Team:Goettingen/NoteBook w9

Inoculation of 8 Clones from each, Riboswitch, DarR and E0840 (were put in the fridge on friday, plated 23. and 24.7.)

 ->E0840 heavily contaminated with something that looks like fungus. Plates discarded.

->This leaves DarR and the Riboswitch for inoculation in 4ml Cam cultures for a miniprep tomorrow.

->in addition, a masterplate was plated.

Hybridization of iGEM_63 and iGEM_64 to generate the Promoter(part3)-Operator construct

The primers were first diluted in HPLC-H2O according to the QC-sheet (100µM). Then a 1:20 dilution stock was pipetted. For this experiment, the 1:20 dilution stocks were used.

Protocol:

10µl iGEM_63

+         10µl iGEM_64

           20µl reaction

->80°C for 10 minutes (heatblock)

->after 10 minutes, turn the heatblock off and let the sample slowly cool down in it

-> The ends of the oligos are “digested” with EcoRI and SpeI and phosphorylated. The product of this experiment can directly be ligated in front of the GFP-part!

See the gel tomorrow

Restriction digestion of Part 1-4 and Part 7

-          Part 1-4 digested with SpeI FD

-          Part 7 digested with EcoR1+SpeI FD + Phosphatase treatment (last 20 minutes of digestion)

-          40 µl setups, standard assay.

->for part 1-4, restriction with PstI (not FD, because empty) on 30.7.