Appendix N. 
399 
As summarized above, most if not all cloned animals are abnormal 
because of faulty reprogramming after nuclear transfer. Does this 
epigenetic dysregulation affect the potential of ntES cells to generate 
functional somatic cells that can be used for cell therapy? To address 
this question, I will first compare the in vivo development of embryos 
with the in vitro process of ES cell derivation from explanted 
embryos. This will be followed by discussing the epigenetic state of 
the ES cell genome. Finally, I will contrast the phenotype of cloned 
mice derived from ES cell donor nuclei with that of chimeric mice 
generated by injection of ES cells into blastocysts. 
fi) The phenotype of an emhrvo is determined hv its donor nucleus 
As mentioned repeatedly above, embryos can be derived from the 
fertilized egg or from a somatic nucleus by SCNT. The potential of the 
resulting blastocyst, when implanted into the womb, to develop into 
a fetus and a postnatal animal depends strictly on the nature of the 
donor nucleus (Fig 5): (i) When derived fiom the zygote, most 
embryos develop to birth and generate a normal animal; (ii) Similarly, 
most blastocysts cloned from an embryonic stem cell donor nucleus 
develop to birth but, in contrast to the normally fertilized embryo, the 
great majority of the cloned animals v\nll be abnormal ("Large 
offspring syndrome") (Eggan et al., 2001; Humpherys et al., 2001); (iii) 
The great majority of cloned blastocysts derived from somatic donor 
nuclei such as fibroblasts or cumulus cells will die soon after 
implantation and only a few clones wfill survive to birth and these too 
will be abnormal, suffering once again from the Large offspring 
syndrome (Wakayama and Yanagimachi, 2001); (iv) Finally, the 
likelihood of cloned blastocysts derived from another type of somatic 
donor nuclei - those present in terminally differentiated lymphoid 
cells - to generate a cloned animal is extremely low and has not been 
achieved except by using a two step procedure involving the 
intermediate generation of embryonic stem cells (Hochedlinger and 
Jaenisch, 2002a). These observations suggest that a blastocyst 
retains an “epigenetic memory" of its donor nucleus. This memory 
determines its potential for fetal development: while a fertilized 
embryo develops normally, any embryo derived by SCNT will be 
abnormal though the efficiency of a given clone to develop to birth is 
strongly influenced by the differentiation state of the donor cell (see 
Table 1). In other words, the cloned embryo after implantation into 
the womb will be abnormal because the cloned blastocyst retained 
an epigenetic memory of its donor nucleus and this causes faulty 
fetal development. This epigenetic memory is erased when a 
blastocyst, either derived by nuclear cloning or from the fertilized 
egg, is explanted into tissue culture and grown into an embryonic 
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