Construction and use of derivatives of transposon Tn4001 that function in Mycoplasma pulmonis and Mycoplasma arthritidis

Previous attempts to introduce transposon Tn4001 into Mycoplasma pulmonis a nd Mycoplasma arthritidis have not been successful, possibly due to functio nal failure of the transposon's gentamicin resistance determinant. Tn4001C and Tn4001T were constructed, respectively, by insertion of a chloramphenic ol acetyltransferase gene and the tetM tetracycline resistance determinant into Tn4001. Both Tn4001C and Tn4001T transposed in M. pulmonis, and Tn4001 T transposed in M. arthritidis. The incorporation of a Tn4001T derivative t hat contained lacZ into either Mycoplasma species resulted in transformants with readily detectable LacZ activity. Tn4001T may be of general utility f or use as a mycoplasma cloning vehicle because tetM functions in all specie s of Mycoplasma examined thus far.