Week from July 13rd, to July 19th, 2009

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July, 13rd

• This week we planned to assemble GFP protein generator under the control of Plac in our IPTG/lactose sensor (i.e. A11) in order to test it. We also planned to build up another aTc sensor in which tetR is expressed constitutively, but in smaller amount than in A9 BioBrick. Finally, we planned to put lacZ protein generator under the control of the lactose sensor in order to build up one of the final parts for our project: this part will express beta-galactosidase when lactose is present in the medium and this expression will be glucose-independent (see R0011 page in the Registry for more info).

• We infected 5 ml of LB + Amp with 8 ul of the following glycerol stocks:

• We incubated the inocula at 37°C, 220 rpm overnight.

July, 14th

• Miniprep for:

• Digestion for:

• Gel run/cut/purification: DNA yield was good for all the extracted parts, except A11-1(E-S), which was quantified as 3 ng/ul after Nanodrop measurement. Anyway, we decided to try to ligate it.

• Ligations:
  • A12 = J23118(S-P) + A4(X-P) in ~pSB1A2
  • A13 = A11(E-S) + A3(E-X) in pSB1AK3
  • A14 = A11(S-P) + E0240(X-P) in pSB1A2

• We incubated the ligations at 16°C overnight.

July, 15th

• We received pdc and adhB genes from Mr Gene! we will call them MRGENE1 and MRGENE2.
  • MRGENE1: 5 ug of liophylized DNA, Kan resistance plasmid;
  • MRGENE2: 5 ug of liophylized DNA, Amp resistance plasmid.

• We transformed the overnight ligations in TOP10 and plated the transformed bacteria. We incubated the plates at 37°C overnight.

July, 16th

July, 17th

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