IGEM:Harvard/2008/Lab Notebooks/DailyBook/Week10/Chemical and Light

Transformations in Shewie

About 250 ng of DNA added each time.

First Set 8/22

Plate	Colonies	Size	Color
S1 P102 (P108+45) Kan	5 colonies	3mm to 5 mm	Pink
S1 P121+P38 (P124) 1:2.5 Kan A	1 colony	5mm to 7.5mm	Pink
S1 P28 (ColE1) Kan	0 colonies
S1 P121+P38(P124) 1:2.5 Kan B	0 colonies
S1 P121+P39 1:2.5 Dephos Kan A	0 colonies
S1 Topo vector (puc) 2ul Kan X-gal	>50 colonies	1mm	White
S1 Topo vector (puc) 4ul Kan X-gal	>50 colonies	1mm	White
S1 P121+P39 1:2.5 Kan D	0 colonies
S1 (-) ctrl (just cells) Kan	0 colonies
S1 (+) ctrl (E1 p59b) Kan	2 colonies	5mm	Pink

Poor transformation efficiency in the positive control suggests that there may be a problem with our cells. All samples, except for P102, were retransformed the following week with fresh cell cultures.

Second Set (with new cells) 8/27

Plate	Marker	Description
S1 P124 1:2.5 A	Kan	14 2-3mm pink colonies, ~100 0.5mm white colonies
S1 P124 1:2.5 B	Kan	2 pink 2mm colonies, >100 0.5mm white colonies
S1 P125 1:2.5 Dephos A	Kan	2 pink 2mm colonies, >100 0.5mm white colonies
S1 P125 1:2.5 D	Kan	8 2mm pink colonies, ~100 0.5mm white colonies
S1 P28	Kan	~8 1-2mm pink colonies, ~50 0.5mm white colonies
S1 TOPO 2uL	Kan	Lawn of (>200) 0.5mm white colonies
S1 TOPO 4 uL (w/ Xgal)	Kan	Lawn of (>200) 0.5mm white colonies
S1 P59b (+) ctrl	Kan	~50 2mm pink colonies, ~50 0.5mm white colonies
S1 (-) ctrl	Kan	50-100 0.5mm white colonies

Getting mtrB onto thermo and lac inducible systems

Ligating mtrB TOPO with RBS

Grew up more mtrB TOPO for next week.

PCR of mtrB with RBS + mtrB primers

Scheduled for next week.