Why denature of DNA is necessary during isolation of DNA?

Question

by denaturing DNA it split into single stranded.say in plasmid isolation before getting the plasmid all the DNA including the plasmid should be denatured.is it not possible to get the plasmid without denaturing the DNA?

Answer 1

The reason that DNA is denatured during isolation is that typically, PCR (polymerase chain reaction) will be used to replicate the desired strand.  To understand why it is denatured, here is a step-by-step process of PCR:

1. heat targeted sequence of DNA to separate stands

2. cool to allow primers to bind

3. DNA polymerase extends the 3' end of each primer

This continues on and on with the number of identical strands of DNA doubling with each 'cycle'.  The reason that DNA is replicated using PCR is that when it is being isolated, it is typically being isolated to splice into another organism or to perfom electrophoresis (used for DNA comparison, i.e. crime scene).  For both of those, the amount of DNA required is typically more than a single strand--thus the denaturing portion of PCR only serves to further the process towards completion.