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Characterisation and Gene Expression Profiling of a Stable Cell Line Expressing a Cell Cycle GFP Sensor
Nick Thomas, Mike Kenrick, Theresa Giesler, Gretchen Kiser, Hayley Tinkler and Simon Stubbs
volume 4 | issue 1
january 2005Pages: 191 - 195
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The use of stable cell lines expressing fusions with green fluorescent protein (GFP) has increased significantly in recent years. In this study we have used a range of complimentary analytical techniques to examine the characteristics of a cell line stably expressing a EGFP cell cycle sensor relative to parental U2OS cells. Analysis of cell cycle duration and cell cycle phase distribution by cell growth assays and flow cytometry revealed that the two cell lines had identical doubling times and cell cycle distributions. Measurement of EGFP fusion protein mRNA by quantitative RT-PCR indicated a EGFP sensor expression level equivalent to endogenous Cyclin B1 (7000 copies/cell in G2). Microarray analysis showed a 0.9% (>2 fold at p<0.001 across 20,000 genes) difference in global gene expression levels between parental and EGFP expressing U2OS cells, with no significant differences in expression of A, B, C, D, E, F, G, H, I, K, L, M or T type Cyclins between the two cell types. These results confirm that engineering a stable cell line for low expression of a EGFP cell cycle sensor is minimally perturbing to the cell cycle and cellular gene expression.
We now provide open access to journal articles published online for one year or more. This article may be downloaded at the following link:
Download PDF If the document does not open, please right-click on the link (control-click on a Macintosh) and select the option to save the file to disk.