重组腺相关病毒（recombinant adeno-associated virus, rAAV）载体具有安全高效的特点，已成为基因疗法的首选递送载体之一。基于哺乳动物细胞瞬时表达系统生产rAAV是目前的主流方法，由于生产效率低和给药剂量大导致基因疗法成本居高不下，因此限制了其规模化应用。随着大量基于rAAV的基因疗法进入临床试验和上市阶段，对rAAV的产能要求越来越高。通过深入认识哺乳动物细胞瞬时表达系统和rAAV生物合成特征来提高rAAV的合成效率，是建立高效、经济的基于动物细胞培养的rAAV生产工艺的关键。此文综述了rAAV的基本特性及其生产方式，并重点探讨了基于多质粒的瞬时表达过程中影响rAAV生物合成的关键因素和宿主细胞特性、以及相应的调控优化手段，为进一步开发和优化rAAV表达系统和生产工艺、实现高效生物制造提供理论参考。

Recombinant adeno-associated virus (rAAV) vectors have become one of the preferred delivery vectors for gene therapy due to their safety and effectiveness. Currently the production of rAAV is mainly based on transient expression system in mammalian cells. However, the low production efficiency and high administration dose result in the high cost of gene therapy, which limits the scale-up of its application. As a large number of rAAV-based gene therapies enter the clinical trial and marketing stage, requirement on the production capacity of rAAV is increasingly urgent. Improving the synthesis efficiency of rAAV through in-depth understanding of transient expression system and rAAV biosynthesis characteristics of mammalian cells is the key to establish an efficient and economical rAAV production process based on animal cell culture. This paper reviews the fundamental characteristics and production methods of rAAV, focusing on the key factors affecting rAAV biosynthesis and host cell characteristics during the transient expression based on multiple plasmids, as well as the corresponding regulatory optimization strategies, which provide theoretical references for further development and optimization of rAAV expression system and production process to achieve efficient biomanufacturing of rAAV.