用于枯草芽孢杆菌分子克隆的新型质粒标记拯救转化系统，可直接筛选重组体。

Novel plasmid marker rescue transformation system for molecular cloning in Bacillus subtilis enabling direct selection of recombinants.

作者信息

Haima P, Bron S, Venema G

机构信息

Department of Microbiology, Center of Biological Sciences, Haren Groningen, The Netherlands.

出版信息

Mol Gen Genet. 1990 Sep;223(2):185-91. doi: 10.1007/BF00265052.

DOI:10.1007/BF00265052

PMID:2123518

Abstract

A versatile plasmid marker rescue transformation system was developed for homology-facilitated cloning in Bacillus subtilis. It is based on the highly efficient host-vector system 6GM15-pHPS9, which allows the direct selection of recombinants by means of beta-galactosidase alpha-complementation. The system offers several advantages over previously described cloning systems: (1) the convenient direct selection of recombinants; (2) the ability to effectively transform B. subtilis competent cells with plasmid monomers, which allows the forced cloning of DNA fragments with high efficiency; (3) the availability of 6 unique target sites, which can be used for direct clone selection, SphI, NdeI, NheI, BamHI, SmaI and EcoRI; and (4) the rapid segregational loss of the helper plasmid from the transformed cells.

摘要

开发了一种通用的质粒标记拯救转化系统，用于枯草芽孢杆菌中同源性促进克隆。它基于高效的宿主-载体系统6GM15-pHPS9，该系统允许通过β-半乳糖苷酶α-互补直接选择重组体。与先前描述的克隆系统相比，该系统具有几个优点：（1）方便地直接选择重组体；（2）能够用质粒单体有效地转化枯草芽孢杆菌感受态细胞，这允许高效地强制克隆DNA片段；（3）有6个独特的靶位点可供使用，可用于直接克隆选择，即SphI、NdeI、NheI、BamHI、SmaI和EcoRI；（4）辅助质粒从转化细胞中快速分离丢失。