FIGURE 2.1 Preimplantation development. The oocyte (unfertilized egg) combines with sperm to form a zygote (fertilized egg). Each gamete (oocyte or sperm) is haploid (has a single set of chromosomes); the zygote and all later cells are diploid (have two sets of chromosomes). The zygote then divides approximately once a day. Since there is no growth during this period of cell division (cleavage), the cells become progressively smaller. By 3-4 days, a ball of cells (morula) has formed. By 5 days, it has become hollowed out to form a blastocyst, which consists of a sphere 0.1-0.2 mm in diameter comprising two cell types—an outer shell of trophectoderm cells and an inner collection of 30-34 cells called the inner cell mass. By day 6, the blastocyst would normally implant into the uterine wall, the trophectoderm would begin to form the placenta, and the inner cell mass would begin to form the cells and tissues of the fetus. At the blastocyst stage, cells of the inner cell mass are undifferentiated and pluripotent; that is, they have the potential to differentiate into all cells of the fetus except the placenta. If separated from the blastocyst and cultured, the cells of the inner cell mass can be converted into embryonic stem cells that are also pluripotent and can be propagated extensively while maintaining that potential. Blastocyst picture from http://stemcells.nih.gov/info/scireport/chapter3.asp.

organisms. The latter subject raises possibilities of cell-based therapies to treat disease, often referred to as regenerative medicine.

Scientists discovered how to obtain or derive embryonic stem cells from mouse blastocysts in the early 1980s (Evans and Kaufman, 1981; Martin, 1981) by culturing inner cell masses on feeder layers of mouse fibroblasts. It was later discovered that feeder cells could be replaced with culture medium containing the growth factor leukemia inhibitory factor (LIF)(Smith et al., 1988; Williams et al., 1988). Mouse ES cells (mES cells) have been studied in the laboratory, and a great deal has been learned about their essential properties and what makes them different from specialized cell types.

mES cells are shown to be pluripotent using three kinds of tests. The first and most rigorous test is to inject mES cells into the blastocoel cavity of a blastocyst (Stewart, 1993). The blastocyst is then transferred to the uterus of a pseudopregnant female (a female primed to accept implanted blastocysts). If the mES cells are pluripotent, the resulting progeny will be a chimera because it consists of a mixture



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