首页> 外文期刊>Proceedings of the National Academy of Sciences of the United States of America >Nonhomologous chromosomal integration of foreign DNA is completely dependent on MUS-53 (human Lig4 homolog) in Neurospora
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Nonhomologous chromosomal integration of foreign DNA is completely dependent on MUS-53 (human Lig4 homolog) in Neurospora

机译:外源DNA的非同源染色体整合完全依赖Neurospora中的MUS-53(人类Lig4同源物)

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摘要

Homologous integration of a foreign DNA segment into a chromosomal target sequence enables precise disruption or replacement of genes of interest and provides an effective means to analyze gene function. However, integration after transformation is predominantly nonhomologous in most species other than yeast. Here, we show that homologous integration in the filamentous fungus Neurospora requires the homologous-recombination proteins MEI-3 (yeast Rad51 homolog) and MUS-25 (yeast Rad54 homolog), whereas nonhomologous integration requires nonhomologous end-joining protein MUS-52 (yeast Ku80 homolog). Two additional minor integration pathways are present, one MEI-3-independent and homologous, the other MUS-52-independent and nonhomologous. Homologous and nonhomologous mechanisms compete when external DNA is integrated. In Neurospora, both nonhomologous integration pathways, MUS-52-dependent and MUS-52-independent, require MUS-53 (a homolog of human Lig4), which functions in the final step of nonhomologous end-joining. Because nonhomologous integration is eliminated in a LIG4-disrupted strain, integration occurs only at the targeted site in mus-53 mutants, making them an extremely efficient and safe host for gene targeting.
机译:将外源DNA片段同源整合到染色体靶序列中,可以精确破坏或置换目的基因,并提供分析基因功能的有效手段。但是,除了酵母外,大多数物种中转化后的整合主要是非同源的。在这里,我们显示丝状真菌Neurospora中的同源整合需要同源重组蛋白MEI-3(酵母Rad51同源物)和MUS-25(酵母Rad54同源物),而非同源整合则需要非同源末端连接蛋白MUS-52(酵母Ku80同系物)。存在两种另外的次要整合途径,一种独立于MEI-3并且是同源的,另一种独立于MUS-52并且是非同源的。整合外部DNA时,同源和非同源机制相互竞争。在Neurospora中,依赖于MUS-52和不依赖MUS-52的非同源整合途径都需要MUS-53(人Lig4的同源物),该功能在非同源末端连接的最后一步起作用。因为在LIG4中断的菌株中消除了非同源整合,所以整合仅发生在mus-53突变体的目标位点,这使其成为基因靶向的极其有效和安全的宿主。

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