Transcription Initiation Activity Sets Replication Origin Efficiency in Mammalian Cells
Figure 2
Sensitivity of the ORI identification method.
Array profiles and nascent strand abundance measurements by Q-PCR of 18 positive regions located at 5′ends of genes (A), at less than 200 bp of exons (B), including one at the 3′ UTR of two genes of convergent transcription (ORI 67065), or at intergenic zones (C). Similar analysis was performed for 3 negative regions (D). The maps above each graph show the annotated genomic features and probe distribution of the regions analysed. Blue and red rectangles indicate exons transcribed from the upper or the lower strand, respectively, and black arrows show the position of the major annotated TSS. Grey rectangles represent array probes. The red dashed line depicts the threshold of the array duplicates. Q-PCR experiments were carried out in duplicate in at least two independent preparations of 300–800 nt long nascent strands and values were normalised to the flanking primer pair detecting the lowest amount of nascent strands at each region. Standard deviation bars are indicated. Primer pairs were designed to amplify across the array probes in all possible cases and their sequences are shown in Table S3. ORI 67276 corresponds to the CpG island region of the Mecp2 gene.