Lab Oct 16 2009

12.30am

Assemblies started.

K235034 (Chl) = K235033(Kan) const. promoter+32oC temp. sensing RBS + J04450(Kan) Lac controlled RFP + P1010-Chl

K235035 (Chl) = K235033(Kan) const. promoter+32oC temp. sensing RBS + K235032(Kan) lactose promoter+RBS+GFP+stop + P1010-Chl

K235038 (Chl) = J23102(Amp) const. promoter + B0034 (Amp) RBS + P1010-Chl

K235038 (Kan) = J23102(Amp) const. promoter + B0034 (Amp) RBS + P1010-Kan

1.30am

New subcultures started for K235036 and 37 for 30, 34, 37 and 42°C. (8 in total.) Used 30µL culture in 2mL LB.

5am

Transformations plated.

3pm

Disappointing results from the flourescence microscope, none of the temperature sensitive parts were working, however the lac regulated GFP (K235032) was. The interesting thing is that the ones with IPTG seemed to have slightly less flourescence than the ones without, which is backwards to our expectations.

Temperature 	Room temperature 	30°C 	34°C 	37°C 	42°C K235032 A expected result 	g 	g 	g 	g 	g 	This should be repressed (limited green) without lactose or IPTG independent of temperature. K235032 A actual result 	G 	G 	G 	G 	G 	These were quite green. K235032 B + IPTG expected 	G 	G 	G 	G 	G 	This should be induced by IPTG and produce green independent of temperature. K235032 B +IPTG actual 	g 	g 	g 	g 	g 	These were green. K235033 A expected 	/ 	/ 	/ 	/ 	/ 	Negative control to account for microscope artifacts - there is no colour producing part at all. K235033 A actual 	/ 	/ 	/ 	/ 	/ 	This is as expected. K235036 A expected 	/ 	/ 	R 	R 	R 	This should be controlled by the 32°C thermometer and red should be repressed under 32°C. K235036 A actual 	/ 	/ 	/ 	/ 	/ 	No flourescence. K235037 A expected 	/ 	/ 	G 	G 	G 	This should be controlled by the 32°C thermometer and green should be repressed under 32°C. K235037 A actual 	/ 	/ 	/ 	/ 	/ 	No flourescence. K235005 A expected 	R 	R 	R 	R 	R 	This has no promoter but might produce limited red independent of temperature. K235005 A actual 	/ 	/ 	/ 	/ 	/ 	No flourescence. J04630 expected 	G 	G 	G 	G 	G 	This has no promoter but might produce limited green independent of temperature. J04630 actual 	/ 	/ 	/ 	/ 	/ 	No flourescence.

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