An internal standard improves the reliability of transient expression studies in plant protoplasts

Transient expression of foreign genes introduced on a plasmid into isolated plant protoplasts is widely used to study the control of gene expression. Unfortunately, many experimental variables implicated in this technique are difficult or impossible to control, resulting in a disturbing degree of variability between otherwise identical experiments. We have studied the co-expression of two constitutively expressed genes located on the same plasmid. This has allowed us to identify the lot of plasmid DNA as an important source of variation, along with the protoplast lot. Plasmid DNA concentration was found to be of minor importance. Since the variation of expression level of the two genes was identical for the two genes in all experiments, we propose the use of an internal standard in all comparative transient expression studies, which allows the reduction of the variation between experiments by one order of magnitude.