Spi- selection:: An efficient method to detect γ-ray-induced deletions in transgenic mice
Spi- selection:: An efficient method to detect γ-ray-induced deletions in transgenic mice
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DOI：
10.1002/(sici)1098-2280(1999)34:1
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发表时间：
1999-01-01
影响因子：
2.8
通讯作者：
Sofuni, T
中科院分区：
文献类型：
--
作者：
Nohmi, T;Suzuki, M;Sofuni, T
尽管基因组重排在癌症和人类遗传疾病的病因学中很重要，但转基因啮齿动物致突变性测试很难检测到缺失突变。为了促进体内缺失突变的检测和分子分析，我们建立了含有 lambda EG10 穿梭载体的转基因小鼠模型，该载体包括用于 Spi(-)（对 P2 干扰敏感）选择的 red 和 gam 基因 [Nohmi 等人，2017]。 [1996]环境。梅尔.诱变剂。 28:465-470]。这种选择比其他遗传系统有很大的优势，因为噬菌体缺失突变体可以优先选择为Spi(-)噬菌斑，然后可以对其进行分子分析。在这里，我们展示了 41 个由伽马射线照射诱导的缺失突变连接点的核苷酸序列。与自发缺失突变体不同，超过一半的大缺失发生在 1 至 8 bp 的短同源序列之间。其余的连接点没有这样的同源序列。有趣的是，两个 Spi(-) 突变体在断点处有类似 P（回文）的核苷酸添加，这在 V(D)J 重组的编码连接处经常观察到，这表明具有发夹结构的断裂 DNA 分子可能是修复辐射诱导的双链断裂的中间体。我们得出的结论是，Spi(-) 选择对于有效检测体内缺失突变非常有用，并且小鼠中伽马射线诱导的大多数重排是通过 DNA 末端连接的非法重组介导的。环境。摩尔。诱变剂。 34:9-15, 1999。(C) 1999 Wiley-Liss, Inc.
Despite the importance of genome rearrangement in the etiology of cancer and human genetic disease, deletion mutations are poorly detectable by transgenic rodent mutagenicity tests. To facilitate the detection and molecular analysis of deletion mutations in vivo, we established a transgenic mouse model harboring a lambda EG10 shuttle vector that includes the red and gam genes for Spi(-) (sensitive to P2 interference) selection [Nohmi et al. [1996] Environ. Mel. Mutagen. 28:465-470]. This selection has a great advantage over other genetic systems, because phage deletion mutants can be preferentially selected as Spi(-) plaques, which can then be subjected to molecular analysis. Here, we show nucleotide sequences of 41 junctions of deletion mutations induced by gamma-irradiation. Unlike spontaneous deletion mutants, more than half of the large deletions occurred between short homologous sequences from one to eight bp. The remaining junctions had no such homologous sequences. Intriguingly, two Spi(-) mutants had P (palindrome)-like nucleotide additions at the breakpoints, which are frequently observed in the coding junctions of V(D)J recombination, suggesting that broken DNA molecules with hairpin structures can be intermediates in the repair of radiation-induced double-strand breaks. We conclude that Spi(-) selection is useful for the efficient detection of deletion mutations in vivo and that most rearrangements induced by gamma-rays in mice are mediated by illegitimate recombination through DNA end-joining. Environ. Mol. Mutagen. 34:9-15, 1999. (C) 1999 Wiley-Liss, Inc.