Robert Page

Here’s a very cheap, rapid, accessible, and scalable DNA purification method suitable for viral RNA or bacterial DNA. It's been validated for use in SARS-CoV-2 (COVID-19) detection, and has more recently been used to purify DNA to allow the detection of toxin-producing E. coli in leafy greens. The method, by Page et al. (2022), was developed as a response to the global shortage of reagents such as guanidium thiocyanate, and presumably silica membrane spin columns, during the COVID-19 pandemic. Instead, their method is an adaption of the long-established glassmilk DNA extraction method, in which DNA is bound to a ground silica slurry in the presence of a chaotropic salt (for an example of an early application, see Boom et al., 1990 at the bottom of this post). Their method uses: ⭐4% SDS (sodium dodecyl sulfate) to inactivate viral particles ⭐1.25M NaCl, isopropanol, and glassmilk to bind the RNA ⭐Centrifugation to pellet the glassmilk, and pipetting to remove the supernatant ⭐70% ethanol to wash the glassmilk pellet, and pipetting to remove the supernatant ⭐Air-drying to evaporate residual ethanol ⭐Water to resuspend the RNA, followed by pipetting to remove it from the glass-milk solution and store in a fresh tube. During their tests to confirm that sodium chloride was effective as a binding salt, the authors found no significant differences in the results regardless of which chaotropic salt was used. They also found no significant differences between their protocol and one involving paramagnetic bead DNA purification. The use of sodium chloride as the binding reagent not only makes this method much more robust to supply chain issues, but also makes it much cheaper, much more accessible outside of research laboratories, much safer, and also more environmentally friendly. You can read the article here: Page et al. (2022). Homebrew: An economical and sensitive glassmilk-based nucleic-acid extraction method for SARS-CoV-2 diagnostics. Cell Reports Methods, 2(3). ncbi.nlm.nih.gov/pmc/article… Even better, since its publication, the above method has been modified and used successfully as part of an point-of-need onsite-detection PCR assay for the detection of Shiga toxin-producing Escherichia coli bacteria in leafy greens. In this method, by Costa-Ribeiro et al. (2024), there is an initial heat lysis step of 5 min at 99 °C but the rest of the process is very similar. It allowed them to detect less than 10 colony forming units of Shiga toxin-prodicing E. coli per 25 g of leafy greens using a LAMP assay. You can read this article here: Costa-Ribeiro et al. (2024). Moving towards on-site detection of Shiga toxin-producing Escherichia coli in ready-to-eat leafy greens. Current Research in Food Science, 100716. ncbi.nlm.nih.gov/pmc/article… And for an early example of glassmilk DNA extraction using guanidium thiocyanate rather than sodium chloride, you can read Boom et al. (1990) below: Boom et al. (1990). Rapid and simple method for purification of nucleic acids. Journal of clinical microbiology, 28(3), 495-503. journals.asm.org/doi/abs/10.… *** ncbi.nlm.nih.gov/pmc/article…