Generating mammalian cells with specific mtDNA-nDNA combinations is desirable but difficult to achieve and would be enabling for studies of mitochondrial-nuclear communication and coordination in controlling cell fates and functions. We developed 'MitoPunch', a pressure-driven mitochondrial transfer device, to deliver isolated mitochondria into numerous target mammalian cells simultaneously. MitoPunch and MitoCeption, a previously described force-based mitochondrial transfer approach, both yield stable isolated mitochondrial recipient (SIMR) cells that permanently retain exogenous mtDNA, whereas coincubation of mitochondria with cells does not yield SIMR cells. Although a typical MitoPunch or MitoCeption delivery results in dozens of immortalized SIMR clones with restored oxidative phosphorylation, only MitoPunch can produce replication-limited, non-immortal human SIMR clones. The MitoPunch device is versatile, inexpensive to assemble, and easy to use for engineering mtDNA-nDNA combinations to enable fundamental studies and potential translational applications.

中文翻译：

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通过高通量、加压分离的线粒体递送稳定移植人线粒体 DNA

产生具有特定 mtDNA-nDNA 组合的哺乳动物细胞是可取的，但难以实现，并且将有助于研究线粒体-核通讯和协调以控制细胞命运和功能。我们开发了“MitoPunch”，一种压力驱动的线粒体转移装置，可同时将分离的线粒体输送到众多目标哺乳动物细胞中。MitoPunch 和 MitoCeption 是一种先前描述的基于力的线粒体转移方法，它们都产生稳定的分离线粒体受体 (SIMR) 细胞，这些细胞永久保留外源性 mtDNA，而线粒体与细胞的共孵育不会产生 SIMR 细胞。尽管典型的 MitoPunch 或 MitoCeption 递送会产生数十个具有恢复氧化磷酸化的永生化 SIMR 克隆，只有 MitoPunch 可以生产复制受限的非永生人类 SIMR 克隆。MitoPunch 设备用途广泛，组装成本低廉，并且易于用于设计 mtDNA-nDNA 组合，以实现基础研究和潜在的转化应用。