Genotype/phenotype analysis in patient DBA019. (A) RPL35A mutational analysis in this family showed 2 first-degree relatives, a father and sister, with the heterozygous RPL35A 97 G>A mutation. Both of these patients had macrocytic anemia suggestive of subclinical DBA carriers. Hemoglobin (Hb), mean red cell volume (MCV), and eADA activity are indicated; the hemoglobin value indicated for the proband is during treatment with steroids. Erythrocyte ADA was normal in all tested members of this pedigree. Black shading represents the DBA proband; white shading, hematologically unaffected members of the paternal lineage with normal RPL35A sequence; diagonal lines, deceased members with unknown RPL35A status; cross-hatched symbols, clinically normal persons who were not tested for RPL35A mutations. (B) 2 RPL35A RNA products were amplified from patient DBA019 using RT-PCR with primers designed to amplify the full-length RPL35A message. The shorter product results from an alternative splicing event between exons 3 and 4, leading to a truncated protein. The arrows above the transcript diagram show the normal splicing event, leading to wild-type RPL35A and removal of the approximately 2691 nucleotide intron. The lines above exon 3 indicate the wild-type codon sequence within exon 3; the 97 G/A mutation is indicated in gray. The arrows below the line demonstrate the abnormal splicing event. The 97 G>A mutation results in selection of a cryptic splice donor site within exon 3 immediately upstream of the change, causing removal of 70 base pairs of 3′ exon 3 coding sequence in addition to the intron. The predicted amino acid sequence of wild-type, simple amino-acid substitution, and the splicing variant are shown below.