||How natural selection, the driving force of evolution first proposed by Charles Darwin, occurs in real ecosystem is a fundamental question in evolutionary biology. Classical theories in population genetics assume that adaptive evolution occurs when a beneficial mutation arises and then increases in frequency under constant selective pressure until it becomes fixed in the population. However, our empirical and theoretical analyses suggest that selective pressure on a beneficial variant is frequently not constant but negative frequency-dependent. First, when we detected the footprint of natural selection or “selective sweep” from DNA sequence variation in African populations of fruit flies (Drosophila melanogaster), we found that at many genes the same beneficial mutations arose to intermediate frequencies in multiple geographic regions, which cannot be explained unless the pressures of positive selection diminished as mutations’ frequencies increased. Mathematical analyses and computer simulations confirmed this result. Second, our mathematical model of seasonally fluctuating selection revealed that a variant conferring advantage in one season but disadvantage in the other season can be maintained in intermediate frequencies when the variant is partially protected from selection. This condition that leads to the emergence of negative frequency-dependent selection is called the “storage effect”. Computer simulations showed that this effect can maintain the high levels of phenotype-changing genetic variation, the fundamental ingredient of adaptive biological evolution, at many loci in the genome.