Protocol

Phenotypic Analysis of Arabidopsis Mutants: Brassinosteroid Hormone Response

This protocol was adapted from "How to Analyze a Mutant Phenotypically," Chapter 4, in Arabidopsis: A Laboratory Manual (eds. Weigel and Glazebrook). Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY, USA, 2002.

INTRODUCTION

Brassinosteroid (BR) is required for normal plant growth and development; most notably, BR promotes cell elongation. Thus, mutants with defects in BR biosynthesis or response have defects in cell elongation. They fail to undergo skotomorphogenesis (etiolation) in the dark and they are dwarves when grown in the light. Whether a cell elongation defect can be rescued, unaffected, or alleviated by BR treatment provides an important criterion for whether a mutant has a defect in BR biosynthesis or BR response or whether the mutant phenotype is unrelated to BR. On the other hand, mutants with increased BR signaling are less sensitive to the BR-biosynthesis inhibitor brassinazole (BRZ). The hypocotyl length of dark-grown seedlings and petiole length of light-grown plants are often measured to determine BR responsiveness; BRZ inhibition of BR biosynthesis can reduce hypocotyl and petiole elongation. Wild-type seedlings grown on 1 µM BRZ in the dark have hypocotyls approximately one-third the length of those on medium lacking BRZ. This protocol describes the measurement of BR responsiveness, with or without BRZ, in light- and dark-grown seedlings.