New visible and selective DNA staining method in gels with tetrazolium salts

Abstract

DNA staining in gels has historically been carried out using silver staining and fluorescent dyes like ethidium bromide and SYBR Green I (SGI). Using fluorescent dyes allows recovery of the analyte, but requires instruments such as a transilluminator or fluorimeter to visualize the DNA. Here we described a new and simple method that allows DNA visualization to the naked eye by generating a colored precipitate. It works by soaking the acrylamide or agarose DNA gel in SGI and nitro blue tetrazolium (NBT) solution that, when exposed to sunlight, produces a purple insoluble formazan precipitate that remains in the gel after exposure to light. A calibration curve made with a DNA standard established a detection limit of approximately 180 pg/band at 500 bp. Selectivity of this assay was determined using different biomolecules, demonstrating a high selectivity for DNA. Integrity and functionality of the DNA recovered from gels was determined by enzymatic cutting with a restriction enzyme and by transforming competent cells after the different staining methods, respectively. Our method showed the best performance among the dyes employed. Based on its specificity, low cost and its adequacy for field work, this new methodology has enormous potential benefits to research and industry.