Optimized Method for Efficient DNA Extraction from Agricultural Soils.

Soil harbors the highest concentration of microorganisms in ecosystems, and their molecular characterization through high-throughput sequencing is essential for metagenomic studies. However, obtaining high-quality, high-concentration DNA is limited by physicochemical properties (pH, heavy metals, humic acids) and adsorption to clay minerals. Although standardized commercial protocols exist, they present variable limitations depending on soil type. This study developed and validated the National Center for Genetic Resources-Microorganism Collection (CNRG-CM) method, which incorporates innovative pre-washing steps using phosphate-buffered saline (PBS) and sodium phosphate to effectively remove inhibitory humic acids and metal ions, combined with cetyltrimethylammonium bromide (CTAB)/chloroform extraction to achieve high-molecular-weight metagenomic DNA isolation. The CNRG-CM method was applied to three diverse soil types with variable physicochemical properties, recovering DNA concentrations ranging from 1000 to 1300 ng/μL ith a yield of 30 to 48 µg/g-1, significantly exceeding those obtained with a standard commercial kit with maximum DNA concentrations of 360 ng/μL and a yield of 43 µg/g-1. The CNRG-CM protocol is established as an effective and adaptable alternative for metagenomic DNA extraction across diverse agricultural and ecological contexts. It enables subsequent metagenomic studies of soil microbial communities.