Quantitative assay of recombinant vaccinia virus-encoded neomycin phosphotransferase in infected eukaryotic cell lysates.
A method for the detection and quantitation of neomycin phosphotransferase (NPT II) activity in recombinant vaccinia virus (VV)-infected eukaryotic cell lysates is described. The assay is linear with respect to both protein concentration and time of incubation. Cytoplasmic extracts of cells infected with a recombinant VV expressing the bacterial neo gene exhibited NPT II levels more than 50-fold higher than those detected in extracts from either uninfected or VV-infected cells. These results indicate that interference from cellular or viral-induced ATPase activities is sufficiently low that NPT II enzyme activity can be measured in crude cell lysates without employing additional protein purification procedures.
D000123:Acetyltransferases / Q000032:analysis; D000251:Adenosine Triphosphatases / Q000378:metabolism; D002478:Cells, Cultured; D015500:Chloramphenicol O-Acetyltransferase; D005786:Gene Expression Regulation; D019868:Kanamycin Kinase; D010770:Phosphotransferases / Q000032:analysis* / Q000235:genetics; D011994:Recombinant Proteins / Q000032:analysis*; D014616:Vaccinia virus / Q000235:genetics*
D011994:Recombinant Proteins; D000123:Acetyltransferases; D015500:Chloramphenicol O-Acetyltransferase; D010770:Phosphotransferases; D019868:Kanamycin Kinase; D000251:Adenosine Triphosphatases