比较两种穿梭质粒在五种细胞中的克隆形成

将两种靶基因和选择基因不同的穿梭质粒以电穿孔法转入五种哺乳动物细胞,pMci3质粒有较强的宿主特异性,pESnx宿主范围较大。含质粒的HT克隆细胞自发突变率为10~(-1)～10~(-2);pM-ci3在FL、LTK、PA317、Wg3-h细胞中不能形成克隆,而pESnx在上述四种细胞中可形成克隆,但后三者克隆细胞的质粒DNA转入细菌后,转化子<30个/皿。只有pESnx/FL克隆细胞自发突变率低于10~(-4),转化子为50～200个/皿,质粒较稳定地存在于克隆细胞,且pESnx质粒的显色底物邻苯二酚价廉物美,所以pESnx/FL克隆细胞可成为一个比较好的测试系统。

A Comparison of the Clone Formation of Two Shuttle Vectors Among Five Mammalian Cell Lines

Two shuttle vectors based on Epstein-Barr virus: pMci3 and pESnx were transfected into cells with electroporation. The target gene and selective gene are different between the two vectors. The hoster's range of pMci3, which carries the bacterial lac I gene as target for mutation, is limited. While the shuttle vector with pESnx, xylE gene as the mutation target, poses wilder hosters. Both HT resistant clonal cells including pMci3 or pESnx show the spontaneous mutation frequencies of 10-2-10-1. There are no resistant clonal cells of PMci3 in cell lines of FL, LTK, PA317 and Wg3-h. On the contrary, we have gained resistant clonal cells of PESnx in the four call lines. But the transformants number of pE-Snx extracted from LTK,PA317,Wg 3-h clonal cells are less than 30 per Amp Plate.Only the pESnx/FL clonal cells render low spontaneous mutation frequency of less than 10-4 and the transformants more than 50 in per Amp plate. On the other hand, the substrate of Cat O2ase in pESnx, catechol, is cheaper than that of lad in pMci 3 , x-gal.So pESnx/FL clonal cells will be a more suitable testing system for detecting mutagens.