Haynes Lab:Notebook/RNA-seq of PcTF Transfected U2OS & SK-N-SH cell lines/2014/12/17

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Owwnotebook icon.pngRNA-seq of PcTF Transfected U2OS & SK-N-SH cell lines Report.pngMain project page
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Summary

Today we're checking the transfected U2OS cells in the 6 well plates for transfection efficiency. This is done by putting the sample under the fluorescence microscope and observing mCherry fluorescence. mCherry is a fluorescent marker tagged to the Pc-TF protein expressed in the plasmid. The more cells with fluorescent red nuclei (Pc-TF has a nuclear colocalization tag), the greater the transfection efficiency.

Goals

  • Determine transfection efficiency of U2OS cells transfected the previous day

Protocol

None

Materials

None

Notes

Composite image of phase contrast + mCherry fluorescence, adherent U2OS cells:

2014-12-17 U2OS cells transfected with mCherry-PcTF.jpg

Results/Conclusions

Transfection efficiency appears to be low, <10%. Could be due to low-quality plasmid DNA, or operator error. Going to repeat the transfection procedure, starting by splitting U2OS cells into 6-well plates tomorrow.