Mitsubishi Kagaku Institute of Life Sciences, 11 Minamiooya, Machida, Tokyo 194-8511, Japan.
J Biochem. 2010 Jun;147(6):819-22. doi: 10.1093/jb/mvq012. Epub 2010 Feb 9.
DNA in the environment is a source to mediate horizontal gene transfer (HGT). Present molecular cloning methods are based on this HGT principle. However, DNA in the extracellular environment, particularly with high molecular-weight, is thought to be prone to shearing or digestion by nucleases. Here we discovered that extracellular plasmid DNA released from lysed Escherichia coli remained intact and stable. Furthermore, it was demonstrated that plasmids up to 100 kb in size were taken up by co-present competent Bacillus subtilis cells. The detailed kinetics of the process together with sensitivity to added DNase I indicated that plasmid DNA released from lysed E. coli into the culture medium was stable enough for quantitative efficacy in the transformation of B. subtilis. Our results will be useful for the development of methods to transfer giant DNAs from general host E. coli without their biochemical purification.
环境中的 DNA 是介导水平基因转移 (HGT) 的来源。目前的分子克隆方法基于这一 HGT 原理。然而，细胞外环境中的 DNA，特别是具有高分子量的 DNA，容易被核酸酶剪切或消化。在这里，我们发现从裂解的大肠杆菌中释放的细胞外质粒 DNA 保持完整和稳定。此外，研究表明，大小达 100kb 的质粒被同时存在的感受态枯草芽孢杆菌细胞摄取。该过程的详细动力学以及对添加的 DNA 酶 I 的敏感性表明，从裂解的大肠杆菌释放到培养基中的质粒 DNA 足够稳定，可以定量有效地转化枯草芽孢杆菌。我们的研究结果将有助于开发不经过生化纯化从普通宿主大肠杆菌中转移巨型 DNA 的方法。
