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Bangalore The Company The Mission e-Notebook


The Bangalore iGEM Journal, 07




Contents

e-notebook

May 07 June 07 July 07
S M T W T F S S M T W T F S S M T W T F S
1 2 3 4 5 1 2 1 2 3 4 5 6 7
6 7 8 9 10 11 12 3 4 5 6 7 8 9 8 9 10 11 12 13 14
13 14 15 16 17 18 19 10 11 12 13 14 15 16 15 16 17 18 19 20 21
20 21 22 23 24 25 26 17 18 19 20 21 22 23 22 23 24 25 26 27 28
27 28 29 30 31 24 25 26 27 28 29 30 29 30

Current Status

Tasks Accomplished

Experiments so far

  • Calibration of the number of colonies obtained at different optical densities.
  • Transformation of competent E.coli (strain K12Z1) cells with constructs.
  • Induction of the construct pLac cfp

E.coli cells transformed with the construct pLac cfp were cultured for 10 hours in Luria Bertani medium containing 50mg/ml Ampicillin.

The cells were then transferred into a minimal medium (M9 medium) and were induced with the following concentrations of IPTG. A stock concentration of 100mM was used.

Sample 100mM IPTG (ul) 1mM IPTG (ul) Inoculum volume from LB (ul) Volume of M9 added (ml)
pLac cfp (0 uM) - - 3 ul 3 ml
pLac cfp (1 uM) - 3 ul 3 ul 3 ml
pLac cfp (10 uM) - 30 ul 3 ul 3 ml
pLac cfp (50 uM) 1.5 ul - 3 ul 3 ml
pLac cfp (100 uM) 3 ul - 3 ul 3 ml
pLac cfp (1000 uM) 30 ul - 3 ul 3 ml
K12Z1 (Autoflourescence) - - 3 ul 3 ml

Cells were allowed to grow till the optical density was in the range of 0.05-0.1 (early exponential phase) and were imaged using a phase contrast microscope.

Results: The mean fluorescence of CFP was seen to increase with increasing concentrations of the IPTG.

  • Induction of the construct pLac yfp

E.coli cells transformed with the construct pLac yfp were cultured for 10 hours in Luria Bertani medium containing 50mg/ml Ampicillin.

The cells were then transferred into a minimal medium (M9 medium) and were induced with the following concentrations of IPTG. A stock concentration of 100mM was used.

Sample 100mM IPTG (ul) 1mM IPTG (ul) Inoculum volume from LB (ul) Volume of M9 added (ml)
pLac yfp (0 uM) - - 3 ul 3 ml
pLac yfp (1 uM) - 3 ul 3 ul 3 ml
pLac yfp (10 uM) - 30 ul 3 ul 3 ml
pLac yfp (50 uM) 1.5 ul - 3 ul 3 ml
pLac yfp (100 uM) 3 ul - 3 ul 3 ml
pLac yfp (1000 uM) 30 ul - 3 ul 3 ml
K12Z1 (Autoflourescence) - - 3 ul 3 ml

Results: The mean fluorescence of YFP was seen to increase with increasing concentrations of the IPTG.

The Week Ahead

The to-do List

Discussions