20
edits
Changes
no edit summary
#Add the DNase I stock solution to 70 uL buffer RDD. Mix by gently inverting the tubem, and centrifuge briefly to collect residual liquid fro tmt the sides of the tube.
#Add the DNase I incubation mix (80uL) directly to the RNeasy spin column embrane, and place on the benchtop (20-30<sup>o</sup>C) for 15 min.
#Add 350 uL buffer RW1 to the RNeasy spin column. Close the lid gently, and centrifuge for 15 s at >8000 x g or >10,000 rpm. Discard the clow-through.
'''Elution'''
#Add 500 uL buffer RPE to the RNeasy spin column. Close the lid gently, and centrifuge for 2 min at >8000 x g or >10,000 rpm. to wash the spin column membrane. This dries the spin colujmn membrane, ensuring that no ethanol is carried over during RNA elution. Residual ethanol may interfere with downstream reactions.
# Place the RNeasy spin oclumn in a new 2 ml collection tube, and discared the flowthrough. Close the lid gently, and centrifuge at full speed for 1 min.
# Place the RNeasy spin column in a new 1.5 ml collection rube. Add 30 uL RNase-free water directly to the center of the spin column membrane. Close the lid gently, and centrifuge for 1 min at >8000 x g or >10,000 rpm to elute the RNA.
# If the expected RNA yield is >30 ug, repeat the previous step using another 30-50 uL RNase-free water, or using the eluate from the previous step. Reuse the collection tube from th eprevious step.
