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Fig. 7 | BMC Genomics

Fig. 7

From: The Trypanosomatid Pr77-hallmark contains a downstream core promoter element essential for transcription activity of the Trypanosoma cruzi L1Tc retrotransposon

Fig. 7

Binding analysis of T. cruzi nuclear proteins to DPE-bearing DNA as double strand DNA and single strand DPE-bearing oligos (sense and antisense) in native and heat denatured conditions. a DNA corresponding to the double-stranded DPE-bearing Pr77 sequence (dsDPE, nucleotides 12 to 33 of Pr77), single stranded sense DPE oligo (sDPE), single stranded antisense DPE oligo (asDPE) and the heat denatured sDPE and asDPE oligos, was 5′ end radiolabelled with γ-ATP32 and 80.000 cpm of each radiolabeled probe loaded onto native 12 % polyacrylamide gels with (+) or without (−) incubation with T. cruzi nuclear proteins (NP). The mobility of the highest amount of radiolabelled free probe is indicated as free Probe (FP). Asterisks indicate the shifted bands that are formed when nuclear proteins are incubated with both 32P- single stranded sense DPE and heat denatured 32P- single stranded sense DPE. b Binding of nuclear proteins to the Pr77 sequence and competition assays with DPE-bearing DNAs. Double-stranded DNA corresponding to the entire Pr77 sequence (dsPr77 probe) was 5′ end radiolabelled with γ-ATP32 and loaded onto native 6 % polyacrylamide gels with incubation (+) or without (−) incubation with T. cruzi nuclear proteins (NP). Competition reactions were performed by adding to the reaction non-labelled DNAs corresponding to the dsDPE sequence, the single stranded DPE oligo (sDPE) or the single stranded DPE antisense oligo (asDPE) as competitors (C) at ratios of 1:10. Shifted bands are indicated by black arrows on both sides of the gel (). Gel wells are indicated as ‘w

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