Team:Paris Bettencourt/Notebook/Phage Sensor/Thursday 18th July.html

Phage Sensor

ASDF

Thursday 18^th July

Calculating the transformation efficiency for the E.coli NEB and E.coli NNEB strains.

· 20 µL of cell culture was inoculated in 200 µL of LB, where 0.5 µL of DNA sample (?) was added, giving the total volume of 220.5 µL.
· After the heat shock transformation 10 µL of the culture was diluted in 90 µL of LB, giving the 10-1 dilution (d); then 10 µL of this dilution was transferred in 90 µL of LB, giving the 10-2 dilution. 10 µL of each dilution (v) was plated on ampicillin media and left for incubation (24 h, 37 ̊C).


· After incubation, colonies were counted:

	NEB	NNEB
10-¹	0	433
	0	368
Average	0	400.5
10-¹	0	62
	0	40
Average	0	51

Obviously, NEB strain was not transformed and only NNEB was included in further caluculations.
· The number of the transformed colony forming units per mL of the culture was calculated following the formula: colony number / d*v.

Calculation from the 10-1 dilution: 0.4 * 109 CFU/mL
Calculation from the 10-2 dilution: 0.51 * 109 CFU/mL

· The transformation efficiency was calculated by dividing the number of transformed colony forming units (CFU) by the amount of the DNA used (mDNA).

CFU1 = 0.4 * 109 CFU/mL * 0.22 mL (total V of the transformation culture) = 0.088*109 CFU
CFU2 = 0.51 * 109 CFU/mL * 0.22 mL = 0.1122*109 CFU

The concentration of the DNA solution which was used was 0.323 µg/µL, while the total DNA mass which was added to the transformation culture was: 0.323 µg/µL * 0.5 µL = 0.1615 µg.

So the transformation efficiency, calculated from the 10 times diluted (TE1) and 100 times diluted (TE2) cultures is:

TE1 = CFU1/mDNA = 0.55 * 109 CFU/µg
TE2 = CFU2/mDNA = 0.69 * 109 CFU/µg

TE1 and TE2 are almost the same