Team:UNIPV-Pavia/Calendar/June/settimana5

June, 28th

First time in wet lab

11 LB agar plates, 35 LB agar + Amp plates and 15 LB agar + Cm 12.5 were prepared.

• <partinfo>BBa_C0060</partinfo> was resuspended from iGEM 2011 kit distribution, Plate 1, well 4K in 15 μl of ddH₂O
• <partinfo>BBa_C0061</partinfo> was resuspended from iGEM 2011 kit distribution, Plate 1, well 4M in 15 μl of ddH₂O
• <partinfo>BBa_J04450</partinfo> was resuspended from iGEM 2011 kit distribution, Plate 1, well 5E in 15 μl of ddH₂O
• <partinfo>BBa_K081022</partinfo> was resuspended from iGEM 2011 kit distribution, Plate 2, well 12N in 15 μl of ddH₂O

For each of them 1 μl was transformed in 100 μl of home-made TOP10 competent cells. Transformants were plated on LB agar plates with the right antibiotic and incubated ON at 37°C.

June, 29th

In the morning all plates showed colonies (<partinfo>BBa_J04450</partinfo> showed light red colonies); for each we picked one colony and inoculated it in 1 ml of liquid LB with the proper antibiotic (ampicillin for <partinfo>BBa_C0060</partinfo>, <partinfo>BBa_C0061</partinfo> and <partinfo>BBa_K081022</partinfo>, kanamycin for <partinfo>BBa_J04450</partinfo>). Cultures were grown at 37°C, 220 rpm.

In the afternoon glycerol stocks were prepared (750 μl of cultures and 250 μl of 80% glycerol) and stored at -80°C except for <partinfo>BBa_C0061</partinfo> which was not sufficiently grown (we let it grow until next morning); then cultures were refilled with 5 ml of liquid LB with the proper antibiotic and incubated ON at 37°C, 220 rpm.

June, 30th

All cultures were in saturation growth phase; glycerol stock for <partinfo>BBa_C0061</partinfo> was prepared and stored at -80°C.

Plasmids were extracted with MiniPrep kit; purified DNA was quantified with NanoDrop Spectrophotometer:

<partinfo>BBa_B0030</partinfo>, <partinfo>BBa_B0031</partinfo>, <partinfo>BBa_B0032</partinfo>, <partinfo>BBa_B0034</partinfo>, <partinfo>BBa_B0015</partinfo>, <partinfo>pSB4C5</partinfo>, <partinfo>BBa_I13501</partinfo>, <partinfo>BBa_I13507</partinfo> were inoculated from freezer glycerol stocks and grown ON at 37°C, 220 rpm for MiniPrep.