Daniel:Notebook/HiResChrPaint/2014-11-4: Difference between revisions

From ZhangLabWiki
Jump to navigation Jump to search
>Djacobse
No edit summary
>Djacobse
 
Line 35: Line 35:
#Allow the rubber cement to air-dry for 5 minutes at room temperature
#Allow the rubber cement to air-dry for 5 minutes at room temperature
#Denature for 2.5 minutes at 92C; keep humid
#Denature for 2.5 minutes at 92C; keep humid
#Transfer slides to a humidified chamber and hybridize overnight at 37C or 42C
#Transfer slides to a humidified chamber and hybridize overnight at 37C






[[Category:HRCP]] [[Category:Hybridization]]  [[Category:20141104]]
[[Category:HRCP]] [[Category:Hybridization]]  [[Category:20141104]]

Latest revision as of 17:34, 5 November 2014

FISH[edit]

Back to Calendar

Hybridization[edit]

Sample Matrix

Sample uL DMD-A-488 (pmol) uL Cot1-A-546 (ug)
A 3 (25.5) 8 (0.4)
B 10 (85.5) 8 (0.4)
  1. Allow stored slides to warm to room temperature
  2. Incubate slides in 2X SSCT +50% (v/v) formamide for 2.5 minutes at 92 C in a pre-warmed coplin jar
  3. Incubate in coplin jar containing 2X SSCT + 50% formamide at 60C for 20 minutes
  4. Remove slides and allow to cool to room temperature
  5. Add 25 uL hybridization cocktail to a 22x22 #1.5 coverslip
  6. Invert slides on to cocktail-covered coverslips; seal with rubber cement
  7. Allow the rubber cement to air-dry for 5 minutes at room temperature
  8. Denature for 2.5 minutes at 92C; keep humid
  9. Transfer slides to a humidified chamber and hybridize overnight at 37C