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Advanced Gene Editing Techniques in T-Cell Modeling with I-SceI and H2-Kd Targeting

This study explores the application of I-SceI genome editing for modifying T-cells expressing CD4 and CD8 markers. We investigate the efficiency of introducing specific mutations using NHEJ pathways and assess the co-expression of genes like eGFP and Puro under CMV promoters. Data reveals nuanced differences in targeting and expression levels, particularly focusing on H2-Kd and GFP metrics. The findings underline the potential for enhanced T-cell responses in immunotherapy applications.

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Advanced Gene Editing Techniques in T-Cell Modeling with I-SceI and H2-Kd Targeting

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  1. I-SceI I-SceI CD8 CD8 H2-K CD4 CD4 CD4 Bsd TGC and NHEJ I-SceI CD8 CD4 CMV H2-K eGFP Puro GFP-, H2-K+, CD8-, CD4- I-SceI trGFP eGFP Puro H2-K- CD4+ CMV GFP+ H2-K- CD8+ CMV H2-K- CD4/8- CM 2-K

  2. I-SceI I-SceI CD8 CD8 H2-K CD4 CD4 CD4 Bsd TGC and NHEJ I-SceI CD8 CD4 CMV H2-K eGFP Puro GFP-, H2-K+, CD8-, CD4- I-SceI trGFP eGFP Puro H2-K- CD4+ CMV GFP+ H2-K- CD8+ CMV H2-K- CD4/8- CM 2-K

  3. Targeting @ A  Mutation @ B no I-SceI H2-Kd stained + I-SceI stained H2-Kd + I-SceI unstained 0.42% 0.37% 0.004% GFP 0.29% 0.003% 0.23% 0.11% 14.5% 84.4% 2.4% 97.3% H2-Kd (PE)

  4. Targeting @ A  Mutation @ B no I-SceI H2-Kd stained + I-SceI stained H2-Kd + I-SceI unstained 1.96% 1.62% 0.001% GFP 1.34% 0% 0.77% 0.67% 10.9% 86.6% 9.7% 89.4% H2-Kd (PE)

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