IGEM:British Columbia/2009/Notebook/Biosensor Sensitivity/2009/06/10

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Miniprep of RBS, J23100, RFP and YFP Biobricks

  • Used a Invitrogen ChargeSwitch Pro filter Plasmid miniprep kit
  • Followed the protocol given with the kit except for the following changes:
    • The precipitate that formed when adding the precipitation buffer clogged the mini column filter of YFP, J23100 and RBS. As a result the liquid solution was not able to completely pass through the filter
    • Repeated the centrifuge spin at 8500xg (10200 rpm on the Ambl Galaxy 16 centrifuge), still no change in liquid level
    • Used a pipette to try and resuspend the precipitate, and spun again at 8500 x g. Still no change in the liquid level
    • Pipetted the liquid out of the mini column and directly into the binding column and spun again at 8500 x g, all solution went through the filter. Proceeded to the next stepa of the protocol
    • When washing the column with 750uL of Wash Buffer 1, the bottom tube was filled so the entire wash could not go through the filter. Emptied the flow through and spun again pass all of the Wash Buffer through the binding column

Results of Miniprep

Sample ng/ul 260/280 260/230
RBS 144.44 1.99 2.57
J23100 121.77 1.95 2.53
RFP 171.78 2.00 2.59
YFP 144.35 1.98 2.53
  • NOTE: All results were done using a nanodrop 1000 with 1uL of sample
    • The Blank was pure water, not the buffer that the plasmids were suspended in.