I’m in the process of developing stable Huh-7 cell line for luciferase purposes. My experiment is mostly based on Connelly’s (2012) work, except I’m using pmirGLO vector instead of psiCHECK-2. Is there some major difference which promotes using the latter one for this purpose?
Anyway, I cotransfected my construct with pcDNA vector and exposed cells to the G418 selection agent. Everything went as expected - cells which survived formed ‘islands’ - so than I moved 1 whole ‘12 well’ to 1 smaller ‘96 well’ plate and now they are growing in 1 ‘48 well’. It looks ok, cells are alive and growing, but the process of growing is very slow (compared to non-treated Huh-7), it takes more than a week to make such small well confluent enough. Is it normal? Current photo in the attachment. Maybe you could point some dangerous symptoms from this one?
Thank you for your time.
HUH7 is a relatively slow growing cell line, and having been transfected may affect its growth further. I wouldn't worry too much about it!
Which media are you using to culture the cells?
Thank you for your answers. I'm using RPMI (line was purchased from CLS [http://www.clsgmbh.de] as they recommend).
Ohh OK, we generally culture these cells in DMEM. It is a liver tumour cell line and DMEM being comparatively rich in glucose, helps with the cell culture. But again if your protocol suggests the use of RPMI, it is upto you! Hope this helps!
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