Department of Biology

The Wisconsin Crop Innovation Center - In the public sector, moving plant transformation forward

Dr. Ray Collier
Wisconsin Crop Innovation Center, University of Wisconsin-Madison

July 25, 2023
2:00 pm Sullivan 101

Hosted By Dr. Ayalew Osena

Abstract

In January of 2017 the Monsanto company (now Bayer Crop Sciences) gifted to the University of Wisconsin – Madison the Agracetus campus, located in Middleton, Wisconsin, in which a majority of Monsanto dicot transformation projects were conducted between 1996 through 2016. This ~100000 ft2 facility now operates as the largest public sector fee-for-service plant transformation facility in North America. Direct meristem transformation, which helps to mitigate germplasm related transformation recalcitrance, is the core technology upon which the WCIC dicot transformation pipelines have been established, allowing for, relative to more traditional approaches, expedited inception to bag-of-seed project timelines. Clients work first with the Molecular Technologies Department (MTD) on project design. The MTD uses gene synthesis and Golden Gate cloning to generate binary vectors (or naked plasmid for bombardment projects) to produce Agrobacterium stocks for hand off to the Production Transformation Team (PTT). Transgenic plants generated by the PTT are delivered to the Plant Analysis Team for endpoint PCR confirmation of selection marker presence and/or transgene copy number enumeration via droplet digital PCR (ddPCR). Verified transgenic events are raised to maturity by the Greenhouse Team, who works with the client on completion of USDA-APHIS permits allowing for shipment of the transgenic seeds. The WCIC offers fee for service transformation of soybean, maize, wheat, sorghum, Brachypodium, barley, Cannabis (hemp, less than 0.3% THC content), chickpea, cowpea, and banana, and are actively developing protocols for common bean, chickpea and field pea. While the MTD does provide fee-for-service assembly of bespoke plant transformation plasmids, the new user friendly DIRECTCLONE plasmids, featuring the WCIC synthetic binary plasmid backbone loaded with a WCIC preferred plant selectable marker (with optional seed expressed tdTomato or RUBYv1 scorable markers) have been designed and assembled by the MTD and are scheduled to be deposited in Addgene in Summer 2023. DIRECTCLONE plasmids provide end users with a onestep path, using either T/A cloning (gene of interest overexpression or promoter GUSPlus options) or Golden Gate cloning (CRISPR/Cas9 gene editing), to generate finished binary plasmids for shipping to WCIC for use in their contracted fee-for-service plant transformation projects.