Direct removal in the mouse of a flexed neo gene from a three-loxP conditional knockout allele by two novel approaches
Direct removal in the mouse of a flexed neo gene from a three-loxP conditional knockout allele by two novel approaches
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DOI：
10.1002/gene.1025
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发表时间：
2001-05-01
期刊：
影响因子：
1.5
通讯作者：
Deng, CX
中科院分区：
文献类型：
--
作者：
Xu, XL;Li, CL;Deng, CX
PlxP-neo-loxP盒内含子的存在往往会严重干扰基因的表达。因此，许多研究人员通过在ES细胞中瞬时表达Cre来选择性地移除ploxP-neo-loxP盒，虽然添加ES细胞的操作有效，但可能会降低克隆通过生殖系传播的可能性，因此，我们开发了两种新的方法，通过Cre介导的小鼠重组来移除ploxP-neo-loxP。首先，将含ploxP-neo-loxP基因的小鼠与Ella-Cre转基因小鼠杂交。其次，将Cre表达载体注射到携带ploxP-neo-loxP等位基因的受精卵原核中，两种方法都产生了部分和完全切除的嵌合体小鼠。然后，这些嵌合体小鼠交配，在只筛选了几窝小鼠后，在后代中发现了新基因缺失的条件性基因敲除等位基因。除了通常使用的删除ES细胞中的NEO的方法外，这些程序还提供了直接在鼠标中移除NEO的选项。创世纪30：1-6,2001，出版于2001 Wiley-Liss，Inc.(Dagger)
The presence in an intron of the ploxP-neo-loxP cassette often results in severe interference with gene expression. Consequently, many investigators selectively remove the ploxP-neo-loxP cassette by transient expression of Cre in ES cells, Although effective, the added manipulation of the ES cells may reduce the likelihood that a clone will be able to transmit via the germline, Therefore, we developed two novel approaches that remove the ploxP-neo-loxP by Cre-mediated recombination in mouse. First, the ploxP-neo-loxP-containing mice were crossed with Ella-Cre transgenic mice. Second, a Cre-expression plasmid was injected into pronuclei of fertilized eggs bearing the ploxP-neo-loxP allele, Both approaches produced mosaic mice with partial and complete excision. These mosaic mice were then mated, and the neo-less conditional knockout allele was found in the offspring after screening only a few litters. These procedures provide options for removing neo directly in the mouse in addition to the commonly used approach that deletes neo in ES cells. genesis 30: 1-6, 2001, Published 2001 Wiley-Liss, Inc.(dagger)