Transient transfection analysis A. The sequence of the wild-type hTERC core promoter (-107/+69) is shown at the top. The name of each mutant construct is indicated on the left hand side. The number on either side of the sequence is related to the transcriptional start site. Dashes indicate an identical sequence to wild-type. Mutated nucleotides are shown below the wild type sequence. B. Scanning mutational analysis Sp1 sites in the hTERC core promoter Promoter activities of site-replaced mutant constructs: The various symbols or circles represent the different transcription factor binding sites indicated at the top. Transcriptional start site indicated as broken line. The constructs are shown with a black ellipsoid shape indicating a site-replaced mutation in one or more positions and open ellipsoids representing unmodified sites. The promoter activity is shown on the right hand side. Three micrograms of each plasmid were used for transient transfection analysis in 5637 cells. Promoter activities of the mutant constructs were assayed by transfection and compared to the wild-type promoter. The pRL-SV40 vector was used as an internal control to normalise the transfection efficiency. For each transfection the mean and standard deviation of data from three experiments are shown. C. Mutation of Sp1.2 site from PNH patient in 117 bp in length of minimum hTERC promoter increases promoter activity.