FAQ about AquaPlasmid
1. Do I need to store the AquaPlasmid kit at 4 °C?
No, AquaPlasmid solution should be stored at room temperature (~22 °C). If the temperature is below 18 °C, precipitates may develop. If so, incubate the solution at 37-55 °C for a few minutes and vortex to re-solubilize it before use.
2. How does AquaPlasmid work?
AquaPlasmid combines the functions of traditional P1 buffer (cell suspension), P2 buffer (cell lysis), N3 buffer (debris removal), and silica column (plasmid DNA purification) in a single solution. It lyses the cells without denaturing the DNA, extracts the plasmid DNA while keeping other cellular contaminants in the cell debris. The plasmid DNA is subsequently precipitated from the clear lysate with isopropanol.
3. Does AquaPlasmid contain guanidine salts?
No, AquaPlasmid does not contain guanidine salts that are required for column-based purification. Guanidine salts are not biodegradable. When mixed with Bleach, they can release toxic and mutagenic fumes into the environment, which may be harmful to lab workers and aquatic lives.
4. How should I scale up and down the reagent for other culture volumes?
We recommend using 200 ul of AquaPlasmid for each 1.5 ml of overnight bacterial culture. However, if the cell density is too high, it may result in poor cell lysis, incomplete debris removal and decreased DNA yield. An indication of cell density exceeding the capacity of AquaPlasmid is a large DNA pellet following isopropanol precipitation. In that case, you should use 300 ul of AquaPlasmid for each 1.5 ml of bacterial culture or use 200 ul of AquaPlasmid for each ml of culture.
5. Can I use AquaPlasmid to extract plasmid DNA from endA1 strains?
We recommend AquaPlasmid only for plasmid extraction from E. coli strains carrying the endA1 mutation, such as TOP10, DH5a, XL-1 Blue, JM109, and SURE, etc. If the genotype is unknown, you should incubate an aliquot of the purified DNA in a restriction enzyme buffer at 37 °C for 12 hours to confirm there is no DNA degradation. If the DNA is degraded, you may try incubating the purified DNA at 75-85 °C for 30 min to inactivate the contaminating nucleases.