RNA polymerase III (Pol III) promoters express short non-coding RNAs and have been adopted for expression of microRNA, interference RNA, and CRISPR single guide RNA (sgRNA). Vectors incorporating H1 and U6 Pol III promoters are being applied for therapeutic genome editing, including multiplexed CRISPR/Cas9 effects. We report a nucleosome-depleted, minimal U6 promoter, which when embedded within lentiviral long terminal repeat (LTR) regions, supports high level transcriptional activity. Furthermore, duplex minimal H1 & U6 promoters transcribed dual sgRNAs for simultaneous disruption of T cell receptor (TCR) and human leukocyte antigen (HLA) molecules, supporting efficient generation of ‘universal’ CAR T cells.

RNA 聚合酶 III (Pol III) 启动子表达短非编码 RNA，已用于表达 microRNA、干扰 RNA 和 CRISPR 单引导 RNA (sgRNA)。包含 H1 和 U6 Pol III 启动子的载体正用于治疗性基因组编辑，包括多重 CRISPR/Cas9 效应。我们报告了一个核小体耗尽的最小 U6 启动子，当它嵌入慢病毒长末端重复 (LTR) 区域时，支持高水平的转录活性。此外，双链最小 H1 和 U6 启动子转录双 sgRNA，同时破坏 T 细胞受体 (TCR) 和人类白细胞抗原 (HLA) 分子，支持“通用”CAR T 细胞的高效生成。