A simple, low-cost, and universal gold nanoparticle (AuNP) aggregation-induced photothermal biosensing platform has been developed for the first time and applied for the visual quantitative genetic detection using a common thermometer. By exploiting the photothermal effect of target-induced gold nanoparticle aggregation, visual quantitative biochemical analysis can be achieved by simply recording temperature signals using a common thermometer. Compared to conventional genetic testing methods, it is label- and amplification-free and can be completed in 40 min without the aid of any advanced analytical instruments. Mycobacterium tuberculosis (MTB) DNA was used as a model target to demonstrate the application of this photothermal biosensing platform. Although no costly instrument was used, high sensitivity and specificity were achieved with the limit of detection (LOD) of 0.28 nM, which was nearly 10-fold lower than that of the colorimetric method using a spectrometer. This AuNP aggregation-induced photothermal biosensing strategy provides a simple, low-cost, and universal platform for broad application of visual quantitative detection of nucleic acids and many other biomolecules, particularly in point-of-care (POC) biosensing applications.