This study aimed to propose an effective positions of CO
2 supply in tomato plants grown by high-wire system to reduce CO
2 leakage from a naturally ventilated greenhouse. CO
2 was supplied at two different positions within the tomato canopy, and CO
2 concentration was measured at different heights above the ground within or outside the canopy under ventilated or unventilated conditions in the greenhouse. CO
2 supply started below 400 µmol mol
-1 of the CO
2 concentration measured at 2.4 m above the ground and stopped at 450 µmol mol
-1. When CO
2 was supplied at the base of the canopy (0.6 m above the ground), the CO
2 concentration near the top of the canopy (2.4 m above the ground) was below 400 µmol mol
-1 in some cases. On the other hand, when CO
2 was supplied at the middle layer of the canopy (1.2 m above the ground), the CO
2 concentration within the canopy (1.2 to 2.4 m above the ground) was maintained around 450 µmol mol
-1 regardless of ventilated or unventilated conditions. The CO
2 concentration below the roof windows (4.2 m above the ground) was hardly increased by changing the position of CO
2 supply from 0.6 to 1.2 m above the ground. The CO
2 concentration at 4.2 m above the ground under unventilated condition was slightly above 400 µmol mol
-1, which was almost the same as that under ventilated condition. In summary, it was suggested that the CO
2 supply at the middle layer of the canopy was more appropriate than at the base of the canopy to elevate the CO
2 concentration within a canopy of tomato grown by high-wire system. In addition, the CO
2 supply at the middle layer of the canopy was seemed to have a low CO
2 leakage from the top windows even in the ventilated greenhouse.
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