Plasmid mediated mutagenesis of a cellular gene in transfected eukaryotic cells.
Plasmid mediated mutagenesis of a cellular gene in transfected eukaryotic cells.
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质粒介导的转染真核细胞中细胞基因的诱变。
DOI：
10.1093/nar/15.2.561
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发表时间：
1987
影响因子：
14.9
通讯作者：
Galloway,DA
中科院分区：
文献类型：
--
作者：
Brandt,CR;Buonaguro,FM;McDougall,JK;Galloway,DA
NIH 3 T3细胞广泛用于转化试验，并易于摄取转染的DNA。使用NIH 3 T3细胞设计了一个系统来测量转染DNA对受体细胞基因的致突变作用。NIH 3 T3细胞可以以表明至少一部分细胞仅具有HGPRT基因的一个功能性拷贝的频率突变为6-硫代鸟嘌呤抗性。它们具有低自发背景突变频率（101 ×10−7）。将三种不同的质粒转染到NIH 3 T3细胞中诱导6-硫鸟嘌呤抗性突变体，其频率范围为背景的3至11倍。突变体表型稳定，几个突变体的回复频率≤1×10−7。Southern杂交分析表明，26个突变体中有4个（15.4%）的HGPRT基因的结构有可检测的改变。有趣的是，通过转染HSV-2形态转化区获得了4个显示重排的突变体中的3个。
NIH3T3 cells are widely used in transformation assays and readily take up transfected DNA. A system has been devised using NIH3T3 cells to measure the mutagenic effect of transfected DNA on recipient cell genes. NIH3T3 cells can be mutated to 6-thioguanine resistance at a frequency which suggests that at least a portion of the cells have only one functional copy of the HGPRT gene. They have a low spontaneous background mutation frequency (≈1×10−7). Transfection of three different plasmids into NIH3T3 cells induced 6-thioguanine resistant mutants at frequencies ranging from 3 to 11 fold above background. The mutant phenotype is stable and reversion frequencies of several mutants are ≤1×10−7. Southern blot analysis of the HGPRT gene in several mutants showed that 4 of 26 mutants (15.4%) had detectable alterations in the structure of the HGPRT gene. Interestingly 3 of the 4 mutants showing rearrangements were obtained by transfection of the HSV-2 morphological transforming region.