论文部分内容阅读
过去有人提出EBV—DNA是通过病毒颗粒感染,或者是携带EBV基因的细胞与宿主细胞融合这两种途径之一,导致病毒基因转输入宿主细胞DNA。近年我们注意到:基因片段转染,是基因转移和导致细胞转化的一种重要形式。 本实验应用DNA介导的基因转移的钙沉淀改良法,在体外培养将EBV重复序列区的BamHl—W及其相邻片段转染宿主细胞(大鼠成纤维细胞,Rat—1),3周内出现转化灶,经半固体培养基进行转化选择,从而获得多株高效转化克隆,其中W_4和W_(10)通过裸鼠体内接种与鼠间移植,最后建立高移植成功率(94—100％)的可移植性裸鼠瘤(CH—1,CH—2)。 经SP6/T7RNA多聚酶诱导RNA探针、缺口翻译DNA探针和亚细胞定位的原位杂交探针综合应用,测知经EBV基因片转转染后多次传代的转化细胞(W_(10),W_4)和多次移植的裸鼠瘤(CH—1,CH—2)DNA中存在着EBV基因片段。另一方面,测知人类的鼻咽癌组织和Burkitt淋巴瘤细胞株以及绒猴的EBV转化细胞株中,恒定地整合着该基因片段。因此,EBV基因片段的活性及其在癌变过程中的作用,与癌基因激活等问题值得进一步研究。
In the past, it has been suggested that EBV-DNA is infected by virus particles, or it is one of the two ways in which cells carrying EBV genes fuse with host cells, resulting in the transfer of viral genes into host cell DNA. In recent years we have noticed that transfection of gene fragments is an important form of gene transfer and cell transformation. In this experiment, a DNA-mediated calcium precipitate-improving method for gene transfer was used to transfect EBV repeat region BamHl-W and its adjacent fragment into host cells (rat fibroblasts, Rat-1) in vitro for 3 weeks. Transforming foci appeared in the medium and were selected for transformation by semi-solid medium. Several highly efficient transformed clones were obtained, among which W_4 and W_10 were inoculated in nude mice and transplanted between mice. Finally, a high success rate of transplantation (94-100%) was established. ) Transplantable nude mouse tumor (CH-1, CH-2). Through the combined application of RNA probes, nick translation DNA probes and subcellular localization in situ hybridization probes induced by SP6/T7 RNA polymerase, the transformed cells transfected with EBV gene transfected cells were detected (W_(10), There are EBV gene fragments in W_4) and nude mouse tumor (CH-1, CH-2) DNA that has been repeatedly transplanted. On the other hand, in the human nasopharyngeal carcinoma tissues and Burkitt lymphoma cell strains as well as marsupial EBV transformed cell strains, the gene fragments were constantly integrated. Therefore, the activity of EBV gene fragment, its role in carcinogenesis, and oncogene activation deserve further study.