Figure 3

Exon trapping analysis. (A) Structure of the exon trapping vector pSPL3 containing the BRCA2 exon 21 and 449 bp of intron 20 and 408 bp of intron 21, respectively containing the wild-type or the nucleotide 8982+1 G→A/c.8754+1 G→A variant. (B) COS-7 cells were transfected with pSPL3-BRCA2-exon 21 wild-type or pSPL3-BRCA2-exon 21 mutant plasmids. Total RNA was isolated, RT-PCR analysis was performed and the PCR products (in duplicates) were resolved on a 2% agarose gel. The 299 bp product corresponds to wild-type exon 21 (unaltered splicing), while the 345 bp product corresponds to exon 21 and the inclusion of 46 bp of intron 21. The sizes of the DNA marker are indicated to the right. (C) Sequence of exon 21 (345 bp band). The nucleotide 8982+1 G→A/c.8754+1 G→A mutation and the TAA stop codon are underlined (sense strand).