Team:Cambridge/Experiments/Initial Exercise Group One

Initial Exercise: Katy, Heather, Marta and Veronica

Fusion of Div IVA protein with GFP

Role of Div IVA

Div IVA controls septum positioning in Bacillus Subtilis

Mutants show

• inhibition of cell division initiation*
• abnormally placed septum*
• some small anucleate cells*

Reasons for Choice

• constitutively expressed*
• Distinct characteristic distribution that is easy to observe*
• successful divIVA-GFP fusion documented by Edwards and ERrington, 1997*

Construct Design

We wish to:

• keep the original promotor for the Div IVA*
• remove GFP promotor sequence*
• remove DivivA stop codon*
• Remove ComGA stabiliser sequence*
• ‘Linkers’ were described by Edwards and Errington - we suspect that this is related to the restriction enzyme assembly process they used and hence didn’t include it in our design*

Primer Design

used the Gibthon Construct Designer developed by the 2010 iGEM Cambridge team

Left side of the Div IVA coding sequence(site 1)

Right side of the Div IVA coding sequence(site 2)