Analyses of spontaneous mutations of cloned gene 49 of phage T4.
Holliday structure resolving enzyme endonuclease VII (endo VII) of phage T4 is highly toxic for E. coli when expressed outside of the phage infection environment. As a consequence, plasmids with a mutated gene 49, the gene which encodes for endo VII, can be easily isolated and characterised. We have isolated and characterised 400 survivors from independent transformations with a plasmid carrying gene 49 under the control of the T7 promoter. The majority had mutated gene 49 by IS10 insertions which almost exclusively mapped to a distinct site. When this site was mutated other insertion sites were observed as well as an increase in other mutational events including large deletions. Neither of the observed insertion sites mapped matched the consensus IS10 sequence completely. Additionally when the level of expression of gene 49 was altered the distribution of mutations was changed suggesting that other elements apart from the target sequence are necessary for determining IS10 insertion. The expression of gene 49 in E. coli provides a particularly useful tool for the analysis of mutational events.
Hartung
M
M
Institute for Genetics of the University of Cologne, Zülpicher Strasse 47, 50674 Köln, Germany.
Slack
M
M
Kemper
B
B
D017122:Bacteriophage T4 / Q000235:genetics*; D004252:DNA Mutational Analysis; D004251:DNA Transposable Elements / Q000235:genetics; D004274:DNA, Recombinant; D004706:Endodeoxyribonucleases / Q000235:genetics*; D016254:Mutagenesis, Insertional; D009154:Mutation; D009690:Nucleic Acid Conformation; D010957:Plasmids; D017354:Point Mutation; D017384:Sequence Deletion; D014170:Transformation, Genetic
D004251:DNA Transposable Elements; D004274:DNA, Recombinant; D004706:Endodeoxyribonucleases; C035571:endodeoxyribonuclease VII