Analysis of Physcomitrella Phytochrome Mutants via Phototropism and Polarotropism.

In mosses such as Physcomitrella patens phytochrome photoreceptors steer directional/vectorial responses to unilateral/polarized light. In this chapter, we describe procedures to assay phototropism and polarotropism quantitatively in wild type and mutant lines. Protonemata are placed on agar-based medium in square Petri dishes in darkness for 1 week, allowing caulonemata to develop and grow negatively gravitropically. For phototropism, the dishes are placed vertically in black boxes and unilaterally irradiated with continuous red light. For polarotropism, Petri dishes are placed horizontally and irradiated with linearly polarized red light from above. After irradiation, the filaments are photographed using a macroscope with CCD camera and the bending angles measured using image processing software. The data are transfered to a spreadsheet program, placed into 10° bending angle classes and illustrated using a circular histogram.