Concentration of DNA for Transformation - (Oct/24/2009 )

Hi All,

What should be teh optimum concentration of plasmid DNA in order to have successful transformation and get well isolated colonies.
I have been facing problems in determining the amount of DNA needed for transformation and most of the time i end up with smear on the plates rather than well separated colonies so I then take some smear from that and re-plate them and so on....

If you are transforming miniprep plasmid, then 100 pg of plasmid will yield hundreds or thousands of transformants, assuming you have good transformation efficiency. Usually people plate two or more plates, with different amounts or dilutions of the transformed cells, using the one with the isolated colonies. Are you sure your antibiotic selection is working? Do you plate untranformed cells to check?