Fragile X [fra(X)] cell lines have been established from an expressing 46,XX amniocyte culture. The amniocyte cells were transformed by the addition of an origin defective pSV40 vector. Fra(X) expression was observed at a frequency of 3% in the parental amniocyte cell line, and the transformed clones expressed the fra(X) site at a frequency of 9-13%. These cell lines maintain the cytogenetic phenotype and can serve as positive controls for testing the efficacy of the inducing systems during prenatal diagnostic studies.