TnphoA is a derivative of Tn5 which makes gene fusios to the gene for E. coli alkaline phosphatase upon insertion into a target gene. This allows one to obtain gene fusions specifically to secreted proteins since these are the only targets that give phoA(+) gene fusions. TnphoA was developed by Colin Manoil and John Beckwith.
This plasmid is a suicide vector since the oriR6K is functional only in the presence of the pir gene (which is suplemented by lambda-pir prophage). pRT733 is provided in SM10 carrying this prophage. Mobilization of pRT733 into various gram-negative species has given mixed results. Km (R) colonies which are Ap(S) are transpositon events. However, depending upon the organism, we have also seen Km(R) Ap(R) colonies. We believe these are cointegrates but other possibilities exist. If you are lucky your transconjugates will be resistant to only Km.
|RT733||1379||SM10 (lambda-pir(+))/ pRT733 (oriR6L, Tra(-), Mob(+), Amp(R), Kan(R)||The plasmid pRT733 is mobilized into gram-negative species. It is a suicide vector, able to replicate only in the presence of pir(+)|
|TA2361||167||PhoN2||Pho (-), non-specific acid phosphatase|
|TA2362||168||PhoP12||Pho (-), non-specific acid phosphatase|
Hoffmann, C. and Wright, A.. 1985. PNAS 82: 5107-5111
Manoil, C. and Beckwith, J. 1985. PNAS 82: 8129-8133
Taylor et. al.1987. PNAS 84: 2833-2837