How do you think scientist use resistance to antibiotics to identify bacteria which contain plasmids?
Hi, Ive been working with bacterial transformations lately, and I believe I can answer your question. I use T/A cloning methods from a kit, and will contain my answer within the context of my commercial kit (T/A cloning kit from invitrogen). The plasmid comes with the kit. However, the plasmid is linear, not circular. If a linear plasmid is incorporated into a bacterium, the plasmid will not be replicated, as only circularized plasmids get amplified. One important step in the cloning process is to re-circularize the linear plasmid, and of great interest is to re-circularize plasmid DNA that contains your insert DNA. This step is called a ligase reaction, and uses an enzyme (ligase) that will re-circularize the linear plasmid. It is possible to re-circularize two molecules in this step. Re-circularize the plasmid that does not contain your insert (a bad thing), and/or re-circularize the plasmid that is linear and DOES contain your insert (good thing). After you perform the ligation reac
