Preparation of RNA Samples from Mouse Tissues: Difference between revisions
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#Add 1 mL TRIzol reagent to each 2 mL tube. | #Add 1 mL TRIzol reagent to each 2 mL tube. | ||
#Cut ~50-100 mg of tissue. If tissue is in RNAlater, cut at room temperature. If tissue is frozen cut on dry ice. Weigh into a fresh 2mL eppendorf tube and keep on ice. | #Cut ~50-100 mg of tissue. If tissue is in RNAlater, cut at room temperature. If tissue is frozen cut on dry ice. Weigh into a fresh 2mL eppendorf tube and keep on ice. | ||
#Using tissue grinder, homogenize tissue for 3 | #Using tissue grinder, homogenize tissue for 3 min at 25Hz (make sure there are no remaining clumps). | ||
#Incubate 5 minutes at room temperature. | #Incubate 5 minutes at room temperature. | ||
#Add 200 uL Chloroform and shake vigourously by hand for 15s. Do not vortex. | #Add 200 uL Chloroform and shake vigourously by hand for 15s. Do not vortex. | ||
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#Spin 1 min on max to dry the cartridge. Discard the collection tube and place into a clean recovery tube. Add 100 uL RNAase free water to the center of each tube. This can be adjusted to between 30 and 300 uL elution buffer if necessary. | #Spin 1 min on max to dry the cartridge. Discard the collection tube and place into a clean recovery tube. Add 100 uL RNAase free water to the center of each tube. This can be adjusted to between 30 and 300 uL elution buffer if necessary. | ||
#Incubate at room temperature for 1min. | #Incubate at room temperature for 1min. | ||
#Spin 2 min 12500 rpm to get purified RNA. | #Spin 2 min at 12500 rpm to get purified RNA. | ||
#Quantify the RNA using the nanodrop. | #Quantify the RNA using the nanodrop. | ||