Q1：Why is plasmid DNA yield low?
A1：
1) Plasmid copy number is low, plasmid >10 kb or gram-positive bacterial plasmid. The amount of bacteria used should be increased, 300~500 mL of overnight culture can be used, and the eluent Buffer EB should be preheated in a 60°C water bath, and the adsorption and elution time can be appropriately extended to increase the extraction efficiency;
2) The problem of bacteria species. There is a loss of plasmids during the storage process of the strains. Before culturing the bacteria, it is recommended to streak and activate to stabilize the yield;
3) The bacteria are not fully lysed. Bacteria must be fully resuspended in Buffer I/RNase A or the bacterial cells should not be too large to avoid clumping or excessive bacteria that cannot be lysed and reduce the yield.
Q2：Why is there genomic DNA contamination in plasmid DNA?
A2：
1) The bacterial culture time is too long. The culture time of bacterial liquid should be controlled within 12-16 h;
2) The cracking problem. When adding Buffer II, it must be mixed by gentle inversion; when processing multiple samples, the total time should not exceed 5 min from the time of adding Buffer II.
