Week from July 27th, to August 1st, 2009

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July, 27th

• Screening for the 11 miniprepped DNA samples for B1: digestion S-P for all.

• Gel results:
  • Samples 1, 4, 10, 11, 14, 19 and 20 showed an extra band for the non ligated plasmid;
  • Samples 2, 9, 13 and 18 were pure! we decided to keep B1-13 (lane 8) to perform future ligations.

• NOTE: we had a pure sample for B1 (i.e. B1-13)and three almost-pure sample for B2. Anyway, we decided to perform ligation reactions for these samples and the extra band of B2 will be eliminated during gel cut/purification. WE DECIDED TO KEEP B2-5. Next weeks we will think about purifying B2-5 itself.

• We transformed 20 pg of B1-13 purified DNA (stored at -20°C) in TOP10 in order to prepare a glycerol stock for this construct. We incubated the plate at 37°C overnight.

Preparation of experiment with Tecan F200

• We infected 5 ml of LB + Amp with 10 ul of A14pg, A8pg and A9pg glycerol stocks.

• We also infected 5 ml of LB + Amp with a single colony taken from B0030 native plate (stored at +4°C).

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

July, 28th

• We streaked LB agar plates + suitable antibiotic with iGEM stabs:

K116001	K116002	K112405
P0412	I746902	I746903
K101017	F2620MIT1	F2620MIT2

• We incubated these "single colonies" plates at 37°C overnight.

• We picked a single colony from B1-13 plate to infect 1 ml of LB + Amp and incubated this inoculum for 5 hours and 1/2.

• We prepared a glycerol stock for B1-13.

• We aliquoted the remaining 250 ul of B1-13 bacterial culture in two different falcon tubes and re-filled them with 5 ml of LB + Amp.

• We also infected 5 ml of LB + Amp with 10 ul of B0015(X2) and B2-5(X2) glycerol stocks.

• We incubated these six cultures at 37°C, 220 rpm overnight.

Preparation of experiment with Tecan F200

• We diluted 1:1000 the overnight cultures of A14pg, A8pg, A9pg and B0030.

• We incubated the diluted cultures for 5 hours (37°C, 220 rpm).

• After 5 hours, we adjusted the OD600 at 0.025.

Experiment with Tecan F200

• Description
• Purpose:
• Materials & Methods
• Protocol
• Results

July, 29th

July, 30th

July, 31st

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