When using a lentiviral vector for inducible expression of genes using the Tet-On system.
In the literature, I saw a widely used of pLVX-TetOne-Puro Plasmid, is there a reason why people prefer to work with puro and not using copGFP as the selection marker?
A good question, actually, there are pLVX-TetOne-EGFP-Puro vectors.
For GFP, you need a cell sorter to get many pure GFP+ cells.
For copGFP, I am wondering the harm of copGFP aggregates as copGFP tends to form aggregates in mammalian cells especially in long-term cultures.
Regards
Both flow sorting and antibiotic selection have their uses in purification of transduced cells, but achieve this from two different directions.
Fluorescence-activated cell sorting selects cells based on a user-defined/adjusted threshold. This positive selection is particularly useful in establishing a clonal cell lines, which reduces the variability of transgene expression.
Antibiotic selection removes cells that do not possess the resistance gene included in the viral cassette. This negative selection is commonly employed over positive selection by FACS due to the simplicity and low labor requirement. Generally, unless clonal selection is required antibiotic selection is sufficient to produce a transgenic cell line. In high-throughput designs, negative selection is far more useful than positive selection due to the scalability compared to FACS.
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