Team:UCL/Labbook

June

Week 1-3

No lab work

Week 4

26th June - We are given safety training in the Advanced Centre for Biochemical Engineering in all relevant laboratories, as well as general procedures in case of emergency.

July

Week 5-6

No lab work

Week 7

Bacterial Lab

15th July - The team is introduced to the laboratories which will be used during the summer for both bacterial and mammalian experiments.

16th July - 5X M9 salts [link to protocol], minimal agar [link to protocol], 1.4% molten agar solution [link to protocol] and 0.1M CaCl2/15% glycerol [link to protocol] were prepared for the generation of competent cells. Minimal agar plates were poured and streaked [link to streaking protocol] with W3110 Escherichia coli cells and left overnight to incubate at 37C.

17th July - Very little colony growth was observed from W3110 E.coli streaked plates. Plates were therefore left to incubate for a further 17 hours.

18th July - Sufficient colony growth allowed for the selection of a single colony from each plate. This was then inoculated in 5ul LB media [link to LB recipe] + 100ul 1M MgSO4 and left to incu-shake overnight at 37C.

19th July - Cultures re-suspended in new LB media and 100µl aliquots placed into individual eppendorf tubes for placement at -80C.

Mammalian Lab

17th July - Mammalian cell culture and maintenance [link to mammalian protocol] training by Mrs. Ludmilla Ruban. Passaged primary MEF (mouse embryonic fibroblast) cells. Passage 3.

18th July - MEF passage 4

19th July - MEF passage 5

Week 8

Bacterial Lab

22nd July -Transformation [link to transformation protocol] of our competent cells with plasmid YB3110 was carried out.

23rd July - No colony growth was observed on Ampicillin plates [link to ampicillin plate protocol] indicating no plasmid uptake. Transformation was repeated with YB3110.

Vial	Ampicillin Plate	Plasmid Insertion	Colony Count
1 (Main)	Yes	Yes	0
2 (Positive Control)	No	Yes	100+
3 (Negative Control)	Yes	No	0

24th July - No colony growth from main experiment indicating no plasmid uptake. Transformation was repeated.

Vial	Ampicillin Plate	Plasmid Insertion	Colony Count
1 (Main)	Yes	Yes	0
2 (Positive Control)	No	Yes	100+
3 (Negative Control)	Yes	No	0

25th July - No colony growth and hence no plasmid uptake. Transformation was repeated.

Vial	Ampicillin Plate	Plasmid Insertion	Colony Count
1 (Main)	Yes	Yes	0
2 (Positive Control)	No	Yes	100+
3 (Negative Control)	Yes	No	0

26th July - Once again our cells were unsuccessful in taking up the YB3110 plasmid. We concluded that our batch of competent cells were not competent and so these will not be used in any further experiments. A new batch of competent cells are to be generated.

Vial	Ampicillin Plate	Plasmid Insertion	Colony Count
1 (Main)	Yes	Yes	0
2 (Positive Control)	No	Yes	100+
3 (Negative Control)	Yes	No	0

Mammalian Lab

22nd July - MEF passage 6. Training ends.

Week 9

Bacterial Lab

August