NUCLEO-CYTOPLASMIC INTERACTIONS IN DEVELOPMENT

我们建议研究核质间的相互作用。 在正常的发育过程中，并确定这些细胞 互动是可控的。不同组件的作用 发育中的哺乳动物受精卵(雄性基因组、雌性基因组、卵细胞质) 将使用最近开发的核移植技术进行分析。我们的 第一个主要目标将是确定发展的原因 双亲雄激素(带有两个男性原核的胚胎)和 双亲雌核生殖(两个雌性原核)。嵌合体的研究进展 由雌核发育或雄核发育与正常的聚集产生的胚胎 将对胚胎进行监测，以确定是胚胎还是 细胞致死性是等压胚胎死亡的最终原因。 从雄配子发生的连续阶段获得的单倍体核将 转移到去掉雄性核的受精卵中。这些 实验将解决出现功能差异的时间问题 在男性和女性基因组之间。第二个主要目标将是 去核受精卵的发育分析 发育阶段被转移。我们将确定是否会重复 核移植将促进这类胚胎的发育。对.的使用 来自较老胚胎(4-和8-细胞期)的细胞质作为核接受者 将会被探索。使用二维凝胶分析，我们将监测 核移植后的基因组表达。核形态、变化 在核蛋白(层蛋白)和有丝分裂器的结构中 这些核移植胚胎将被研究。

We propose to investigate nucleo-cytoplasmic interactions which take place during normal development and to determine the mechanism by which these interactions are controlled. The role of various components of the mammalian zygote (male genome, female genome, egg cytoplasm) in development will be analyzed using recently developed nuclear transfer techniques. Our first major objective will be to determine the reasons for developmental failure of biparental androgenones (embryos with two male pronuclei) and biparental gynogenones (two female pronuclei). Development of chimeric embryos produced by aggregation of gynogenetic or androgenetic and normal embryos will be monitored in order to determine whether embryo- or cell-lethality is ultimately responsible for death of isoparental embryos. Haploid nuclei obtained from successive stages of male gametogenesis will be transferred into zygotes from which male nuclei were removed. These experiments will address the timing of appearance of functional differences between the male and the female genome. The second major objective will be to analyze development of enucleated zygotes into which nuclei from older developmental stages are transferred. We will determine if repeated nuclear transfer will improve the development of such embryos. The use of cytoplasm from older embryos (4- and 8-cell stage) as the nuclear recipient will be explored. Using 2D gel analysis we will monitor the changes in genome expression following nuclear transfer. Nuclear morphology, changes in nuclear proteins (lamins) and the structure of the mitotic apparatus in these nuclear transfer embryos will be investigated.