Registry/Measurement kit/Instructions: Difference between revisions
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=Promoter Measurement Kit Instructions= | |||
The steps for using the Promoter Measurement Kit for testing a promoter. | |||
=Oligo design= | |||
We recommend building your promoter from oligo annealing. If you are testing a part or device larger than ~80bp you can use miniprep and digest in place of oligo annealing. Just be sure to gel purify the part from the plasmid it was on if that plasmid has KanR -- if it doesn't contain KanR a purification step should probably be skipped (due to the inefficiency of a typical gel isolation). | |||
Oligos should be designed to attach a BioBrick prefix and suffix to your promoter, and to create an EcoR1 overhang on the 5' end and a SpeI overhand on the 3' end. You should order your oligos with a 5' phosphate or [[PNK Treatment of DNA Ends|add one with PNK]]. | |||
=Ligation= | |||
=RBS Measurement Kit Instructions= | |||
=Old Notes= | |||
Just posting notes here from going through the process of building the 16 test constructs. Since this is essentially the process that a user of the kit would be doing when they inserted their RBS or Promoter. | Just posting notes here from going through the process of building the 16 test constructs. Since this is essentially the process that a user of the kit would be doing when they inserted their RBS or Promoter. | ||
Revision as of 09:45, 14 January 2008
Promoter Measurement Kit Instructions
The steps for using the Promoter Measurement Kit for testing a promoter.
Oligo design
We recommend building your promoter from oligo annealing. If you are testing a part or device larger than ~80bp you can use miniprep and digest in place of oligo annealing. Just be sure to gel purify the part from the plasmid it was on if that plasmid has KanR -- if it doesn't contain KanR a purification step should probably be skipped (due to the inefficiency of a typical gel isolation).
Oligos should be designed to attach a BioBrick prefix and suffix to your promoter, and to create an EcoR1 overhang on the 5' end and a SpeI overhand on the 3' end. You should order your oligos with a 5' phosphate or add one with PNK.
Ligation
RBS Measurement Kit Instructions
Old Notes
Just posting notes here from going through the process of building the 16 test constructs. Since this is essentially the process that a user of the kit would be doing when they inserted their RBS or Promoter.
Example miniprep of measurement kit components
- GFP Promoter Tester Version 1 E/S <BBpart> I20244</BBpart> - 27.2
- GFP Promoter Tester Version 2 E/X <BBpart> I20245</BBpart> - 29.8
- RFP Promoter Tester Version 1 E/S <BBpart> I20246</BBpart> - 23.2
- RFP Promoter Tester Version 2 E/X <BBpart> I20247</BBpart> - 23.1
- GFP RBS Tester Version 1 E/X <BBpart> I20248</BBpart> - 31.5
- GFP RBS Tester Version 2 S/X <BBpart> I20249</BBpart> - N/A
- RFP RBS Tester Version 1 E/X <BBpart> I20250</BBpart> - 25.4
- RFP RBS Tester Version 2 S/X <BBpart> I20251</BBpart> - 30.2
