Figure 6

Regulatory elements in Exon 7 and flanking intronic sequences are important for controlling CEACAM1 alternative splicing. (A) The DUP5-1 minigene shown here contains three exons and two introns derived from human β-globin gene and is essentially identical to previously described version [46] except that vector backbone is pcDNA 3.1/Myc-His(-) C. The middle exon is a chimera of β-globin exon 1 and 2. The name of each minigene and exon and intron sizes are indicated. Minigene βg-ΔiE7-βg is a derivative of DUP5-1 in which the middle exon 2 is replaced by exon 7 of CEACAM1. Minigene βg-E7-βg was created by digesting pDUP5-1 with ApaI and BglII and inserting PCR amplified CEACAM1 fragment that contains exon 7 and the flanking introns 6 and 7 (thick line) except first 150 bp in intron 6 and last 94 bp in intron 7. (B and C) RT-PCR analysis of MDA-MB468 and ZR75 cells transfected with the indicated minigenes or empty vector. -RT represents omission of reverse transcriptase during cDNA synthesis. The data shown is representative of three independent experiments. Bar diagrams represent the mean ± SD of at least three independent experiments. *P < 0.001 versus DUP5-1.