Allogeneic cell therapies derived from induced pluripotent stem cells (iPSCs) are an exceptionally promising class of cellular therapeutic, as they can greatly reduce the manufacturing complexities of autologous and donor-derived allogeneic cell therapies such as scalability, batch-to-batch consistency, and cost. However, host immune cell recognition and clearance of exogenous cells can lead to iatrogenic toxicities and ineffective therapeutic responses. This immune rejection is often caused by CD8+ T cells recognizing foreign HLA-I molecules, while a complete lack of HLA-I expression can result in NK-mediated immune rejection (the “missing self” response). Modulation of HLA class I elements for evasion of clearance by both CD8+ T cells and NK cells has been explored as a strategy to improve in vivo persistence. We previously reported generation of iPSCs using an mRNA-based process that avoids the genomic integration and instability risks of DNA and viral reprogramming methods. We also previously reported a novel mRNA-encoded dimerizing gene-editing endonuclease, UltraSlice™ for high-specificity gene editing and demonstrated its use in inactivating β2microglobulin (B2M), a key component of HLA class I molecules. Here, we report the development of an mRNA-engineered iPSC line developed using UltraSlice™ to express a B2M-HLA-E fusion transgene in lieu of the endogenous B2M gene product to mimic native B2M expression (i.e., upregulation when exposed to proinflammatory stimuli). Using UltraSlice™, we optimized the conditions for insertion of a flexible linker-containing HLA-E transgene upstream of the stop codon of the native B2M gene in mRNA-reprogrammed iPSCs. Specifically, a single-stranded DNA template encoding a (GGGGS)4 flexible linker and HLA-E flanked by 125-nt sequences homologous to the B2M locus was co-delivered to iPSCs along with UltraSlice™ mRNA targeting exon 3 of the B2M gene. We found that the sequence of the flexible linker was susceptible to hairpin formation, and initial experiments resulted in insertion of a truncated sequence. We then designed and tested two new linkers with GC content reduced from 83.3% to 71.7% and 61.7%, which increased the theoretical Gibb’s free energy from -6.0 to -3.92 and 0.64, respectively. We then analyzed insertion, finding no significant difference in the rate of insertion but observing successful insertion of the full repair template when using the two templates with reduced GC content. These edited populations were plated as single cells to obtain clonal mRNA-reprogrammed iPSC lines that express B2M-(GGGGS)4-HLA-E fusion protein. These cells may prove useful for the rapid development of therapeutics with the potential for increased efficacy and safety owing to the immune-evasive nature of the cells.