The role of the pre-T cell receptor (TCR) in lineage commitment to the gammadelta versus alphabeta lineage of T cells was addressed by analyzing TCRbeta chain rearrangements in gammadelta T cells from wild-type and pre-TCR-deficient mice by single cell polymerase chain reaction. Results show that the pre-TCR selects against gammadelta T cells containing rearranged Vbeta genes and that gammadelta T cell precursors but not gammadelta T cells express the pre-TCRalpha protein. Furthermore, pre-TCR-induced proliferation could not be detected in gammadelta T cells. We propose that the pre-TCR commits developing T cells to the alphabeta lineage by an instructive mechanism that has largely replaced an evolutionary more ancient stochastic mechanism of lineage commitment.