Thermo Fisher Scientific
mirVana miRNA isolation kit
AM1560
The aim of this experiment was to extract total RNA from skin biopsies (approx. 4mm) for downstream microRNA quantification (qRT-PCR).
microRNA and gene expression assays
Skin biopsies
Total RNA was extracted from clinical skin biopsies using the mirVana miRNA Isolation kit. Skin biopsy was homogenised on a sterile petri-dish, in 60 µl of Lysis Buffer using a sterile scalpel. The lysate was transferred into a microcentrifuge and made up to 300 µl with the Lysis Buffer. Then the Homogenate Additive was added and the sample was incubated on ice for 10 minutes. Phenol:chloroform was then carefully added to the sample, vortexed and then centrifuged. The upper phase was carefully transferred to a fresh tube and 100% Ethanol added to it. The mix was then loaded onto a Filter Cartridge and centrifuged. The sample was washed several times using the indicated Wash Buffers 1 & 2. The Filter Cartridge was dried by centrifugation and the Filter was transferred to a new Eppendorf and pre-heated nuclease-free water (100 µl) at 95ºC was used to elute the total RNA.
Initially, homogenise the skin using sterile scalpel.
Good quality RNA yield.
None
Easy to use.
Phenol-based.
Good yield, great if you only need to extract total RNA and microRNA.