Optimising transfection methods for genome editing using iPSCs and CRISPR/Cas9
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Date
2026-02-28
Authors
Beltran, Jhoan Sebastian
Journal Title
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Publisher
University College Cork
Published Version
Abstract
The emergence of induced pluripotent stem cells (iPSCs), together with major advances in genome editing technologies, has significantly expanded the potential of cell-based therapies in regenerative medicine and immunotherapy. Since their first description by Shinya Yamanaka and Kazutoshi Takahashi in 2006, iPSCs have been widely explored as a promising platform for the treatment of a broad range of diseases, including Parkinson’s disease, Alzheimer’s disease, and organ failure. Their capacity for unlimited self-renewal and differentiation into virtually any somatic cell type make them an attractive and versatile cell source for therapeutic applications.
Despite these advances, significant clinical challenges remain. One of the principal barriers to the clinical translation of iPSC-derived therapies is the potential immune rejection associated with allogeneic cell products. Consequently, multiple strategies have been proposed to mitigate immune recognition and improve the compatibility of transplanted cells.
This project aimed to contribute towards the development of hypoimmune iPSC lines by targeted knock-out of human leukocyte antigen (HLA) class I and class II molecules. To achieve this, the study evaluated alternative approaches for the delivery of genome editing cargo into iPSCs, assessing their potential as less invasive alternatives to electroporation. Transfection efficiencies obtained through cell-penetrating peptide (CPP)-mediated delivery and Lipofectamine-based methods reached approximately 40–50%%, with CPPs allowing higher viability and preservation of cellular morphology. In contrast, photoporation yielded lower transfection efficiencies, averaging around 22%. Overall, these findings suggest that CPPs may provide improved balance between cargo uptake and viability for iPSCs and will support future efforts to generate hypoimmune cell lines research and therapeutic applications.
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Keywords
iPSCs , Transfection , Optimization , Lipofection , HLA
Citation
Beltran Guiza, J. S. 2026. Optimising transfection methods for genome editing using iPSCs and CRISPR/Cas9. MRes Thesis, University College Cork.