MicroRNAs are non-coding small RNAs that regulate gene expression by Watson-Crick base pairing to target messenger RNA (mRNA). They are involved in most biological and pathological processes, including tumorigenesis. The binding of microRNA to mRNA is critical for regulating the mRNA level and protein expression. However, this binding can be affected by single-nucleotide polymorphisms that can reside in the microRNA target site, which can either abolish existing binding sites or create illegitimate binding sites. Therefore, polymorphisms in microRNA can have a differing effect on gene and protein expression and represent another type of genetic variability that can influence the risk of certain human diseases. Different approaches have been used to predict and identify functional polymorphisms within microRNA-binding sites. The biological relevance of these polymorphisms in predicted microRNA-binding sites is beginning to be examined in large case-control studies.