Cell synchronisation protocole using Nocodazole or Thymidine?

12 November 2024 0 7K Report

I have 2 cell lines (Hek-293 and MDA-MB-231) and i have a DNA construct that contains a selection marker (Puro) which I wanted to check cell lysates in G2/M,

1/ should I prepare the viral partical in HEK cells (DNA + packaging plasmids) after that infected in MDA-MB-231 and selected after that I add nocodazole? or no need and I use just one cell line and how?

2/ after I add nocodazole (16 h) i just need to was the plate and add RIPA buffer and collect cell lysate as always or there is specific step?

3/ Is it needed to make double blocking (nocodazole +thymidine) or use nocodazole alone is enough?

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