dehybridization

Dehybridization, in molecular biology, is the process by which a double-stranded nucleic acid duplex dissociates into two single strands, reversing the action of hybridization. It is commonly achieved by heating (thermal denaturation) or by exposure to chemical denaturants and is a fundamental step in many laboratory techniques that require access to individual strands.

Mechanism and determinants: Dehybridization disrupts hydrogen bonding between complementary bases and the base-stacking interactions that stabilize the duplex. The rate and extent depend on sequence composition, length, and temperature. The melting temperature (Tm) is the point where half of the duplexes are denatured. High GC content, longer sequences, and lower salt concentration raise Tm; alkali, urea, or formamide lower it. Denaturation can be partial or complete, and it is typically reversible under appropriate rehybridization conditions.

Applications and methods: In PCR and RT-PCR, a denaturation step separates strands for primer annealing. In blotting, sequencing, and microarray workflows, dehybridization permits probe-target dissociation or strand release. It is also used in nucleic acid hybridization assays, in situ hybridization, and gene detection methods. Rehybridization or annealing can occur if conditions are re-established, enabling probe reassociation or reformation of duplexes.

Terminology and related concepts: Dehybridization is sometimes referred to as denaturation or melting. It can involve thermal, chemical, or mechanical methods. In broader contexts, analogous processes disrupt duplex or duplex-like nucleic acid structures in RNA folding or protein–DNA interactions. See also: denaturation, DNA melting, hybridization.