NUCLEAR RECONSTITUTION AROUND PURIFIED HELA CHROMOSOMES IN A CELL-FREE SYSTEM FROM XENOPUS EGGS
Zhang Chuan-mao Qu Jian Zhai Zhong-he (Department of Biology, Peking University, Beijing 100871)
Activated Xenopus egg extracts are capable of reconstituting nucleus around the added DNA or chromatin. To understand better the nuclear reconstitution process and the structure of the assembled nucleus, we have developed a cell-free system using activated Xenopus egg extracts and metap-hase chromosomes of HeLa cells. We demonstrate that the extracts are efficient promoting the decondensation of the metaphase chromosomes and the nuclear reconstitution around the chromosomes, and that the nuclear reconstitution is a precise process. The reassembled nucleus possesses a typical nuclear envelope, pore complexes, nuclear lamina, nuclear matrix, chromatin, etc., as shown by electron microscopy, embedment and embedment-free electron microscopy, and immunofluorescence method. We also found that the annulate lamellae directly take part in nuclear envelope and pore complex reconstitution by fusing with the double membrane, having reassembled on the surface of chromatin/chromosomes.