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Maize is a low-temperature (LT)-sensitive plant and its physiological responses towards LT of temperate regions developed is an adaptive trait. To further our understanding about the response of maize to LT at the physiological and photosynthesis level, we conducted Infrared Gas Analysis (IRGA using LICOR6400-XT in 45-day-old grown two maize genotypes, one from temperate region (Gurez-Kashmir Himalayas), viz., Gurez local (Gz local), and another from tropics (Gujarat), viz., GM6. This study was carried out to evaluate the underlying physiological mechanisms in the two differentially temperature-tolerant maize genotypes. Net photosynthetic rate (A/PN), 18.253 in Gz local and 25.587 (µmol CO2 m−2 s−1) in GM6; leaf conductance (gs), 0.0102 in Gz local and 0.0566 (mmol H2O m−2 s−1) in GM6; transpiration rate (E), 0.5371 in Gz local and 2.9409 (mmol H2O m−2 s−1) in GM6; and water use efficiency (WUE), 33.9852 in Gz local and 8.7224 (µmol CO2 mmol H2O−1) in GM6, were recorded under ambient conditions. Also, photochemical efficiency of photosystem II (PSII) (Fv/Fm), 0.675 in Gz local and 0.705 in GM6; maximum photochemical efficiency (Fv′/Fm′), 0.310234 in Gz local and 0.401391 in GM6; photochemical quenching (qP), 0.2375 in Gz local and 0.2609 in GM6; non-photochemical quenching (NPQ), 2.0036 in Gz local and 1.1686 in GM6; effective yield of PSII (ФPSII), 0.0789 in Gz local and 0.099 in GM6; and electron transport rate (ETR), 55.3152 in Gz local and 68.112 in GM6, were also evaluated in addition to various response curves, like light intensities and temperature. We observed that light response curves show the saturation light intensity requirement of 1600 µmol for both the genotypes, whereas temperature response curves showed the optimum temperature requirement for Gz local as 20 °C and for GM6 it was found to be 35 °C. The results obtained for each individual parameter and other correlational studies indicate that IRGA forms a promising route for quick and reliable screening of various stress-tolerant valuable genotypes, forming the first study of its kind.