Tracking the cyclin B1-GFP sensor to profile the pattern of mitosis versus mitotic bypass

Abstract

This chapter provides a method for quantitative single cell analysis to track the transition of single cells from G 2, indicated by high cyclin B1 levels, to G 1 polyploidy phase (G 1 (p)), indicated by low cyclin B1 levels, in a 4 n population. The cell tracking methodology described provides a fl uorescence fingerprint suitable for deriving G 2 /M or G 2 /G 1 p transitions. Notably, during late G 2 the absolute cyclin B1-eGFP reporter levels obtained were high and the switch-off point identifiable, with destruction rates of a similar order across all cell cycle routing avenues. The three principle parameters extracted were defined as (1) G 2 -to-G 1 (p) transition duration (tGFP off); (2) rate of sensor destruction (kGFP off), and (3) peak sensor expression (GFP peak).