Transfection of keratinocytes abrogates detectable DNA-binding activity of CCAAT displacement protein.
Transfection of keratinocytes with plasmid DNA leads to the loss of detectable DNA-binding activity of CCAAT displacement protein but not of Yin Yang 1, as monitored by electrophoretic mobility shift assay. This phenomenon was found to be attributable to the presence of plasmid DNA in the nuclear extracts prepared from transfected cells. Treatment of these nuclear extracts with DNase I restored the ability to monitor DNA-binding activity of CDP. This report documents a new pitfall associated with transfection.
Narahari
Janaki
J
Department of Microbiology and Immunology, Indiana University School of Medicine, and Walther Cancer Institute, Indianapolis, Indiana 46202-5120, USA.
Roman
Ann
A
D001667:Binding, Competitive; D002457:Cell Extracts; D002474:Cell-Free System; D002478:Cells, Cultured / Q000187:drug effects / Q000378:metabolism; D004247:DNA / Q000378:metabolism*; D004274:DNA, Recombinant / Q000494:pharmacology*; D004268:DNA-Binding Proteins / Q000378:metabolism; D003850:Deoxyribonuclease I / Q000494:pharmacology; D024202:Electrophoretic Mobility Shift Assay; D050981:Erythroid-Specific DNA-Binding Factors; D006367:HeLa Cells / Q000187:drug effects / Q000378:metabolism; D018398:Homeodomain Proteins; D006801:Humans; D015603:Keratinocytes / Q000187:drug effects / Q000378:metabolism*; D008297:Male; D009687:Nuclear Proteins / Q000037:antagonists & inhibitors* / Q000378:metabolism; D010957:Plasmids / Q000235:genetics; D011485:Protein Binding / Q000187:drug effects; D012097:Repressor Proteins / Q000037:antagonists & inhibitors* / Q000378:metabolism; D014157:Transcription Factors / Q000378:metabolism; D014158:Transcription, Genetic / Q000187:drug effects; D014162:Transfection; D050986:YY1 Transcription Factor
C080632:CUX1 protein, human; D002457:Cell Extracts; D004274:DNA, Recombinant; D004268:DNA-Binding Proteins; D050981:Erythroid-Specific DNA-Binding Factors; D018398:Homeodomain Proteins; D009687:Nuclear Proteins; D012097:Repressor Proteins; D014157:Transcription Factors; D050986:YY1 Transcription Factor; D004247:DNA; D003850:Deoxyribonuclease I