EB04119 (10,917 bp) is a modified version of pEGZ2 REF1,2 domesticated for Golden Gate cloning and containing a DNA barcode. The parental vector was domesticated by removing an internal BsmBI site in the hygromycin cassette via a synonymous mutation (GTC→GTT), and an mScarlet dropout cassette with outward-facing BsmBI sites was inserted downstream of WPRE. EMMA fusion sites at position 17 (GCTC-GCCA) were used. Five fragments were PCR-amplified and assembled by Golden Gate using PaqCI, with overhang fidelity verified via NEBridge. The final construct was sequence-verified. 1.Wojtowicz, E. et al. Panhematopoietic RNA barcoding enables kinetic measurements of nucleate and anucleate lineages and the activation of myeloid clones following acute platelet depletion. Genome Biology 24 (06/27/2023). https://doi.org/10.1186/s13059-023-02976-z 2.Belderbos, M. E. et al. Clonal selection and asymmetric distribution of human leukemia in murine xenografts revealed by cellular barcoding. Blood 129 (06/15/2017). https://doi.org/10.1182/blood-2016-12-758250 3.Martella A, Matjusaitis M, Auxillos J, Pollard SM, Cai Y. EMMA: An Extensible Mammalian Modular Assembly Toolkit for the Rapid Design and Production of Diverse Expression Vectors. ACS Synth Biol. 2017 Jul 21;6(7):1380-1392. doi: 10.1021/acssynbio.7b00016.