STUDIES ON REGULATION OF DEVELOPMENT BY GENOMIC IMPRINTING

本项目旨在通过对小鼠卵母细胞生长过程中基因组印迹基因表达和胚胎发育的研究，阐明基因组印迹何时建立。生长期卵母细胞缺乏进入第二次减数分裂中期成熟和支持发育的能力。为了了解生长期卵母细胞的核能力，采用核移植的方法构建了具有生长期细胞核的成熟卵母细胞。体外受精后，幼鼠的卵母细胞含有直径为60μm的生长细胞核，能够发育足月。以成年小鼠卵巢卵母细胞作为核供体时，直径为50μm的卵母细胞的细胞核支持胚胎发育。为了评估印迹状态对发育的响应，利用RT-PCR检测了构建卵母细胞衍生的胎儿中印迹基因的表达。结果表明，在卵母细胞未生长的等位基因中，父本和母本表达的基因分别被激活和抑制，且这种调控在卵母细胞生长的早期到后期分别建立。这些结果表明，母系基因组在卵母细胞生长过程中发生的修饰对哺乳动物的发育至关重要。

This project was carried out to clarify when the genomic imprinting during oocyte growth is established in mice by investigations on imprinted gene expression and embryo development. Growing stage oocytes lack ability to mature into the metaphase of the second meiosis and support the development. To know nuclear competence of growing stage oocytes, fully-grown oocytes with nuclei from growing stage one were constructed by nuclear transfer. Following in vitro fertilization, the oocytes contained nuclei from growing one, 60μm in diameter, of juvenile mice were able to develop term. When oocytes from adult mice ovary were used as nuclear donors, the nucleus from oocytes of 50μm in diameter supported the term development. To evaluate the imprinted status in response to the developments, the expression of imprinted genes in the fetuses derived from constructed oocytes was examined by RT-PCR. The results showed that paternally and maternally expressed genes were activated and inhibited respectively in non-growing oocyte alleles, and that the regulation was established individually from early to late growing stage in oocytes. These results demonstrate that modifications in the maternal genome that occur during oocyte growth are essential for mammalin development.