Chinese hamster cells frequently have altered karyotypes. To investigate the basis of recent observations that karyotypic alterations are related to telomeric fusions, we asked whether these alterations are due to lack of telomere repeat binding factor/s. Further, Chinese hamster chromosomes contain large blocks of interstitial telomeric repeats, which are preferentially involved in chromosome breakage and exchange, rendering it an interesting model for such studies. Here, we report on the cloning and the chromosomal localization of the Chinese hamster telomere repeat binding factor, chTRF1. The sequence analysis revealed, similar to human TRF1 (hTRF1), an N-terminal acidic domain, a TRF1 specific DNA binding motif and a C-terminal Myb type domain. Unlike mouse TRF1 (mTRF1), chTRF1 shows 97.5% identity to hTRF1. chTRF1 gene was localized on the long arm of chromosome 5. In vitro translation of chTRF1 resulted in protein product similar in molecular weight to hTRF1. Immunostaining of Chinese hamster ovary cells (CHO) with anti-TRF1 antibody revealed punctate nuclear staining. At metaphase, antibodies failed to detect TRF1 on most of the chromosome ends and the interstitial telomeric repeat bands. These studies suggest that chTRF1 does not bind the interstitial telomeric repeats, and its presence at the metaphase chromosome ends is limited. The later could be a factor contributing to frequent karyotypic alterations observed in Chinese hamster cells.