视网膜母细胞瘤细胞系SO—Rb50瘤细胞Rb基因突变动态变化研究

目的：研究SO－Rb50细胞系瘤细胞Rb基因突变的动态变化。方法：1．用Southern blot杂交法分析SO－Rb50细胞系第327代瘤细胞DNA。2．用PCR－SSCP法对SO－Rb50细胞系第415代及第713代瘤细胞的Rb基因27个外显子和1个启动子逐个筛查。3．将SO－Rb50细胞系的第775代克隆出3个细胞株：MC2、MC3及MC4。用PCR－SSCP－HA法对MC2、MC3及MC4第11代细胞和MC3第138代细胞的Rb基因的27个外显子逐个筛查。结果：SO－Rb50细胞系第327代细胞DNA缺失3．5Kb、2.9Kb及1.0Kb的三条带，证明SO－Rb50细胞系瘤细胞Rb基因缺失。第415代瘤细胞Rb基因第23外显子少一条单链；第713代第25外显子发生新的突变。SO－Rb50细胞系第775代的3个克隆株MC2、MC3及MC4的第11代细胞，只有MC4第24外显子突变；而MC3第138代第24外显子突变，提示MC3经多次传代后第24外显子发生新突变。结论：SO－Rb50细胞系在长期培养传代过程中，瘤细胞的Rb基因突变有动态变化。

Study on the Dynamic Changes of Retinoblastoma Gene of SO-Rb50 Cell Line

Purpose: To study the dynamic changes of Retinoblastoma Gene of SO-Rb50 cell line. Methods: 1) Southern blot hybridization was used to analyse the tumor cell DNA of 327th passage of SO-Rb50 cell line. 2) A promoter and 27 exons of Rb gene were screened exon-by-exon by using PCR-SSCP at 415th and 713th passages of SO-Rb50 cell line. 3) Three cell cloning strains named as MC2, MC3 and MC4 were isolated by single cell cloning technique from the SO-Rb50-775, and mutation of Rb gene were also screened exon-by-exon by using PCR-SSCP-HA in MC2-11, MC3-ll, MC4-11 and MC3-138.Results: The 3.5Kb, 2.9Kb and 1.0Kb bands were deleted in the DNA of SO-Rb50-327 tumor cells, showing the deletion of Rb gene in SO-Rb50 cell line. Exon23 of 451th passage cells decreased one band; but exon 25 of 713th passage cells decreasedtwo bands, indicating that exon 25 had new mutation. PCR-SSCP-HA analysis of ex-on24 showed that MC4-11 and MC3-138 had abnormal bands, but MC2-11 and MC3-ll weren't found mutation of Rb gene. This result suggested that new mutation occurred to exon24 of MC3 during a long-term culture in vitro.Conclusion: Retinoblastoma gene mutation of SO-Rbso cell line had dynamic changes during a long-term culture. Eye Science 2001; 17: 111 - 113.