Stimulation of gene transfer efficiency by treatment of human cells with topoisomerase II inhibitors

用拓扑异构酶 II 抑制剂处理人体细胞来刺激基因转移效率

• 批准号: 10660317
• 负责人: ARATANI Yasuaki
• 金额: $ 1.6万
• 依托单位: Yokohama City University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 1998
• 资助国家: 日本
• 起止时间: 1998 至 1999
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-10660317/
• 关键词: mammalian cell; gene transfer; topoisomerase; bleomycin; nonhomologous recombination; DNA

In mammalian cells, nonhomologous recombination is a predominant pathway to repair DNA double-strand breaks. We have shown that DNA topoisomerase II inhibitors are capable of enhancing random integration of foreign DNA via nonhomologous recombination. Since this enhancement is likely due to stabilized DNA strand breaks, we examined the effect of radiomineric antitumor drug, bleomycin, on nonhomologous recombination. We found that bleomycin greatly enhances the random integration of transfected plasmids into human cells. Also, we analyzed the integration junctions between transfected DNA and human chromosomes in clones derived from topoisomerase II inhibitor-treated and-untreated cultures as well as the characterization of inserted DNA found in integration junctions between transfected DNA and human chromosomes. These studies will facilitate the understanding of DNA integration events through nonhomologous recombination and the development of transfection protocols with higher efficiencies.

在哺乳动物细胞中，非同源重组是修复DNA双链断裂的主要途径。我们已经证明，DNA拓扑异构酶II抑制剂能够通过非同源重组促进外源DNA的随机整合。由于这种增强可能是由于稳定的DNA链断裂，我们检查了放射性抗肿瘤药物博莱霉素对非同源重组的影响。我们发现，博莱霉素极大地增强了转基因质粒进入人类细胞的随机整合。此外，我们还分析了经拓扑异构酶II抑制剂处理和未处理的克隆中转基因DNA与人类染色体之间的整合连接，以及在转基因DNA与人类染色体之间整合连接中发现的插入DNA的特征。这些研究将有助于通过非同源重组了解DNA整合事件，并开发更高效率的转染方案。

项目成果

小山秀機: "培養細胞ラボマニュアル(小山秀機編)"シュプリンガー・フェアラーク東京. 164 (1999)

小山秀树：“培养细胞实验室手册（小山秀树编辑）”Springer Verlag 东京 164（1999）。

Noriyuki Sugo: "Neonatallethality with abnornal neurogenesis in mioe deficient in DNA polymerase β"EMBO J. In press.

Noriyuki Sugo：“DNA 聚合酶 β 缺陷的 mioe 中神经发生异常导致的新生儿致死率”EMBO J. 正在出版。

Yasuaki Aratani: "Severe impairment in early host defense against Candida albicansin mice deficientin myeloperoxldase"Infect.Immun.. 67. 1828-1836 (1999)

Yasuaki Aratani：“早期宿主针对缺乏髓过氧化物酶的白色念珠菌小鼠的防御严重受损”Infect.Immun.. 67. 1828-1836 (1999)

Masao Kobayashi: "Characterization of the 3'untranslated region of mouse DNA topoisomerase Iia RNA"Gene. 215. 329-337 (1998)

Masao Kobayashi：“小鼠 DNA 拓扑异构酶 Iia RNA 3非翻译区的表征”基因。

Noriyuk Sugo: "Neonatal lethality with abnormal neurogenesis in mice deficient in DNA polymerase β."EMBO J. (in press).

Noriyuk Sugo：“缺乏 DNA 聚合酶 β 的小鼠神经发生异常导致新生儿死亡。”EMBO J.（出版中）。