The study presents an analysis of constrained elements in the mammalian genome using a multiple sequence alignment of 29 mammalian species, capturing approximately 3.9 neutral substitutions per site and spanning about 1.9 million base pairs (Mbp) of the human genome. The authors identify constrained elements ranging from 3 bp to over 1 kilobase (kb) in length, covering about 5.5% of the human locus. They find that the total amount of nonexonic constraint is roughly twice that of exonic constraint. Constrained elements tend to cluster and overlap with known functional elements, such as repeats and exons. The density of constrained elements inversely correlates with mobile element density, and some constrained elements overlap with mammalian ancestral repeats. The study also describes a number of elements that have undergone intense purifying selection throughout mammalian evolution, more numerous than previously thought. These findings are obtained using Genomic Evolutionary Rate Profiling (GERP), a statistically rigorous and biologically transparent framework for identifying constrained elements. GERP identifies regions with nucleotide substitution deficits, measured as "rejected substitutions," which reflect the intensity of past purifying selection. The authors anticipate that GERP and similar analyses will provide further insights and improved annotation for the human genome as more mammalian genome sequence data becomes available.The study presents an analysis of constrained elements in the mammalian genome using a multiple sequence alignment of 29 mammalian species, capturing approximately 3.9 neutral substitutions per site and spanning about 1.9 million base pairs (Mbp) of the human genome. The authors identify constrained elements ranging from 3 bp to over 1 kilobase (kb) in length, covering about 5.5% of the human locus. They find that the total amount of nonexonic constraint is roughly twice that of exonic constraint. Constrained elements tend to cluster and overlap with known functional elements, such as repeats and exons. The density of constrained elements inversely correlates with mobile element density, and some constrained elements overlap with mammalian ancestral repeats. The study also describes a number of elements that have undergone intense purifying selection throughout mammalian evolution, more numerous than previously thought. These findings are obtained using Genomic Evolutionary Rate Profiling (GERP), a statistically rigorous and biologically transparent framework for identifying constrained elements. GERP identifies regions with nucleotide substitution deficits, measured as "rejected substitutions," which reflect the intensity of past purifying selection. The authors anticipate that GERP and similar analyses will provide further insights and improved annotation for the human genome as more mammalian genome sequence data becomes available.