When calculating the melting temperature (Tm) of a short probe, which formula is used?

• A. Tm = (2C x GC pairs) + (4C x AT pairs)
• B. Tm = (4C x GC pairs) x (2C x AT pairs)
• C. Tm = (2C x GC pairs) - (4C x AT pairs)
• D. Tm = 5C + probe length

Answer: (A)

Short DNA probes are often estimated using a simple additive rule: each base pair type contributes a fixed amount to the melting temperature, and you add those contributions together. The form shown uses 2°C for each GC pair and 4°C for each AT pair, then sums them. To use it, count how many GC pairs there are and multiply by 2°C, count how many AT pairs there are and multiply by 4°C, and add the results. This gives a quick estimate of Tm that reflects why GC-rich sequences tend to be more stable than AT-rich ones. For example, a probe with 3 GC pairs and 4 AT pairs would have an estimated Tm of (2×3) + (4×4) = 6 + 16 = 22°C. The other options either multiply counts, subtract, or add a term not tied to base composition, so they don’t provide a valid quick estimate for short probes.