Factors affecting rAAV titers during triple-plasmid transient transfection in HEK-293 cells.
Martina PistekPeter AndorferReingard GrabherrBarbara KrausJuan A Hernandez BortPublished in: Biotechnology letters (2024)
The efficiency of triple-plasmid transfection in recombinant Adeno-Associated Virus (rAAV) production was analyzed by examining two distinct HEK-293 cells lines. These were categorized as high producer (HP) and low producer (LP) based on their differing levels of productivity under identical conditions. Analysis of RNA expression levels of viral genes revealed disparities in plasmid derived gene expression between the cell lines. Further assessment of transfection efficiency utilizing labeled plasmids revealed lower plasmid uptake and less efficient nuclear transport in LP cell line. Additionally, we observed inferior translation activity in LP, contributing to its shortcomings in overall productivity. In our attempt to optimize plasmid ratios to enhance fully packaged rAAV particle yield, we discovered cell-line-specific optimization potential. The findings highlight the transfection's complexity, urging tailored strategies for improved rAAV production based on each cell line's characteristics, enhancing understanding and guiding further efficiency optimization in rAAV production.
Keyphrases
- escherichia coli
- crispr cas
- induced apoptosis
- gene expression
- cell cycle arrest
- climate change
- poor prognosis
- sars cov
- klebsiella pneumoniae
- endoplasmic reticulum stress
- dna methylation
- single cell
- oxidative stress
- computed tomography
- cell death
- genome wide
- healthcare
- binding protein
- cerebral ischemia
- blood brain barrier
- multidrug resistant
- positron emission tomography