I received a pCMV5 vector of a size of 6.6 Kb and an insert of 1.9 Kb, It contains a C-terminal flag. My question is how can I assess the efficiency of mammalian cells transfection using this vector? Can I detect the β-galactosidase activity? and if Yes, what is the best method to do so.
This is the link of the backbone of the vector (without the insert)
https://www.addgene.org/vector-database/2242/
Thank you
https://www.researchgate.net/post/How_can_I_assure_the_transfection_of_Hek_cells_with_pCMX_plasmid
Please go through last reply.
Dear Razan,
You can always clone eGFP in pCMV5 vector under the same promoter and then check the GFP positive cells by flow cytometry. I think this might help you.
https://www.researchgate.net/post/What_antibiotic_concentration_should_I_use_for_transient_expression_in_CHO_K1_Or_is_it_not_necessary_to_use_antibiotics_for_selection?_tpcectx=profile_answers
Hello,
I don't know what is encoded by Yours insert, but, if the protein is not present in non-transfected cells, You can use specific antibody with any fluorescent probe, and image cells on microscope. Than count total number of cells and number of stained cells - that will be yours positive transfection. However, you will have to wait 24-48h for protein to appear.
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