Efficient and stable gene transfer following microinjection into nuclei of synchronized animal cells progressing from G1/S boundary to early S phase
In: Biochemical and biophysical research communications, Jg. 249 (1998-09-10), Heft 3
Online
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Zugriff:
We examined the possible phase(s) of the cell cycle in which a foreign gene can be stably transferred to animal cells. DNA of the plasmid pSV2neo containing the neomycin-phosphotransferase gene was microinjected into the nuclei of NIH/3T3 cells synchronized by serum starvation and aphidicolin treatment. The frequency of neo(r)-transformation (expressed as a percentage of microinjected cells) was 6% at the G0 phase and increased with progression of the cell cycle to reach a peak of 76% at the G1/S boundary. When the cells started their growth from the G1/S following release from aphidicolin, the frequency increased or decreased in the parallel with the BrdU-labeling index. Furthermore we developed a simplified method in which asynchronously growing cells were treated with aphidicolin at 10 micrograms/ml fro 16 hrs without serum starvation and subjected to microinjection, and their growth was further induced in aphidicolin-free medium. Using five cell lines (BALB/3T3, BALB/MK-2, NRK, CHO-K1, and HeLa) and one primary culture of chicken embryo fibroblasts (CEF), a 3- to 7-fold increase in the frequency of neo(r)-transformation was consistently detected in aphidicolin-treated cells, compared to non-treated asynchronous cultures. The present study indicates that synchronized animal cells progressing from the G1/S boundary to the early S phase integrate the PSV2neo DNA into their chromosomes with high efficiency.
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Efficient and stable gene transfer following microinjection into nuclei of synchronized animal cells progressing from G1/S boundary to early S phase
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Autor/in / Beteiligte Person: | Sueyoshi, Ritsuko ; Chida, Kazuhiro ; Kuroki, Toshio |
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Zeitschrift: | Biochemical and biophysical research communications, Jg. 249 (1998-09-10), Heft 3 |
Veröffentlichung: | 1998 |
Medientyp: | unknown |
ISSN: | 0006-291X (print) |
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