Hosuk:LabNotes/2013-6-11: Difference between revisions

From ZhangLabWiki
Jump to navigation Jump to search
>Hosuki78
No edit summary
>Hosuki78
No edit summary
Line 9: Line 9:
=====Rolony - Day 2=====
=====Rolony - Day 2=====
*Cell preparation
*Cell preparation
**100% confluence, and coudl wait one more day, and split
**100% confluence, and could wait one more day, and split
 
 
**When cells are transferred from 6 well plate
**When cells are transferred from 6 well plate
**500uL of Trypsin, incubate 5min
**500uL of Trypsin, incubate 5min

Revision as of 15:50, 11 June 2013

Day 2

Image processing
  • Imaris
  • Huygens


Rolony - Day 2
  • Cell preparation
    • 100% confluence, and could wait one more day, and split
    • When cells are transferred from 6 well plate
    • 500uL of Trypsin, incubate 5min
    • Add 500uL media, and mix well, and
    • 100uL media in 96 well, or 300~350uL media in glass bottom dish
    • Add 60uL of cell in 96 well or 200uL in glass bottom dish
    • Culture one or two more day


  • BS(PEG)9
    • 20uL in 1mL PBS
    • Add 200uL of BS(PEG)9 mix per one glass bottome dish
    • Add 50uL per one well of 96 well plate


  • CircLigase II
    • Use 2uL of enzyme, but make 200uL mix by doubling all reagents
    • 128uL H2O
    • 20uL 10x Buffer
    • 10uL MnCl2
    • 40uL Betaine
    • 2uL CircLigase II


  • RCA
    • 1~2hr. for RT or 2hr. for CircLigase II would be enough, but
    • incubate over 12 hour