The central dogma of molecular biology, formally postulated in 1957 by Francis Crick, proposes that the flow of genetic information in a typical organism proceeds from DNA, the genetic blueprint, to RNA, the messenger, which is then ultimately processed by ribosomes to produce a specific protein as the functional end-product. Over 60 years later, it is now recognized that RNA and DNA both play much more dynamic roles than originally proposed by Crick. For example, seminal discoveries in RNA interference and CRISPR technologies have paved the way for developing more efficient methodologies to understand how the gene expression affects cellular homeostasis, in order to potentially generate various pathological conditions. In this Special Issue, we aim to explore the numerous applications of nucleic acids in areas encompassing diagnostics, therapeutics, synthetic biology, and other exciting areas in the chemistry and biology of DNA and RNA. We are proud to have gathered leading experts in each of these fields so as to provide up-to-date reviews in these important areas.