How to manufacture 1000kb of DNA ladder?
gsenviro | Certified Educator
A DNA ladder (or marker) is a control (or standard) commonly used in gel or acrylamide electrophoresis for measuring the size of unknown DNA fragments. It can be manufactured by a number of ways:
- by multiplex PCR: works by simultaneous amplification of DNA target using primer sets. Here, the touchdown PCR is combined with hot start PCR. This method has potential issues of reproducibility because optimal PCR conditions may not be established or maintained easily.
- PCR-synthesized markers: involves separate amplification of different DNA targets using specific primers. This technique is laborious, since we are using separate primer set for particular ladder fragment.
- combination of cloning, PCR and partial digestion: Here a sequence of a certain length can be generated, cloned (to get multimers), amplified by PCR and then partially digested by enzymes to generate ladder of required length.
Hope this helps.