From 2007.igem.org

Contents 1 Test of Ftsz TS strain 2 Transduction of MG1655 with P1 stock made on w121 3 Transformation of Biobricks from the iGEM2007 plates into DH5aplha 4 Microscopy of Acinobacter strain 5 PCRs

Test of Ftsz TS strain

The isolated clones of FtsZ TS all grew at 42°C and at 30°C. They must be mutants... So next time we'll try to find other clones from the source of the strain.

Transduction of MG1655 with P1 stock made on w121

We'll do the transduction only on MG1655 for the moment.
First step for transduction : adsorption
use of 03/07/07 (I) and 12/07/07 (II) phage stock from W121 : X2
MG1655 culture ON

• Control (1mL LB MgSO4 30mM; CaCl2 15mM)
• 5µL Phage + 900µL LB (MgSO4 30mM; CaCl2 15mM) + 100µL MG1655 Culture ON
• 50µL Phage + 900µL LB (MgSO4 30mM; CaCl2 15mM) + 100µL MG1655 Culture ON
• 500µL Phage + 500µL LB (MgSO4 30mM; CaCl2 15mM) + 100µL MG1655 Culture ON

20 min at 37°C
Centrifuge at 12000rpm during 5 min
Resuspend bacteria in 1,5mL LB (NaCitrate 20mM; DAP 300µM)
Incubation 1h10' at 37°C with agitation
Centrifuge 5 min at 5000rpm

Plate bacteria on LB-agar (NaCitrate 20mM; DAP 300µM)

Transformation of Biobricks from the iGEM2007 plates into DH5aplha

• BBa_I0500: Inducible pBad/araC in pSB2K3 (KanR)

well: 9I, Plate 2

• BBa_J23100: strong constitutive promoter in BBa_J61002 (AmpR)

well: 21E, Plate 3

• BBa_B0015: double terminator (B0010-B0012) in pSB1AK3 (AmpR)

well: 1I, Plate 1

Microscopy of Acinobacter strain

We learner how to use the microscope to see single cells (see protocol). Here we show the photos taken from Acinobacter grown in : left : right :

PCRs

We received the oligos !!! But not all of them :-(

3 PCRs:

• Lox71-FtsA-FtsZ1
• FtsZ2
• Lox66-DapAColi