Cell cycle progression and arrest - (May/18/2006 )

HI,
I want to design a exp. about the effects of viral proten in DNA replication progression (2N to 4N) by transient transfection system. However, I don't known which assay condition is better. Please give some suggestion...

Two possible strategies could be used in my assay:

One is to synchronize cells in G0/G1 phase by serum starvation and then release synchronized cells,
(However, I don't know whether serum starvation repress transfected gene expression.)

Second is to synchronize cells in S phase by double-thymidin and then release synchronized cells

Please give some suggestions about my exp. design.. very thanks

Best regards

Chung

You could try enrichment via flow cytometer. I don't have the protocol for isolation of cells at G0/G1 but the following paper could help you search for one.

Widrow & Laird 2000. Enrichment for submitoti cell populations using flow cytometry. Cytometry 39: 126-130