Figure 1

The PCR amplification of AR cDNA (a), the identified intronic AR mutation (b), the schematic representation of the two aberrant mRNAs caused by the deep intronic mutation (c), and the nucleotide sequences of exons 6 and 7 and intron 6 (d). (a) The AR cDNA was PCR-amplified with primers on exon 5 and the 3′ UTR and the products were visualized on a 1.5% agarose gel. Lanes T1 and T2 are amplification products of the patient samples, XX31B and XY31A are control fibroblast samples, and LNCaP is from prostatic cancer cells. (b) The sequence chromatograms showing the intronic mutation which was confirmed to be present as hemizygotic in both patients and heterozygotic in the mother, whereas it was absent in the father and the healthy sister. (c) Schematic drawing of the aberrant AR pre-mRNA splicing leading to the two longer mRNAs. The cryptic exons are marked with Ψ, and the mutation site is marked with an asterisk. (d) On the left are shown the sequence chromatograms showing the borders of the cryptic exonic sequences and the normal exons derived from sequencing of the gel-extracted aberrant PCR products. On the right are shown the nucleotide sequences of exon 6, intron 6, and exon 7, including the c.2450-118A>G mutation marked in bold and in bigger font size. For splice variant 1, the predicted donor site motif is highlighted in red. For splice variant 2, the predicted SRSF1 binding motif is highlighted in green. The intronic sequences that are spliced into the mRNA are in uppercase and highlighted in yellow (or red/green for sequences that are part of a splicing motif) and correspond to the boxes marked with Ψ in panel C. Normal exonic sequences are in uppercase and are not highlighted, intronic sequences are in lowercase.