Conditional touchdown multiplex polymerase chain reaction

A high-throughput and cost-effective method for simultaneous amplification of target DNA sequences with high fidelity and workable rate is achieved by a two-stage amplification incorporating multiplex PCR with conditional touchdown strategies. This improved multiplex PCR comprises a simultaneous PCR and a specific PCR, and either one or both of the amplification steps are performed with a touchdown strategy, of which loose touchdown strategy is applied with a temperature lower than the optimized annealing temperature, and stringent touchdown strategy is applied with a temperature higher than the optimized annealing temperature.