In my capacity as the Research Manager of the Molecular Technologies Department within the Wisconsin Crop Innovation Center, part of the University of Wisconsin – Madison, I am engaged in several projects in which I utilize Golden Gate cloning. Since first adopting Golden Gate cloning during my post-doctoral work in the Ané lab at the UW, I have worked to develop new TypeIIS mediated cloning tools, to expand the applicability of the MoClo Plant Parts kit, for constructing plant molecular biology focused transcriptional units and binary plasmids. I have noticed during the past year that every time I search for information regarding TypeIIS enzymes, or Golden Gate cloning that that I find exactly the paper I am looking for in the literature, with publication dates are often only months or weeks before I have searched for them. I am very happy to be able to tap into such a vibrant and active area of research. The latest examples are a couple of outstanding papers recently published by the fine folks at New England Biolabs (pub 1) (pub 2). These manuscripts provide a treasure trove of empirical data, related to ligation efficiency and fidelity of 4 nucleotide overhanging sequences, that anyone engaged in non-traditional Golden Gate cloning design would do well to read and use in their future work.
After a long waiting period, the work completed during my post-doc with the USDA in California was finally published in 2018. Our unconventional approach to Agrobacterium-mediated plant transformation was met with skepticism and our manuscript was rejected by several journals. Finally, the sacrifice of nearly all my weekends during 2015 and 2016 was rewarded when our manuscript was accepted. The cherry on top was the selection of our work, including a cover image, as Featured Article in The Plant Journal (volume 95, issue 4, August 2018; https://doi.org/10.1111/tpj.13992).
The Molecular Technologies Department that I manage at the Wisconsin Crop Innovation Center, of the University of Wisconsin – Madison, has been fortunate to receive internal funding from the University of Wisconsin – Madison for instrumentation to help us to achieve our goals in helping UW researchers conduct their research. One of the devices that I have installed in the laboratory is an oKtoputre DNA extraction robot, manufactured by LGC (formerly Douglas Scientific). I made and posted a short video of the robot in operation. I recently utilized this well made robot (which I have named “Stephen”, in honor of Stephen Curry, since the robot does a little “shimmy” every time if ejects or picks up tips) to extract DNA from four 96 well plates containing soybean leaf samples from plants that are part of a CRISPR/Cas9 gene editing experiment. The entire procedure took ~ 2 hours, with another hour for cleaning. With the robot and DNA extraction protocol well established, we now offer genomic DNA extraction as one of our fee for service offerings. I hope that this unit will help many researchers to push forward with their projects.