Here are some tips for optimization when using the Monarch PCR and DNA Cleanup kit. The following are some things that you want to keep in mind for your starting sample.
1. We recommend that you start with a volume of at least 20 μl. If your sample is smaller than that you can increase the volume by adding TE buffer.
2. The recommended ratio of binding buffer to sample volume is dependent on the type and the size of the DNA that you are trying to clean up. Please follow the specific recommendations printed on the protocol card.
3. Please note that the maximum sample volume that can be loaded onto the column is 800 μl. Thus, if your volume after dilution with binding buffer exceeds 800 μl, you will need to load a portion of your sample, spin it, and then repeat until all of your sample is loaded and bound to the column.
Tips for optimizing quality of purified DNA
4. When removing the column from the collection tube after the final wash, make sure that the tip of the column doesn't touch the walls of the collection tube. Trace amounts of salt can contaminate your sample and if you observe that the tip of the column came into contact with the collection tube, then we'd recommend an additional 30 second centrifugation step prior to eluting.
5. Ensure that you add the elution buffer to the center of the columns matrix without puncturing it. The elution buffer needs to be in contact with the column matrix for efficient elution. Be sure to incubate the elution buffer in the column for a full minute before centrifugation.
6. For plasmids that are larger than 10 Kb heating the elution buffer to 50°C prior to adding it to the column can improve yields.
7. Please also note that sequential elutions may be performed for quantitatively higher recovery but will result in lower final DNA concentration.
Time saving tips
8. To save time during this protocol the binding and elution spin steps can be reduced from 1 minute to 30 seconds.
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