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From 2013.igem.org

Morning

• Tried to make 1 litre of clear agar using :

5g NaCl

15g Tryptone

15g Agar

5g Soytone

Tryptone, NaCl and soytone were added into a mixer with a magnetic stirrer. (was a yellow colour).

3g of agar were added to two 200ml durans. The Tryptone mix was then added to the durans, filling it up to 200ml.

Mixture was yellow, not colourless - the experiment was not successful. Durans put in the autoclave in the evening to let the agar set overnight.

Afternoon

• Transformation of competent cells

(the standart protocol from iGEM was modified slightly)

1. Competent cells were thawed on ice.

2. 50µl of thawed competent cells was added into a pre chilled 2ml tube, and another 50µl was added to a different 2ml tube which was labelled 'control'.

3. 2µl of the resuspended DNA was added to the 2ml tube. This was pipetted up and down gently a few times. Competent cells were kept on ice.

4. 2µl of the RFP Control was added to the control transformation.

5. Tubes were closed and cells were incubated on ice for 30minutes.

6. The cells were heat shocked by immersion in a pre-heated water bath at 42°C for 60 seconds.

7. Cells were incubated on ice for 5 minutes.

8. SOC media was made by the addition of 500µl of glucose to SOB. 500µl of SOC media was added to each transformation.

9. Cells were incubated at 37°C for 1.5hours while the tubes were shaking.

10. 2 petri dishes were labelled ith LB agar and the appropriate antibiotic(s)