Promoter choice: Who should drive the CAR in T cells?

Effect of promoter, promoter mutation and enhancer on transgene expression mediated by episomal vectors in transfected HEK293, Chang liver and primary cells

The EF-1α promoter maintains high-level transgene expression from episomal vectors in transfected CHO-K1 cells

You may check the above 3 papers. Thanks.

Hi Lukas Hyka

1. First thing to consider here is the amount of DNA transfected if you are relying on random integration. You might play around the ratio of 2.5ug plasmid/ 1.5 million cells.

2. Secondly do a 2 or 3 step screening of the single clones to find out the best expressing clone.

3. If these are already done, you can look for site-directed transfection.

Thanks,

Subham.

1. Select an appropriate cell line: The choice of cell line can have a major impact on the expression of a protein. Different cell lines can exhibit different levels of expression, so it is important to select an appropriate cell line that is known to express the protein of interest. 2. Optimize the growth conditions: Growing cells under optimal conditions is important for achieving high expression of a protein. Factors such as temperature, pH, osmolarity and growth media should be optimized to ensure maximum protein expression. 3. Use an appropriate transfection technique: Different transfection techniques such as lipofection, electroporation and viral infection can be used to introduce DNA into mammalian cells. It is important to select an appropriate transfection technique that will result in efficient and stable expression of the protein. 4. Use optimized expression vectors: Expression vectors can be optimized for high expression of a protein by using the appropriate promoters, enhancers, secretion signals, and other elements. 5. Use post-transcriptional modification techniques: Post-transcriptional modification techniques such as RNAi, antisense, and trans-splicing can be used to improve protein expression levels.