MOLECULAR-CLONING AND EXPRESSION OF DNA ENCODING OVINE INTERLEUKIN-2

We have generated DNA encoding the mature form of ovine interleukin 2 (IL-2) by polymerase chain reaction (PCR) using primers complementary to sequences at the 5' and 3' ends of human, murine and bovine IL-2 cD NA, The predicted PCR product of 400 bp was ligated into the yeast Ty- P1 galactose-inducible expression vector pOGS40 which was used to tran sform yeast spheroplasts. The fusion protein, with a Factor Xa proteol ytic cleavage site between ovine IL-2 and the F1 fusion partner, was e xpressed from galactose-induced transformed yeast. P1:IL-2 fusion prot ein, which self-assembles into virus-like particles (VLPs) due to the interaction of the F1 protein, was purified from lysates of mechanical ly disrupted yeast by centrifugation on a discontinuous sucrose gradie nt, Fusion protein was detected in Western blot analysis with polyclon al antisera raised to recombinant bovine IL-2, Soluble recombinant ovi ne IL-2 was released from the F1 fusion protein by cleavage with Facto r Xa enzyme, After purification recombinant ovine IL-2 was functionall y active as shown by its ability to support the proliferation of Con A -activated T cells and was capable of generating maedi visna virus-spe cific cytotoxic T cells from primed precursor cells, The availability of recombinant ovine IL-2 will greatly help the analysis of the specif icity of pathogen-specific cells in the sheep.