Title: Bacterial Conjugation
1Bacterial Conjugation
- Conjugation will be used as a means to transfer
genetic information from one bacteria to another - Cells with a F plasmid or a R plasmid can
transfer their DNA to other bacteria - On the F and R plasmids, DNA transfer begins at
a specific sequence, OriT, the Origin of
Transfer. - OriT can be knocked out to severely hinder the
efficiency of conjugation, but the efficiency can
be restored by a complementation in trans.
2OriT Complementation
3Conjugation Assays
Matings with recipient cell ec100Dpir, which
is TriR, show if the F or R plasmid transfers by
the TriK plate and if an OriT plasmid transfers
by the TriA plate
Mutation and complementation of oriT for both F
and R plasmids works!
4Sending the Key to the Lock
Mating Results of sending the key plasmid to a
cell holding the locked RFP plasmid,
pJ23006-J23077
Mating Results of sending the key plasmid to a
cell holding a OnRFP plasmid, pSB1A2-J01022
Assay shows the key can be transferred
efficiently to the lock!
5Tecan Fluorescence Assay
The Tecan test shows that the key plasmid results
in the recovery of fluorescence
The controls show that conjugation is equivalent
to a cotransformation of the key and lock
plasmids into a cell
RFP Flurorescence Graph
Graph will NOT BE USED