What is Denaturation, melting temperature? — Biology

Definition 11.9 University Biology — Year 1 · Chapter 11 — Nucleotides and Nucleic Acids

Heating, or extreme pH, separates the two strands of DNA: denaturation (melting), a cooperative transition over a few degrees, whose midpoint is the melting temperature $T_m$ . It is followed by the hyperchromic effect: single strands absorb about $40\,\%$ more ultraviolet light at $260\,\mathrm{nm}$ than the same bases stacked in a helix. $T_m$ rises with the $G + C$ content ( $0.4\,{}^{\circ}\mathrm{C}$ per percent), with the salt concentration, and with length; in $0.15\,\mathrm{mol}/\mathrm{L}$ salt a long DNA of $50\,\%$ $G + C$ melts near $90\,{}^{\circ}\mathrm{C}$ . Cooled slowly, the strands find their complements and re-form the helix: renaturation, or hybridisation when the strands come from different sources.

Melting curves of two DNAs in the same salt. The absorbance rises by 40\,\% as the strands separate, over a few degrees; the midpoint, T_m, is higher for the DNA richer in G–C pairs. — Melting curves of two DNAs in the same salt. The absorbance rises by $40\,\%$ as the strands separate, over a few degrees; the midpoint, $T_m$ , is higher for the DNA richer in G–C pairs.

DNA precipitated by cold ethanol from a cell lysate and spooled onto a glass rod: a few milligrams of the molecule, visible as fibres because each is millions of base pairs long.

Examples

Example 11.11 (A probe finds a gene)

A twenty-nucleotide probe complementary to one gene, at $5\,{}^{\circ}\mathrm{C}$ below its own $T_m$ , binds its exact complement and nothing else in three billion base pairs: a single mismatch in twenty lowers $T_m$ by several degrees and the mismatched hybrid melts. The specificity of base pairing, multiplied over twenty positions, is what every DNA test, from paternity to pathogen detection, relies on.

Read in context →

Examples

Related terms