The phenotypes associated with trisomy 21 at the organismal level are complex. Indeed, individuals with Down syndrome are born with varying severities and higher risks for several human diseases. Two main gene-centric hypotheses have been proposed to explain the biological consequences of trisomy 21. One is that increased expression of a particular gene and its downstream effectors cause the pathophysiology in Down syndrome. For example, increased expression of the amyloid-beta precursor protein (APP), the High Mobility Group Nucleosome Binding Domain 1 (HMGN1), or the transcription factor RUNX1 are associated with the increased incidence of early onset of Alzheimer’s disease, B cell acute lymphoblastic leukemia, or transient myeloproliferative disorder in individuals with Down syndrome, respectively. A second hypothesis is that abnormalities arise due to the indirect effects of increased gene activity on chromosome 21 that alter specific cellular processes. Examples include genes that regulate splicing (U2AF1L5, RBM1, and U2AF1), chromatin regulators (HMGN1 and BRWD1), secretory-endosomal functions (DOPEY2, CSTB, and SYNJ1), protein turnover (USP25),or metabolism (SOD1). Nonetheless, it has been challenging to prove that a third wild-type copy of a given gene on chromosome 21 is the sole driver of a particular disease. Indeed, analysis of human segmental trisomies of chromosome 21 provides evidence against the existence of a single chromosomal region contributing to Down syndrome phenotypes.