Team:Goettingen/week6-2

V06_04

• Experiment:
  For the chemical retransformation the standard protocol for transformation was followed. The following constructs were transformed into E. coli:
  20E - #2
  20E - #3
  18M - #2
  18M - #3
  18C - #1
  18C - #2
  
• Observations & Results:
  The retransformation was successful since all plates showed numeours colonoes except the negative control.
  

V06_05

• Experiment:
  In order to clone flhDC behind further promoters, the purified PCR product was digested with XbaI and PstI, whereas the BioBrick plasmids were cut with SpeI and PstI. The digestion was performed as described in the protocol. This time the follwing promoters were used:
  J23113 - 20G - x21
  J23109 - 2G - x106
  J23114 - 20I - x256
  J23106 - 18O - x1185
  J23104 - 18K - x 1831
  

• Experiment:
  For the chemical transformation the standard protocol was followed. The following constructs were transformed into
  E. coli:
  J23112 - 20E - x1
  J23113 - 20G - x21
  J23109 - 2G - x106
  J23114 - 20I - x256
  J23105 - 18M - x623
  J23106 - 18O - x1185
  J23104 - 18K - x 1831
  J23100 - 18 C - x 2547
  I0500 - 14N - pBAD/araC promoter
• Observations & Results:
  The transformation was successful. Numerous colonies on all plates as well as on the positive control. Now growth could be observed on the negative control plate, so the competent cells are preparation of the competent cells was correct this time. Additionally, no growth could be observed at the 14N plate. This is the only vector containing a kanamycin resistance instead of ampicillin. Eventually, the kanamycin concentration was too strong and inhibited growth.
  The strain (DH10B) grows very fast, thus try not to incubate too long to prevent formation of satellite colonies.
  The cells hosting plasmids with strong promoters are colored red due to the rfp gene fused to the promoter. This phenomenon can be used to verify correct transformation.
  

V05_27

• Experiment:
  In order to isolate the plasmids, over night cultures of the E. coli (DH10B) cells transformed with the different constitutive promoters were prepared.