One of the large, unsolved problems in human genetics is the proportion of functional sequences in genomes. Recently, the encyclopedia of DNA elements consortium revealed that the majority of the genome is biochemically active, which were described as biochemical functions. This has been used as evidence to pronounce the death of the junk DNA concept. In evolutionary biology, junk DNAs are sequences whose gain or loss does not seriously affect fitness of the host organism. In the human genome, a large amount of biochemical activity should be to repress the sequences so as to avoid their harmful expression. The biochemical activity is very different from functionality in the light of evolution. The single nucleotide polymorphism sites associated with disease and other phenotypes may be functional, but their abundance in the active genome regions is not reliable evidence of functionality. Because of sequence-independent functions, the proportion of functional regions would be underestimated when sequence constraints are used alone. Knockout may be the most effective means of distinguishing functional sequences from junk DNA. (C) 2012 Elsevier Inc. All rights reserved.