Conclusion
This work reports, to our knowledge, the first single-cell
RNA-sequencing analysis of Sf9 insect cells during the production of AAV
using a low MOI, dual-baculovirus system. The findings of our work
highlights production bottlenecks of this system, suggesting the need to
reduce cell heterogeneity (e.g., by applying synchronization strategies)
and differences in transgene expression (e.g., by applying similar
promoters), as well as enhance and/or prolong the production process
(e.g., by manipulation of specific pathways) for its improvement.
Overall, this study lays the foundation for further single-cell
transcriptomics studies in insect cell-based bioprocesses and further
elevates our knowledge on the underlying biological mechanism of
multi-vector production processes.