Question

How can you determine denaturation in a DNA sample?

Answer 1

The denaturation of DNA can be measured by monitoring its absorbance of ultraviolet light at 260 nm. On denaturation, single stranded DNA absorbs around 20-30% more ultraviolet light than double stranded DNA. This occurs due to unstacking of the base pairs or hyperchromicity. Single stranded DNA is hyperchromic. Hence, it has a higher A260 values than double stranded DNA. By monitoring the absorbance of DNA at varying temperatures, the melting temperature can be determined. Melting temperature is the temperature at which absorbance increases. It indicates the denaturation point. It is dependent on the GC content of DNA. Higher the GC content, higher is the melting temperature.