GUS Staining and GUS Activity Assays

Glucuronidase (GUS) activities in transfected protoplasts were measured using a Synergy^TM HT microplate reader.

To examine the auxin response of the PRE6p:GUS and effects of PRE6 on the expression of the DR5:GUS reporter gene, 7-day-old seedlings of PRE6p:GUS, DR5:GUS, DR5:GUS/pre6 and DR5:GUS/35S:PRE6 were treated with 10 μM IAA for 12 h, and then used for GUS activity assays either by staining or by quantitative measurement.

For GUS staining, Arabidopsis seedlings or different tissues or organs were incubated in solution containing X-Gluc (5-bromo-4-chloro-3-indolyl-β-D-glucuronide, Rose Scientific Ltd.) as described previously (Ulmasov et al., 1997b). For quantitative measurement, Arabidopsis seedlings were frozen in liquid nitrogen, then proteins were extracted and GUS activity was measured as described previously (Strader and Bartel, 2009).