Duplex-specific nuclease (DSN) from hepatopancreas of the craboid Paralithodes camtschaticus (red king crab) has a unique combination of properties. Along with thermal stability and a high optimal temperature of catalysis, this enzyme exhibits high substrate selectivity, cleaving only DNA in duplexes (DNA-DNA or DNA-RNA). Accordingly, it digests neither single strands (nor single-stranded regions) of DNA, nor RNA strands with any secondary structure. Such properties make it possible to create unique protocols based on DSN, which is also an important object of fundamental research in the field of nuclease evolution. The review considers diverse applications of the red king crab DSN in modern methods of molecular biology.