ZFX, an X chromosome-linked gene encoding a zinc-finger protein, has previously been shown to escape X inactivation in humans. Here we report studies of the inactivation status of the homolog, Zfx, on the mouse X chromosome. We took advantage of both the preferential inactivation of the normal X chromosome in females carrying the T(X;16)16H translocation and the high degree of nucleotide sequence variation between the laboratory strain of mouse [corrected] and Mus spretus genomes. An EcoRV restriction fragment difference between laboratory strain of mouse [corrected] and M. spretus was detected after amplification of Zfx transcripts using the reverse transcription-polymerase chain reaction. Using this allelic variation, we assessed expression of the two Zfx genes in females carrying the T(X;16)16H translocation (from laboratory strain of mouse [corrected]) and an intact X chromosome (from M. spretus). Such females exhibit Zfx transcription from the active laboratory strain of mouse [corrected] chromosome but not from the inactive M. spretus chromosome. These results indicate that the mouse Zfx gene is subject to X inactivation.