Cloning and expression of the 3' portion of the T4 denV gene as a lacZ fusion gene.
Polyclonal antibodies have been raised against endonuclease V from the bacteriophage T4. This rabbit serum, from which endemic E. coli antibodies have been removed, reacts with a single protein from T4-infected E. coli with a molecular weight of 16078 dalton. It was confirmed that these antibodies were directed against endonuclease V through the inhibition of the pyrimidine dimer specific nicking activity of endonuclease V in an in vitro nicking assay. A phage lambda gt11 T4 dC DNA library was screened for phage which produced a beta-galactosidase-endonuclease V fusion protein. Immunopositive clones were detected at a frequency of 0.25% of the plaques in the library. Restriction enzyme analyses of the DNA from 45 of these phage showed that all contained a 1.8 kb T4 EcoRI fragment which had been inserted within lambda gt11 in a single orientation. Western analysis of proteins which were produced from an induction of lysogens made from these phage reveals a single fusion protein band with a molecular weight slightly larger than native beta-galactosidase.
Lloyd
R S
RS
Augustine
M L
ML
D003001:Cloning, Molecular; D043224:Deoxyribonuclease (Pyrimidine Dimer); D004706:Endodeoxyribonucleases / Q000037:antagonists & inhibitors / Q000235:genetics* / Q000276:immunology; D005786:Gene Expression Regulation; D005822:Genetic Vectors; D007763:Lac Operon; D013604:T-Phages / Q000201:enzymology / Q000235:genetics*; D001616:beta-Galactosidase / Q000235:genetics
D004706:Endodeoxyribonucleases; D043224:Deoxyribonuclease (Pyrimidine Dimer); D001616:beta-Galactosidase