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Biology · পরিভাষা

Denaturation, melting temperature কী?

অন্য নাম: denaturation (DNA) · melting temperature · hybridisation

Definition 11.9 University Biology — Year 1 · অধ্যায় 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 TmT_m. It is followed by the hyperchromic effect: single strands absorb about 40 %40\,\% more ultraviolet light at 260 nm260\,\mathrm{nm} than the same bases stacked in a helix. TmT_m rises with the G+CG + C content (0.4 ∘C0.4\,{}^{\circ}\mathrm{C} per percent), with the salt concentration, and with length; in 0.15 mol/L0.15\,\mathrm{mol}/\mathrm{L} salt a long DNA of 50 %50\,\% G+CG + C melts near 90 ∘C90\,{}^{\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 %40\,\% as the strands separate, over a few degrees; the midpoint, TmT_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.
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.

উদাহরণ

Example 11.11 (A probe finds a gene)

A twenty-nucleotide probe complementary to one gene, at 5 ∘C5\,{}^{\circ}\mathrm{C} below its own TmT_m, binds its exact complement and nothing else in three billion base pairs: a single mismatch in twenty lowers TmT_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.

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