Problem in stable expression of target protein using lentiviral vector
Posted 29 December 2010 - 08:37 AM
Posted 29 December 2010 - 11:07 AM
Posted 29 December 2010 - 03:05 PM
Thank you for your comments.Usually, I used cell lysate generated 293TN cells that were transiently transfected with lentiviral vector having target protein as control. I got a strong signaling from above control sample. However, I can not see any specific bans from stable cells having puromycin resistance. Do you think it can be helpful to change host cell type, such as HeLa cell rather than 293TN cells, to improve expression levels?
expression is much weaker in stable cells compared to transient transfection. did your WB have the positive control? you could try to load more protein. make sure the cells 293TN doesnot have the antibiotic resistance gene already. We have used 293FT and they were always in G418 containing media so we could not use this for making cell lines with G418 as selection.
Posted 30 December 2010 - 09:51 AM
by the way, how many cell lines did you screen? You might have to screen many clones before having 1 or 2 with high expression.