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The utilization of pEGFP reporter system in cell-cycle analysis of adherent cells

Martina Deželjin ; Ruđer Bošković Institute, Zagreb, Croatia
Maja Herak Bosnar   ORCID icon orcid.org/0000-0002-8448-4713 ; Ruđer Bošković Institute, Zagreb, Croatia
Klara Dubravčić ; Clinical Hospital »Rebro«, Zagreb, Croatia
Ružica Bago ; Ruđer Bošković Institute, Zagreb, Croatia
Jasminka Pavelić ; Ruđer Bošković Institute, Zagreb, Croatia

Puni tekst: engleski, pdf (94 KB) str. 73-76 preuzimanja: 851* citiraj
APA 6th Edition
Deželjin, M., Herak Bosnar, M., Dubravčić, K., Bago, R. i Pavelić, J. (2008). The utilization of pEGFP reporter system in cell-cycle analysis of adherent cells. Periodicum biologorum, 110 (1), 73-76. Preuzeto s https://hrcak.srce.hr/29092
MLA 8th Edition
Deželjin, Martina, et al. "The utilization of pEGFP reporter system in cell-cycle analysis of adherent cells." Periodicum biologorum, vol. 110, br. 1, 2008, str. 73-76. https://hrcak.srce.hr/29092. Citirano 15.11.2019.
Chicago 17th Edition
Deželjin, Martina, Maja Herak Bosnar, Klara Dubravčić, Ružica Bago i Jasminka Pavelić. "The utilization of pEGFP reporter system in cell-cycle analysis of adherent cells." Periodicum biologorum 110, br. 1 (2008): 73-76. https://hrcak.srce.hr/29092
Harvard
Deželjin, M., et al. (2008). 'The utilization of pEGFP reporter system in cell-cycle analysis of adherent cells', Periodicum biologorum, 110(1), str. 73-76. Preuzeto s: https://hrcak.srce.hr/29092 (Datum pristupa: 15.11.2019.)
Vancouver
Deželjin M, Herak Bosnar M, Dubravčić K, Bago R, Pavelić J. The utilization of pEGFP reporter system in cell-cycle analysis of adherent cells. Periodicum biologorum [Internet]. 2008 [pristupljeno 15.11.2019.];110(1):73-76. Dostupno na: https://hrcak.srce.hr/29092
IEEE
M. Deželjin, M. Herak Bosnar, K. Dubravčić, R. Bago i J. Pavelić, "The utilization of pEGFP reporter system in cell-cycle analysis of adherent cells", Periodicum biologorum, vol.110, br. 1, str. 73-76, 2008. [Online]. Dostupno na: https://hrcak.srce.hr/29092. [Citirano: 15.11.2019.]

Sažetak
Background and Purpose: GFP (green fluorescent protein) is widely used in a variety of fluorescent methods aimed at revealing the fate of proteins in the cell, intracellular transport, transfection efficiency and is also recommended for cell-cycle analysis purposes. In our attempt to evaluate the role of nm23 genes in proliferation of head and neck tumor cells in culture we have decided to use EGFP reporter system and analyze the DNA content by flow cytometry.

Materials and Methods: To optimize the method we either transiently transfected the cells with pEGFPC1-nm23 constructs or cotransfected the cells with an nm23 carrying constructs and pEGFPC1 as a reporter system. We established stable clones with pEGFPC1-nm23 constructs and analyzed them by flow cytometry, as well.

Results and Conclusions:We report our experience for the use of pEGFP reporter system and flow cytometry for determining cell-cycle distribution of transiently and stably transfected adherent tumor cells. We discuss, in brief, the protocol we used and the problems that appeared during our experiments – GFP bleaching, cell clumping and degradation and insufficient number of cells to be analyzed. In conclusion, we suggest useful tips how to avoid or minimize the technical problems of this method and improve the results and analysis.

Ključne riječi
GFP; flow cytometry; cell-cycle analysis; adherent cells; transfection

Hrčak ID: 29092

URI
https://hrcak.srce.hr/29092

Posjeta: 1.081 *