Experimental design PP

In an effort to extend the expression data in Phytozome, we designed an experiment to study Bradi3g27407 gene expression under 5% glucose growth conditions. To explore this, we conducted an experiment to study expression levels in root samples in the presence and absence of 5% glucose. We chose root samples because that is where our gene is most expressed, as indicated by our results from the e-FP browser. We used 8 root samples (5cm, young) from Brachypodium distachyon plants, growing 4 experimental samples in MS medium plates containing 5% glucose and another 4 control samples in MS medium plates without 5% glucose. We grew the plants at optimum temperature and light conditions: 24℃ day, 18 ℃ night. We created primers for the reverse transcription reaction, using primer3 software, so that they flanked introns but bound to exon sequences. The forward primer was 5’-tacaaggggaagatcagggc-3’ and the reverse primer was 5’-ccgcttgatctccttctcca-3’. These were so that the length of the sequence between the primers (not including the intron sequence) was 321bp (Figure S3). In Figure S3, the texts highlighted in yellow are the exon sequences, that highlighted in green is the intron sequence and the texts with red font color are the primers.