Figure 1

Promoter trapping with the hTERT promoter yields a transcriptionally active complex. A. Nucleotide sequence of the hTERT gene regulatory region. Features of known binding regions are noted, such as two E-Boxes, AP2, and SP1 sites. The translational start site (INR), nucleotide +1, is denoted by the bold arrow. B. Full hTERT promoter containing single stranded (GT)₅ tails complexed with TF from HEK293 NE. hTERT specific TF are purified by annealing (GT)₅ tails to a (CA)₅-Sepharose column and eluted with a high salt buffer. C. Transcription Assay of hTERT Promoter Trap Elute. After RNA was transcribed by the promoter trapping eluate, 0.1 pmol ³²P-labeled oligo (5'-cggagcgcgcggcatcgcgg-3') was extended with reverse transcriptase and analyzed on a 6% polyacrylamide gel containing 8 M urea. Visualization was accomplished with autoradiography. (+) Indicates the use of rNTP and (-) denotes where rNTP was not used during transcription, as a negative control. The expected product is 100 base pairs.