Cloned Mice: RESULTS(4)


Substantial differences were also observed between cy-toplasts in their rate of development after nuclear reconstruction (Table 4). Whereas high proportions of MII cy-toplasts cleaved and formed morulae/blastocysts, these stages of development were only reached by 14% and 2% of activated cytoplasts, respectively. Of oocytes parthenogenetically activated with ethanol, 85% and 75% cleaved and reached the morula/blastocyst stage, respectively (n = 40). No cloned blastocysts could be transferred into recipients when activated cytoplasts were used. However, following transfer of 99 cloned morulae/blastocysts from MII cytoplasts into four recipients, 11 pups were born, of which 6 survived. whitening gel

Meiotic Resumption Following Ethanol Activation of B6CBAF1 Oocytes

The efficacy of IE and cytoplast developmental potential were next considered using oocytes from an alternative strain of mice (B6CBAF1). First, the timing of the A-TII transition and PB formation following ethanol activation were examined for this strain (Table 5). Compared with B6D2F1 oocytes, B6CBAF1 oocytes were slow to resume meiosis but quick to catch up. By 15 min pa, most oocytes (97%, n = 35) were still in MII, whereas by this point, we had previously observed 85% of B6D2F1 oocytes to already be at AII (Table 1). By contrast, the majority of B6CBAF1 oocytes (92%, n = 36) had reached the TII stage with the spindle lying parallel to the cell surface by 30 min pa, compared with 52% of B6D2F1 oocytes. However, by 60 min pa and onward, no difference was observed in the timing of meiotic resumption between the strains. Despite the minor initial differences in timing, the same protocol of demecolcine-treatment optimized using B6D2F1 oocytes was applied to B6CBAF1 oocytes. Immunocytochemical assessment of these oocytes at 90 min pa revealed the same PB phenotypes, IE, and compartmentalization (Fig. 2).

TABLE 4. Development following reconstruction of B6D2F1 oocyte cytoplasts with HM-1 embryonic stem cell nuclei.
Table4Cloned Mice Derived from-6

Table 5.
Table5Cloned Mice Derived from-7