Development of a Robotic Platform for Automated Screening and Transformation of Switchgrass Suspension Cultures

Switchgrass (Panicum virgatum) is an important lignocellulosic feedstock with tremendous potential for biofuel production, as well as, a good source of value added products for multiple industrial applications. Associated with the processing of switchgrass biomass is the need to regenerate plants with lower recalcitrance. To achieve this goal, in this study, a robotic platform was developed for high throughput transformation and screening of switchgrass suspension cultures. The robotic platform includes 2 liquid handlers for precise pipetting and washing of 96-well plates, a plate incubator and shaker, a monochromator-based microplate reader, and a robotic arm, all housed within a HEPA-filtered biosafety cabinet. Prior to conducting automated transformation experiments, a FACS system was used to sort viable cells, as measured by fluorescein diacetate (FDA), 1 cell per well, into a 96-well plate. After sorting, the platform was used to transform the cells with Agrobacterium using a protocol developed in this work. After transformation, cells were analyzed for transformation efficiency by monitoring the expression of a GFP reporter at 1 hour intervals over the course of 24 hours. At 24 hours, cells expressing GFP were “cherry-picked” by the liquid handler to generate a plate with 100% transformed cells. Through this work we have demonstrated a proof-of-concept validation for the use of this platform for high-throughput transformation and screening. Future studies will focus on mass screening of various lignin constructs to demonstrate the potential of the system for rapid identification of target transgenic switchgrass.