1 Study of neurodevelopmental diseases necessitates primate models, and the marmoset (Callithrix jacchus) is 2 an ideal species. However, there are limited number of marmoset pluripotent stem cell lines (cjPSCs), and almost 3 no embryo-derived naïve cjPSC lines available to the marmoset research community. This application for an 4 administrative supplement is submitted to meet the objectives of RFA-DA-21-006 and is submitted under that 5 funding mechanism. We are seeking supplemental funds for U01DA056556, “Tools for gene editing in 6 marmosets,” to enable a collaboration with Dr. Ali Brivanlou’s laboratory at Rockefeller University (RU). Both the 7 RU and Johns Hopkins (JHU) groups seek to create genetically modified marmoset models of neurodegenerative 8 diseases. The objective of this collaboration is to derive new PSC cell lines from marmoset embryos through a 9 synergistic collaboration that will benefit both research teams. We will make these lines widely available to the 10 research community. Rationale for this new collaboration: Johns Hopkins (JHU) has developed expertise in 11 IVF in the marmoset with an established marmoset colony that can produce marmoset embryos and Rockefeller 12 University (RU) has expertise in creation of pre-ICM marmoset PSCs called 8-cell stage-like cells (8CLCs) that 13 have a high likelihood of germline transmission. For Aim 1, we will generate new marmoset naïve cjPSCs 14 from IVF-derived 4 to 8-cell stage embryos. Dissociated embryos created at Hopkins will be used to develop 15 new marmoset PSC lines (cjPSCs) and embryo-derived naïve cjPSC lines. These PSC lines will be made 16 available to the research community and will facilitate gene editing in the species. Aim 2 will confirm growth 17 of naïve PSCs and 8CLCs in 8-cell stage embryos and transfer of chimeric embryos into marmoset 18 females. The team at RU have developed 6 transgenic cjPSCs and this aim seeks to convert these lines into 19 8CLCs that will be injected into isochronic 8-cell stage embryos generated by the Hopkins group. We expect that 20 these injections will create chimeras. Following injection, the embryos will be transferred into females to obtain 21 F0 generation with germline transgenesis. In Aim 2A for the pilot phase of this experiment, we will inject one 22 transgenic cjPSC the Rockefeller team has generated, HTT CAG87. We will also inject 3-5 eight cell embryos 23 with other PSCs and confirm expression of the reporter construct and embryo development to the blastocyst 24 stage. For Aim 2B we will transfer 10 embryos to recipient, experienced females that have been programmed 25 and are timed to receive the embryos. A total of 25 embryos are needed for completion of all experiments 26 in Aims 1 and 2. The tools and methods developed will allow this collaborative effort to rapidly generate and 27 characterize stem cell lines in the species. We will leverage highly specialized expertise from both groups to 28 synergistically ...