High-throughput mammalian two-hybrid screening for protein-protein interactions using transfected cell arrays-4

<p><b>Copyright information:</b></p><p>Taken from "High-throughput mammalian two-hybrid screening for protein-protein interactions using transfected cell arrays"</p><p>http://www.biomedcentral.com/1471-2164/9/68</p><p>BMC Genomics 2008;9():68-68.</p><p>Published online 6 Feb 2008</p><p>PMCID:PMC2254387.</p><p></p>ng the Gal4-pZsGreen reporter and plasmids coding for the known interacting p53 (pBD-p53) and SV40-T (pAD-SV40T) hybrid proteins (boxed, line B). As negative control (line A) Gal4-pZsGreen reporter was co-transfected with the non-interacting proteins p53 (pBD-p53) and TRAF (pAD-TRAF). The pBD-NF-κB control plasmid expressing the GAL4 DNA-binding domain fused to the transcription activation domain of NF-κB is used as a positive control to monitor transfection efficiency and reporter performance (line C). A construct expressing EGFP under control of a CMV promoter (line D) is typically printed as a frame at the periphery of the arrays in order to locate the arrayed features. Fig. 1a shows example images of different adherent cell lines transfected using identical microarray slides. : Transfection efficiency as reflected by the level of NF-κB induced reporter expression. Comparable results were obtained for PC-3, HEK293 and HeLa. Data shown are from representative experiments and represent mean fluorescence of 6 features per sample, collected from triplicate spots on two identical slides