Haynes Lab:Notebook/RNA-seq of PcTF Transfected U2OS & SK-N-SH cell lines/2015/01/19

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RNA-seq of PcTF Transfected U2OS & SK-N-SH cell lines Main project page
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Summary

Since the U2OS, SKNSH and KAH126/U2OS cultures were expanded/passaged on Friday last week, today I'll be checking on the cultures to observe their progress. The KAH126/U2OS culture in the 6-well plate should be approaching 80% confluency, at which point I'll replace the medium with fresh medium and induce with doxycycline.



Goals

  • Check U2OS, SKNSH, and KAH126/U2OS cultures for confluency
    • If KAH126/U2OS culture in 6-well plate is 80% confluent, induce expression w/ doxycycline



Protocol

To induce KAH126/U2OS cells: add 1μL doxycycline from the freezer per mL of culture. Mix by gently shaking, then place culture back in the incubator.




Notes

Observations, measurements, placeholder info etc. go here



Results/Conclusions

SKNSH culture (PS-β) - at approximately 20% confluence. Needs a few more days.

U2OS culture (PS-β) - over 80% confluence. Passage into PS-1 and split off a 6-well plate tomorrow.

KAH126/U2OS culture (6-well plate) - 100% confluence. Missed the deadline for inducing doxycycline expression. Need to make another 6-well plate from the stock, meanwhile I'll be inducing this plate to see what happens. I'm inducing 4 wells - 2 for day 5 sampling, 2 for day 10 sampling, leaving 2 for negative controls.

KAH126/U2OS culture (PS-2) - over 90% confluence. Tomorrow I'll passage to PS-3 and split some off for another 6-well plate. This time do a regular dilution - 0.2mL of parent culture per 4mL of new well.


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