Aspergillus fumigatus is the most common opportunistic human fungal pathogen responsible for invasive aspergillosis. Gene manipulation is critical for the investigation of A. fumigatus biology and pathogenesis at the molecular level, and it often requires integration of the introduced DNA into the fungal genome. Here we have searched and identified two potential “safe haven” regions, SH1 and SH2, based on A. fumigatus genome annotation and transcriptome data. When a DNA fragment carrying a fluorescent protein gene mNeonGreen (mNG) and a drug selection marker was inserted into SH1 or SH2, the expression of mNeonGreen was easily detected, indicating that SH1 and SH2 are not surpressive genetic regions. We found that insertion of this DNA fragment into SH1 did not cause any significant changes in the expression of neighboring genes. Insertion of this DNA into either SH1 or SH2 did not significantly alter any of the phenotypes that we analyzed comparing to the wild type control. By comparison, transformants with random ectopic integration of the same DNA fragment showed a wider range of variation in mNeonGreen expression and in virulence in an insect infection model. Having identified predetermined “safe-haven” regions in A. fumigatus could therefore help reduce experimental variations and increase reproducibility, as it has been for the C. neoformans field.

中文翻译：

---

烟曲霉中的一个跨基因“避风港”区域

烟曲霉是引起侵袭性曲霉病的最常见的机会性人类真菌病原体。基因操作对于在分子水平上研究烟曲霉生物学和发病机理至关重要，它通常需要将引入的DNA整合到真菌基因组中。在这里，我们根据烟曲霉基因组注释和转录组数据搜索并确定了两个潜在的“避风港”区域SH1和SH2。当将带有荧光蛋白基因mNeonGreen（mNG）和药物选择标记的DNA片段插入SH1或SH2中时，很容易检测到mNeonGreen的表达，表明SH1和SH2不是抑制性遗传区域。我们发现此DNA片段插入SH1不会导致任何邻近基因的表达发生任何重大变化。与野生型对照相比，将这种DNA插入SH1或SH2不会显着改变我们分析的任何表型。相比之下，在昆虫感染模型中，具有相同DNA片段的随机异位整合的转化子在mNeonGreen表达和毒力方面显示出更大的变异范围。因此，已经确定了烟曲霉中预定的“避风港”区域，就可以帮助减少实验变异并提高可重复性，就像对于新孢梭菌田一样。