(Top) Gene for the A subunit and mutations identified. Exons are indicated by wide vertical bars and Roman numerals, and introns are indicated by capital letters. Each exon and its boundaries and the 5′-flanking regions were amplified one by one using 17 pairs of primers (arrows under exons). Solid and open circles with sequences represent homozygous causative mutations and changes known as common polymorphisms, respectively. Normal sequences are followed by those corresponding to substitutions found in the probands' DNAs. For analysis of the two mutations identified, two amplified fragments were digested with appropriate endonucleases as indicated by their names and arrows. (Bottom) mRNAs for the A subunit in the probands. Three regions of exons II-IV, X-XIII, and XI-XIII were amplified by RT-PCR. A solid line represents the expressed mRNA, whereas a broken line indicates a possible mRNA that was not detected.