Figure 3

Identification and characterization of an alternative Bach2 promoter. (A) Map of the 5' ends for the novel mRNAs determined by 5'RACE and primers employed in 5' RACE PCR and sequencing (1c, 1b, 1e, 1f), and in RT-PCR and Q-PCR (10F and 2R and/or 2F and 2R). Chromosomal locations refer to UCSC version 2006 February. (B) RT-PCR detection of transcripts including intron 2 sequences as a novel Bach2 exon. Tumor material from mouse 1206 was analyzed by RT-PCR using primers 2F and 2R (left panel) or 10F and 2R (right panel) and by ethdium bromide staining. (C) 5'-RACE analysis to detect novel Bach2 promoters. The products of the RACE PCR analysis were visualized by ethdium bromide staining. Arrows indicate the product bands. (D) Sequence around the alternative promoter. Exon sequences are in capital letters and position of mapped transcriptional start sites indicated by arrows. (E) RT-expression analysis of transcripts derived from the alternative Bach2 promoter. Tumor material with (marked by asterisk) or without Bach2 gene proviral integration were analyzed by RT-PCR with primers 10F and 2R and the products visualized by ethdium bromide staining. (F) Q-PCR analyses of transcripts with origin from the alternative Bach2 promoter. Analyses were done using primers 2F and 2R on tumors with (asterisked) or without proviral integration in Bach2 and spleen tissues from untreated NMRI mice (NMRI S). Expression levels were normalized to tbp and NMRI S with the expression level for NMRI S set to 20.