PROMOTER TRAPS IN EMBRYONIC STEM-CELLS - A GENETIC SCREEN TO IDENTIFY AND MUTATE DEVELOPMENTAL GENES IN MICE
PROMOTER TRAPS IN EMBRYONIC STEM-CELLS - A GENETIC SCREEN TO IDENTIFY AND MUTATE DEVELOPMENTAL GENES IN MICE
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DOI：
10.1101/gad.5.9.1513
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发表时间：
1991-09-01
影响因子：
10.5
通讯作者：
SORIANO, P
中科院分区：
文献类型：
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作者：
FRIEDRICH, G;SORIANO, P
已经设计出在小鼠中选择插入突变的一般策略。设计了缺乏启动子并包含β-半乳糖苷酶基因或编码具有β-半乳糖苷酶和新霉素磷酸转移酶活性的蛋白质的报告基因的构建体，使得报告基因的激活取决于其在活性转录单位中的插入。此类插入事件会在标记基因中产生突变，并使其表达后具有β-半乳糖苷酶活性。通过电穿孔或逆转录病毒感染将启动子捕获构建体引入胚胎干（ES）细胞，产生了显示多种β-半乳糖苷酶表达模式的转基因系。来自 24 个表达 β-半乳糖苷酶的菌株的杂合子的交叉鉴定出 9 个菌株，其中纯合性导致胚胎致死。由于在其余菌株中没有检测到明显的表型，这些结果表明通过这种方法鉴定的哺乳动物基因的很大一部分对于发育不是必需的。
A general strategy for selecting insertion mutations in mice has been devised. Constructs lacking a promoter and including a beta-galactosidase gene, or a reporter gene encoding a protein with both beta-galactosidase and neomycin phosphotransferase activity, were designed so that activation of the reporter gene depends on its insertion within an active transcription unit. Such insertion events create a mutation in the tagged gene and allow its expression to be followed by beta-galactosidase activity. Introduction of promoter trap constructs into embryonic stem (ES) cells by electroporation or retroviral infection has led to the derivation of transgenic lines that show a variety of beta-galactosidase expression patterns. Intercrossing of heterozygotes from 24 strains that express beta-galactosidase identified 9 strains in which homozygosity leads to an embryonic lethality. Because no overt phenotype was detected in the remaining strains, these results suggest that a substantial proportion of mammalian genes identified by this approach are not essential for development.