraction workflows have been described, but many current methods are low in extraction yield, throughput and often not tested for NGS or microbiome purposes or optimized for soil (Abdel‐Latif & Osman, 2017; Zou et al, 2017). Commonly used protocols for nucleic acid purification are often column and centrifuge based, which are more laborious, harder to automate and so not easily used in a high‐throughput manner or scaled economically (Hamedi et al, 2016; Miao et al, 2014; Narayan et al, 2016; Oberacker et al, 2019). More recent open‐source protocols for rapid DNA purification using coated magnetic particles (Oberacker et al, 2019) or even cellulose (Zou et al, 2017) address many of these shortcomings.…”

New horizons in developing cell lysis methods: A review

“…Sometimes, some other compounds such as lignin are also exist in the cell wall structure (Somerville et al, 2004). Lignin removal is one of the major challenges in the procedure of DNA extraction from plant or environmental samples (Hamedi et al, 2021). In archaea, the cell wall is mainly composed of pseudo-peptidoglycan, S-layer, or polysaccharides.…”

“…These lysis approaches require more powerful tools to achieve cell disruption and subsequently extraction of DNA. In these circumstances, a combination of the lysis methods are used (Hamedi et al, 2021). Common examples of this approach are metagenomic studies of soil (Nesme et al, 2016), plant roots (Soni et al, 2017), mammalians gut (Maccaferri et al, 2011), and feces (Srivathsan et al, 2016).…”