miRNAs have emerged as key regulators of gene expression in both plants and animals. These small (generally 21–22 nt) RNA molecules, originated from primary “hairpin” transcripts, can induce translational suppression or direct mRNA cleavage. Similar to regular mRNAs, the expression of miRNAs is highly regulated. Their expression pattern could provide critical clues to understanding miRNA functions. However, many previously identified miRNA families have multiple paralogous loci. Within each family, different members are often closely related and sometimes give rise to identical miRNAs. This poses critical challenges in the analysis of individual miRNA genes. This chapter describes several methods that are commonly used for miRNA expression analysis, including high-throughput sequencing, microarrays, and briefly discusses qRT-PCR, northern blotting, and other approaches used for data validation.