STUDIES ON REGULATION OF DEVELOPMENT BY GENOMIC IMPRINTING

本研究旨在通过对小鼠卵母细胞发育过程中印记基因表达和胚胎发育的研究，阐明小鼠卵母细胞发育过程中印记基因的建立。生长期卵母细胞缺乏成熟进入第二次减数分裂中期并支持发育的能力。为了解生长期卵母细胞的核活性，采用核移植法构建了含有生长期1细胞核的成熟卵母细胞。经体外受精后，卵母细胞中含有直径为60μm的幼鼠核，可发育至足月。以成年小鼠卵巢卵母细胞为核供体时，直径为50μm的卵母细胞核可支持胚胎的长期发育。为了评估印迹状态对发育的响应，通过RT-PCR检测印迹基因在来自构建的卵母细胞的胎儿中的表达。结果表明，在非生长期卵母细胞等位基因中，父源表达基因被激活，母源表达基因被抑制，从卵母细胞生长早期到晚期，父源表达基因被激活，母源表达基因被抑制，父源表达基因被激活，母源表达基因被抑制。这些结果表明，在卵母细胞的生长过程中发生的母体基因组的修改是必不可少的哺乳动物的发展。

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.