Illumina HiSeq 2000 paired end sequencing; RNA-seq of Medicago truncatula A17 and the ethylene insensitive sickle mutant interacting with the fungal root pathogen, Rhizoctonia solani AG8.
RNA-seq of Medicago truncatula A17 and the ethylene insensitive sickle mutant interacting with the fungal root pathogen, Rhizoctonia solani AG8.
M. truncatula lines A17 and skl were treated, grown and inoculated according to (Anderson et al., 2010). R. solani AG8 (isolate WAC10335) was prepared according to (Lichtenzveig et al., 2006). Briefly, surface sterilised M. truncatula seeds were germinated on moist filter paper in the dark at 4 °C for two days, then one day at 24 °C. Seedlings were added to pots half filled with vermiculite also containing 5 ml of 2.2mg/ml dry weight equivalent suspension of homogenised fungal mycelium. The seedlings were covered with fine vermiculite and transferred to growth rooms at a constant temperature of 24 °C with 16/8 light/dark photoperiod at a light intensity of 200 mmol m-2 s-1. Mock treated plants were treated the same with the addition of 5 ml sterile water rather than the 5 ml of R. solani suspension. Inoculated plants were harvested at two and seven days after planting into pre-inoculated pots with corresponding mock samples taken at the same time. Root and above ground tissue was collected separately, immediately frozen in liquid nitrogen and stored at -80 °C until RNA extraction. The remaining plants were scored for survival and harvested at 21 days after inoculation to confirm the infection had progressed as previously described (Anderson et al., 2013).  Frozen root tissue was ground in the presence of liquid nitrogen and total RNA extracted using Trizol reagent (Sigma) according to manufacturer’s instructions.  Libraries were prepared using the illumina TruSeq Stranded mRNA Library Prep Kit according to manufacturer’s instructions.
CSIRO Agriculture and Food
E-MTAB-5473:A17 R. solani inoculated  2 dpi Rep3_p
Experimental Factor: A17
ecotype
Experimental Factor: Rhizoctonia solani AG8 inoculated
disease
Experimental Factor: 2
sampling time point