Derivation of embryonic stem cells from wild-derived mouse strains by nuclear transfer using peripheral blood cells

The laboratory mouse (Mus musculus domesticus) has long been used in biomedical research due to its easy maintenance and availability of abundant genetic information. However, laboratory mice have an inherent disadvantage of low genetic diversity caused by their origin from a relatively small colony of “fancy mice” in western Europe. Recently captured wild mice (wild-derived mice) are expected to compensate for this shortcoming because of their greater genetic diversity. On the other hand, wild-derived mice often show poor reproductive performance and are difficult animals in which to apply assisted reproductive technologies. In this study, we examined the technical feasibility of derivation of nuclear transfer embryonic stem cells (ntESCs) from wild-derived mouse strains for their safe preservation. We used leukocytes collected from peripheral blood as nuclear donors without sacrificing wild-derived mice. We successfully established 24 ntESC lines from two wild-derived strains of CAST/Ei and CASP/1Nga (11 and 13 lines, respectively), both belonging to Mus musculus castaneus, a subspecies of laboratory mouse. Most (23/24) of these lines had normal karyotype, and all lines examined showed teratoma formation ability (4 lines) and pluripotent marker gene expression (8 lines). Two male lines examined (one from each strain) were proven to be competent to produce chimeric mice following injection into host embryos from laboratory mice. By natural mating of these chimeric mice, the CAST/Ei male line was confirmed to have germline transmission ability. Our results demonstrate that intersubspecific ntESCs derived from peripheral leukocytes could provide a strategy for preserving genetic resources of wild-derived mouse strains. Gene expression analysis of inter-subspecies ntESCs lines established from leukocyte SCNT blastocysts.