Silhavy T J, Boos W
J Bacteriol. 1974 Oct;120(1):424-32. doi: 10.1128/jb.120.1.424-432.1974.
A procedure has been devised that allows selection of mutants defective in the beta-methylgalactoside transport system (mgl) of Escherichia coli. This procedure utilizes the compound 2R-glyceryl-beta-d-galactopyranoside (glycerylgalactoside), which is known to be transported by only two transport system in E. coli, namely, the lactose and the beta-methylgalactoside transport systems. Mutants lacking glycerol-3-phosphate dehydrogenase (glpD) are sensitive to glycerol. Similarly, mutants lacking uridine diphosphate-galactose-4-epimerase (galE) are sensitive to galactose. Glycerylgalactoside is an inducer of the lactose operon and also a substrate for beta-galactosidase. Thus, a mgl(+)glpD galE lacY strain will not grow in the presence of glycerylgalactoside owing to accumulated glycerol-3-phosphate, galactose-1-phosphate, and uridine diphosphate-galactose. We have constructed such a strain and shown that mgl mutants can be obtained by selecting for those that grow in the presence of glycerylgalactoside.
已设计出一种方法,可用于筛选大肠杆菌β-甲基半乳糖苷转运系统(mgl)存在缺陷的突变体。该方法利用化合物2R-甘油-β-D-吡喃半乳糖苷(甘油半乳糖苷),已知它在大肠杆菌中仅通过两种转运系统进行转运,即乳糖和β-甲基半乳糖苷转运系统。缺乏甘油-3-磷酸脱氢酶(glpD)的突变体对甘油敏感。同样,缺乏尿苷二磷酸-半乳糖-4-差向异构酶(galE)的突变体对半乳糖敏感。甘油半乳糖苷是乳糖操纵子的诱导剂,也是β-半乳糖苷酶的底物。因此,由于积累了甘油-3-磷酸、半乳糖-1-磷酸和尿苷二磷酸-半乳糖,mgl(+)glpD galE lacY菌株在甘油半乳糖苷存在的情况下无法生长。我们构建了这样一个菌株,并表明通过选择那些在甘油半乳糖苷存在下生长的突变体,可以获得mgl突变体。