Diamond Blackfan anemia (DBA) is predominantly an autosomal dominant inherited red cell aplasia primarily caused by pathogenic germline variants in ribosomal protein genes. DBA due to pathogenic RPL35A variants has been associated with large 3q29 deletions and phenotypes not common in DBA. We conducted a multi-institutional genotype-phenotype study of 45 patients with DBA associated with pathogenic RPL35A germline variants and curated the variant data on 21 additional cases from the literature. Genotype-phenotype analyses were conducted comparing patients with large deletions versus all other pathogenic variants in RPL35A. Twenty-two of the 45 cases had large deletions in RPL35A. After adjusting for multiple tests, a statistically significant association was observed between patients with a large deletion and steroid-resistant anemia, neutropenia, craniofacial abnormalities, chronic gastrointestinal problems, and intellectual disabilities (p<0.01) compared with all other pathogenic variants. Non-large deletion pathogenic variants were spread across RPL35A with no apparent hot spot and 56% of the individual family variants were observed more than once. In this, the largest known study of DBA patients with pathogenic RPL35A variants, we determined that patients with large deletions have a more severe phenotype that is clinically different from those with non-large deletion variants. Genes of interest also deleted in the 3q29 region that could be associated with some of these phenotypic features include LMLN and IQCG. Management of DBA due to large RPL35A deletions may be challenging due to complex problems and require comprehensive assessments by multiple specialists including immunologic, gastrointestinal, and developmental evaluations to provide optimal multidisciplinary care.