Preparation of RNA Sample from Cell Culture: Difference between revisions
Created page with "==Materials== *PureLink RNA Mini Kit (Invitrogen cat#12183-018A) *Mouse Tissue (50-100 mg, about a 3mm cube) *TRIZol (Invitrogen cat# 12183-555) *Chloroform (in solvent cabine..." |
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#Incubate at room temperature for 2-3 minutes. | #Incubate at room temperature for 2-3 minutes. | ||
#Centrifuge in a cold eppendorf centrifuge (4C) on maximum for 15 minutes. Sample will separate into a lower, red chloroform phase, an interphase and a colorless upper phase which contains the RNA. | #Centrifuge in a cold eppendorf centrifuge (4C) on maximum for 15 minutes. Sample will separate into a lower, red chloroform phase, an interphase and a colorless upper phase which contains the RNA. | ||
Add 400 uL of 70% ethanol to a fresh tube. | #Add 400 uL of 70% ethanol to a fresh tube. | ||
#Transfer 400 uL of the upper phase to the ethanol tube and mix by vortexing. The remaining chloroform in the previous vial should be disposed of in the phenol waste container under the hood. | #Transfer 400 uL of the upper phase to the ethanol tube and mix by vortexing. The remaining chloroform in the previous vial should be disposed of in the phenol waste container under the hood. | ||
#Remove 700 uL of mixture (invert briefly to mix before taking sample) and add to a spin column in a collection tube. | #Remove 700 uL of mixture (invert briefly to mix before taking sample) and add to a spin column in a collection tube. | ||