Primer extension analysis of PRAT satellite transcripts.

(A) Single extension product is detected on reverse PRAT strand using RATZEB1 primer (line R). The size of the product of 97 nt is determined by comparison with the sequencing reaction of PRAT dimer obtained by the same RATZEB1 primer. (B) Several extension products are detected on the forward strand using primers RATZEB2 and RATZEB4, and their size is indicated (lines R). Products of 122 nt and 127 nt, obtained in a reactions with RATZEB2 and RATZEB4, respectively, correspond to the same start site. (C) Positioning of transcription start sites and polyadenylation sites on PRAT forward and reverse strand. Transcription start sites determined by primer extension are indicated by green bent arrows while those detected by 5′ RLM-RACE are shown by violet bent arrows. Dashed green arrow represents weaker start site corresponding to 108 nt band obtained in reaction with RATZEB2. The sizes of the products obtained in extension assays are indicated under the corresponding nucleotide positions. The lengths of the products from reactions with RATZEB2 are in brackets. Polyadenylation sites are indicated with asterisks. Straight arrows represent primers and are positioned under the sequence they anneal to. Position of putative polymerase II promoter predicted by a neural network method (Reese 2001) is marked in red.