Wu D G, Black L W
Department of Biological Chemistry, University of Maryland School of Medicine, Baltimore 21201.
J Mol Biol. 1987 Jun 20;195(4):769-83. doi: 10.1016/0022-2836(87)90483-9.
Bacteriophage T4 mutants hyperproducing gene 17 protein (Hp17) have been isolated at high frequency by growing gene 17 amber mutants on ochre suppressor strains of Escherichia coli. Most mutants showed the co-hyperproduction of gene 18 protein, although one anomalous mutant hyperproduced a 60,000 Mr partial polypeptide of gene 18. Hybridization of T4 late RNAs to cloned plasmid DNA containing genes 17, 18 or control T4 genes revealed that approximately five times more gene 17 mRNA and two to three times more gene 18 mRNA were synthesized in the Hp17 mutant infections. DNA-DNA hybridizations showed that Hp17 mutant DNA contained two to three times more copies of genes 17 and 18 than wild-type DNA. Southern blot analysis suggested that Hp17 mutants carry a direct tandem repeat of the gene 17-18 region, with variable copy number from one to at least six copies. Hyperproduction of gene 17 and 18 proteins appears therefore to result from gene amplification specific to the gene 17-18 region. In contrast to gene duplications reported in lambda and T4 phage, and numerous cases of gene amplification in bacteria, a similar or identical novel junctional fragment created by the amplification event was observed among 28 independent T4 Hp17 isolates; therefore, the mechanism giving gise to amplified sequences may involve specific sequences in this region of the T4 genome.
通过在大肠杆菌的赭石抑制菌株上培养基因17琥珀突变体，已高频分离出超量产生基因17蛋白（Hp17）的噬菌体T4突变体。大多数突变体显示基因18蛋白也共同超量产生，尽管有一个异常突变体超量产生了基因18的一个60,000道尔顿的部分多肽。将T4晚期RNA与含有基因17、18或对照T4基因的克隆质粒DNA杂交，结果表明在Hp17突变体感染中合成的基因17 mRNA大约多五倍，基因18 mRNA多两到三倍。DNA-DNA杂交显示，Hp17突变体DNA中基因17和18的拷贝数比野生型DNA多两到三倍。Southern印迹分析表明，Hp17突变体携带基因17 - 18区域的直接串联重复，拷贝数从一到至少六个不等，呈可变状态。因此，基因17和18蛋白的超量产生似乎是由基因17 - 18区域特有的基因扩增导致的。与λ噬菌体和T4噬菌体中报道的基因重复以及细菌中大量的基因扩增情况不同，在28个独立的T4 Hp17分离株中观察到了由扩增事件产生的相似或相同的新型连接片段；因此，产生扩增序列的机制可能涉及T4基因组该区域的特定序列。
