BMG Labtech has released an App note on the use of its CLARIOstar microplate reader in finding a promoter for physcomitrella patens.
The 35S promoter of the cauliflower mosaic virus is a strong and constitutive promoter that is widely used in plant systems.
However, in the moss Physcomitrella patens, its promoter strength is weak or all but silent in the dark.
Therefore, it is necessary to fi nd promoters that show higher expression levels and that have a more ubiquitous expression over time, tissues and treatments.
To test promoter strengths, the protoplasts of the moss Physcomitrella patens can be transiently transfected to allow several days of ectopic expression of circular DNA. The moss system can be used to test promoters without the need to produce mutant lines with stable integration.
The App Note demonstrated that the CLARIOstar could be used to measure the GFP fluorescence within living moss protoplasts and that the assay can be used to compare promoter strengths.
The validation of possible new promoters was carried by a promoter-reporter gene construct.
A commonly used system fuses the promoter of choice with GFP and then measures the levels of fluorescence after insertion of the construct into the plant. With the well scanning function of the CLARIOstar it was possible to check the distribution of protoplasts in the well.