A DNA Melting Exercise for a Large Laboratory Class

Abstract

A simple and economical experimental setup is described that enables multiple individuals or groups within a laboratory class to measure the thermal melting of double stranded DNA simultaneously. The setup utilizes a basic spectrophotometer capable of measuring absorbance at 260 nm, UV plastic cuvettes, and a stirring hot plate. Students measure melting curves of short matched and mismatched DNA duplexes, and use quantitative analysis and first derivative curves to determine each duplex’s transition melting temperature, Tm. Student data are highly reproducible among groups and from year to year, and give distinct transition melting temperatures for the different duplexes. The protocol could easily be adapted to investigate the effects of other parameters on DNA melting and stability, such as changes in base composition, chain length, and buffer ionic strength.