1. Please state project objectives and what work was done this quarter to address them: The objectives of this project were to produce disease resistant, commercially & agronomically acceptable, mature citrus transgenics, intragenics, & GMO/non-GMO edited trees using Agrobacterium as a service for research & commercialization. The research focus of this project was to improve transformation efficiencies, so that the mature citrus protocols become more productive, decrease prices for scientists, & contribute more to financial self-sufficiency of our lab. A biolistic transformation protocol was also developed to overexpress all-citrus intragenic sequences using a citrus gene for selection, which is useful for food crops. A final comprehensive report will be submitted in the near future to CRDF. The efficiency of the mature citrus Agrobacterium transformation protocol was increased significantly, primarily due to the introduction of new cultivars developed by the Plant Improvement Team & determining which ones were not recalcitrant to Agrobacterium. The mature citrus Agrobacterium protocol was originally developed for Pineapple sweet orange but Pineapple is not commercially important to the Florida juice industry. Instead we have found several new sweet orange cultivars that are readily transformed with Agrobacterium. Currently we are focusing on producing non-GMO edited mature citrus & have seen some encouraging results. In addition, we continue to produce all-citrus intragenics using biolistic & Agrobacterium transformation. The plants produced using these technologies might not be as strictly regulated compared to plants produced by transformation with genes from foreign organisms. The USDA APHIS, EPA & FDA will work with each scientist on a case by case basis to deregulate superior intragenic trees. Intragenic trees can likely be provided to the growers relatively rapidly. During the last quarter, ~82 transgenics were produced using Agrobacterium transformation of mature rootstocks & ~87 Cas9 trangenic edited scions were produced for another scientist. We have two new potential projects involving scientists from other states & countries. A manuscript describing a protocol for a new liquid selection system in mature rootstock was published, which showed significantly higher Agrobacterium transformation efficiency in liquid than on solid medium. In addition, Agrobacterium transformation efficiency also increased in mature scions using liquid selection & another manuscript is in preparation. Currently, a manuscript is in review about the new citrus, intragenic selectable marker. It is the responsibility of the scientists to field test their trees & to my knowledge, at least ~ two to three projects are being field tested in the Picos Farm during this last granting cycle. The lab transitioned to an Educational Business Account for earning salaries & operating funds, however it is difficult to earn enough money for salaries by selling plants. Prices are being increased again in 2022 since we will no longer receive CRDF funding.