Noël G, Drapeau G R
J Bacteriol. 1986 Feb;165(2):399-404. doi: 10.1128/jb.165.2.399-404.1986.
To facilitate the analysis of the cell division control apparatus in Escherichia coli, we studied extragenic suppressor mutations of a previously characterized temperature-sensitive division mutation, ftsM1. Cells of strain GD40 which harbor this mutation were spread on agar plates and incubated at 42 degrees C, and the surviving cells were analyzed for the presence of a suppressor mutation. One group of suppressed mutants had acquired a new mutation which, by conjugation, was found to be located in the 30- to 40-min region of the E. coli genetic map. The other group comprised revertants carrying a suppressor which appeared to map between thr and leu. This suppressor gene, called sftA, was cloned with a mini-Mu-derived in vivo cloning system by selection for suppression of temperature sensitivity in GD40 cells. Subsequent subcloning of a fragment of the chromosomal DNA from the mini-Mu plasmid into pBR325 resulted in the delineation of the suppressor gene on a 1.8-kilobase XhoI-PvuI fragment. A strain, CV514, which does not express the temperature sensitivity phenotype of the ftsM1 mutation, was found to harbor a natural suppressor of this mutation. UV sensitivity, another known phenotype of the ftsM1 mutation, was also corrected by the presence of the sftA suppressor in the cell. Thus, the characterization of extragenic suppressors may allow the identification of new genes involved in the control of cell division.
为便于分析大肠杆菌中的细胞分裂控制机制，我们研究了先前已鉴定的温度敏感型分裂突变ftsM1的基因外抑制突变。将携带此突变的GD40菌株细胞铺在琼脂平板上，于42℃培养，对存活细胞分析其是否存在抑制突变。一组被抑制的突变体获得了一个新突变，通过接合实验发现该突变位于大肠杆菌遗传图谱的30至40分钟区域。另一组包括携带一个似乎位于thr和leu之间的抑制子的回复突变体。这个称为sftA的抑制基因，通过选择抑制GD40细胞的温度敏感性，利用一个源自mini-Mu的体内克隆系统进行克隆。随后将来自mini-Mu质粒的染色体DNA片段亚克隆到pBR325中，从而在一个1.8千碱基的XhoI - PvuI片段上确定了抑制基因。发现不表达ftsM1突变温度敏感性表型的CV514菌株含有该突变的天然抑制子。ftsM1突变的另一个已知表型紫外线敏感性，也因细胞中存在sftA抑制子而得到纠正。因此，对基因外抑制子的特性分析可能有助于鉴定参与细胞分裂控制的新基因。
