Sutherland P, McAlister-Henn L
J Bacteriol. 1985 Sep;163(3):1074-9. doi: 10.1128/jb.163.3.1074-1079.1985.
An oligodeoxynucleotide specific for a pentapeptide sequence corresponding to amino acid residues 32 through 36 of Escherichia coli malate dehydrogenase was chemically synthesized and used to identify the mdh gene on plasmid pLC32-38 from the Clarke-Carbon recombinant library. Cells transformed with this plasmid exhibited a 10-fold increase in malate dehydrogenase activity. A 1.2-kilobase PvuII fragment which hybridized with the oligodeoxynucleotide probe was subcloned, and the identity of the mdh structural gene was confirmed by partial nucleotide sequence analysis. The expression of the mdh gene, as measured by both Northern blotting and enzyme assays, was found to vary over a 20-fold range with different culture conditions.
化学合成了一种与大肠杆菌苹果酸脱氢酶第32至36位氨基酸残基对应的五肽序列特异的寡脱氧核苷酸，并用于从克拉克-卡尔文重组文库中鉴定质粒pLC32 - 38上的mdh基因。用该质粒转化的细胞中苹果酸脱氢酶活性增加了10倍。将与寡脱氧核苷酸探针杂交的1.2千碱基PvuII片段进行亚克隆，并通过部分核苷酸序列分析证实了mdh结构基因的身份。通过Northern印迹法和酶活性测定发现，mdh基因的表达在不同培养条件下变化范围达20倍。
