Evaluation of ethylene production in tobacco and Arabidopsis induced by particle bombardment

Introduction Particle bombardment is a helpful tool for gene expression studies in plants. It has been successfully used for transforming barley tissues (Skadsen et al. 2002) and characterizing promoter elements that regulate abscisic acid-induced gene expression in barley aleurone (Casaretto and Ho 2003). Researchers have demonstrated that real-time RT-PCR is an effective method for evaluation of mRNA transcript levels in tobacco seedlings after transformation by particle bombardment (Miyamoto et al. 2000). Using this method, they reported that the greatest transcript abundance of the GUS reporter gene in tobacco leaves transformed with a CaMV35S::GUS construct (35S promoter of the cauliflower mosaic virus [CaMV] fused to the b-glucuronidase gene) occurs 2 hr after bombardment. However, bombardment may be a problematic method when evaluating genes that are regulated by ethylene because it causes tissue damage, which potentially induces wounding responses that include increased ethylene production. Woundinduced ethylene production follows a distinct temporal pattern in most tissues, where ethylene production increases after an initial lag, followed by at least one additional peak (Saltveit and Dilley 1978). Consequently, the wound-induced ethylene response may be a confounding variable when studying genes regulated by ethylene.