Plant material and growth conditions
Surface sterilised Arabidopsis thaliana seeds were sown directly onto half strength Murashige and Skoog (MS) (Duchefa Biochemie, Netherlands) 0.8 % bactoagar (BD, USA). Stratification at 4° C in the dark was for two days, before transfer to Sanyo (UK) growth cabinets (19 – 22° C, 100 µmol m-2 s-2; 12 hours light/12 hours dark.
T-DNA lines used in this investigation are described in Supplemental Table 1. Genotyping was carried out according to the instructions athttp://signal.salk.edu/tdnaprimers.2.htmlusing the primers listed in Supplemental Table 2. Left border PCR products were sequenced to determine the precise locations of the T-DNA inserts.
For srt1-4 heterozygous (hete ) and srt1-4 hete x 2-1 mutants, we performed genotyping by PCR to identify heterozygous plants for each experiment. SRT1 artificial miRNAs were designed using the MicroRNA Designer tool of the WMD3 Web site (Ossowski et al., 2008) and were introduced by transformation as described below.