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EM12

EXTRACTME miRNA KIT, 10 preps

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For purification of miRNA from tissue or cultured cells
Available for risk-free testing.
€ 30,00

The EXTRACTME miRNA KIT is designed for the rapid and efficient purification of miRNA with possibility of simultaneous purification of large RNA and DNA. High quality miRNA may be purified from 1-10 mg of tissue (fresh or frozen) and 104-106 cultured cells. The isolation protocol, buffers formulations and columns were optimized for high isolation efficiency and purity of miRNA.

PRINCIPLE

The EXTRACTME miRNA KIT utilizes spin minicolumns with membranes which efficiently and selectively bind nucleic acids at high concentration of chaotropic salts. The isolation procedure consists of 7 steps. In the first isolation step, the tissue is homogenized in order to disintegrate intercellular bonds (epithelial tissue) and fragmentize high-molecular proteins (muscle or connective tissue). Then the homogenate is lysed with guanidine thiocyanate and detergents. Any RNases are inactivated by guanidine thiocyanate and ß-mercaptoethanol. In next step DNA is separated by binding to the first minicolumn. Next the large RNA is bound to the Large RNA Purifcation Column membrane by selective conditions in mixture after addition of ethanol. The miRNA is bound in next step to the miRNA Purifcation Column. The three-step washing stage effectively removes impurities and enzyme inhibitors. The purified miRNA is eluted using a low ionic strength buffer or RNase-free water (pH 7.0-9.0) and can be used directly in all downstream applications such as RT-PCR, Northern blotting, RT-qPCR and so forth.

SAMPLE MATERIAL: 

  • fresh or frozen tissue (stored at -80°C): 1 – 10 mg
  • tissue preserved in RNase inactivating buffers: 1 – 10 mg
  • cell culture: 104 – 106 cells
BINDING CAPACITY: approx. 90 μg

TIME REQUIRED:

  • 25 – 30 minutes (lysis and homogenization time not included)
  • 35 – 70 minutes for homogenization in liquid nitrogen
  • 35 – 50 minutes for mechanical homogenization (ceramic beads)
RNA PURITY: A260/A280 ratio = 1.9 – 2.1
More info on manufacturer's site
PROTOCOL

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