A method for high-concentration agarose gel preparation and its application in high-resolution separation of low-molecular-weight nucleic acids and proteins

Separation of nucleic acids and proteins using gels has always been a crucial part of molecular biology research. For low-molecular-weight nucleic acids and proteins, low- and medium-concentration agarose gels cannot achieve the high resolution as polyacrylamide gels. We found that 6 %-14 % high-concentration agarose gels (HAGs) could be easily dissolved in an autoclave and the vertical gel cast can be effortlessly filled using an easymade plastic box. Coupled with the improved buffer condition, HAG electrophoresis resulted in a good resolution of DNA and protein bands. With conventional TBE buffer plus 0.2 % NaCl, DNA fragments that differ by 2-5-bp within the 50-200-bp size range can be resolved on 6 %-8 % HAGs. By using TBE without NaCl, DNA fragments that differ by 2-bp or 2-nt within the 10-100-bp size range can be well resolved on >8 % HAGs. Using a buffer system comprising 1 M Tris-Cl for gel preparation, 0.2 M Tris-Cl/0.2 % SDS as upper tank buffer, and 0.2 M TrisCl as the lower tank buffer, HAGs achieved good molecular weight separation of total bacterial and plant proteins in the 10-200 kDa range. In conclusion, we developed a method for HAG preparation and electrophoresis of lowmolecular-weight nucleic acids and proteins.

Automated summary

Separation of nucleic acids and proteins is crucial in molecular biology research. This study found that high-concentration agarose gels can be easily dissolved in an autoclave and have good resolution for DNA and protein bands. Improved buffer conditions also contributed to the success of high-concentration agarose gel electrophoresis.