A growth curvature bioassay can be used to determine the concentration of indoleacetic acid (IAA) in plant tissues. In this procedure, shoot tips are placed on agar blocks to allow IAA to diffuse into them. These agar blocks are then placed asymmetrically on one side of cut, decapitated coleoptiles (shoots with their tips removed) in the dark, and the resulting angle of curvature is measured.

Explain the biological mechanism that causes the coleoptile to curve when an agar block containing IAA is placed on only one side of its cut surface.
A student used this bioassay to compare the IAA concentration in the shoot tips of two different cultivars of wheat, Cultivar A and Cultivar B. State three variables (other than the cultivar type) that must be controlled during this investigation to ensure a valid comparison.
Using the calibration curve shown above: