Inner cell mass-free blastocysts, also referred to as trophoblastic vesicles (TVs) could be used as carriers to be injected with in vitro cultured embryonic cells and to generate clones of animals, providing an alternative method of mammalian cloning. Here we compare three methods of obtaining mouse TVs. Culturing preimplantation embryos in heat shock conditions brings about up to 35% of TVs having up to 47 cells and the efficiency is dependent on the stage of cultured embryos. Live birth was obtained after transfer of ES cell-injected TVs to recipient mice. Culture with radioactive precursor yields up to 55% of TVs, but only at concentrations supplying 37% TVs is their cell number sufficiently high. The effects of PMA treatment are 47% of TVs.