Quick and efficient measurement of transient GUS expression using image analysis

An image-analysis procedure was developed for measurement of transient β-glucuronidase (GUS) gene expression events in peanut. Gene transfer via microprojectile bombardment has been effective in peanut. Experiments were performed to improve the efficiency of DNA delivery to embryogenic calli for transformation. Transient GUS expression was assayed 24 hours after bombardment by staining calli and manually counting blue-stained spots under a microscope. These calli were then placed under a dissecting microscope with a video camera that recorded each image onto a computer. The images were then subjected to analysis with software that counted the number of blue-stained GUS-positive spots as well as the area and percent of the total area of each spot. The manual counts and data from the image analysis were compared through correlation statistics and by comparing analyses of variance for fixed parameters of the experiments. Correlation coefficients were high between manual spot counts and image-analysis counts (r=0.77) and between manual counts and percent area (GUS (r=0.91) Results indicate that image analysis provides a quick and efficient method for analysis of transient expression. Much more reliable information is obtained with less effort and less fatigue with the image-analysis procedure. Scientists can use this method to monitor transient expression events in plant transformation experiments.