Direct mRNAdembeads Purification Kit

Supplier
Ademtech
Cat. No
2006021
Kit size
30 preps
Links
ademtech Protocol Information Sheet

Features

• Allow to work with small volume in comparison with mRNA purification from Total RNA
• Ready to use kit: no beads regeneration required
• More convenient for many subsequent RT-PCR
• Scalable system: the quantities of reagents are adapted for each amount of starting material (cells or whole blood)
• Very low ribosomal contamination
• Intact and pure mRNA without degradation
• Purification of all mRNA sizesFast system: less than 30 minutes
• Very low elution volumes: 10 < x < 50µl

A typical mammalian cell contains 10–30 pg total RNA. The majority of RNA molecules, however, are tRNAs and rRNAs. Depending on the cell type, tissue and metabolic state, mRNA accounts for 1–5% of the total cellular RNA. Due to the low proportion of mRNA in the total cellular RNA pool, reducing the amount of rRNA and tRNA in a total RNA preparation increases the relative amount of mRNA. The mRNA enrichment is essential for construction of cDNA libraries and other applications where pure mRNA is highly desirable. The probability of selecting the right clone is greatly increased by reducing the amount of unwanted rRNA and tRNA. With pure, intact mRNA preparations, even low level messengers can easily be detected by in vitro translation, northern hybridization, Nuclease S1 protection analysis, expression-array and expressionchip analysis, or SAGE™ technology. Isolation of pure, intact mRNA is of great importance when characterizing mRNA species with these techniques. The use of the kit relies on the base pairing between the poly A tail of the mRNA and the oligonucleotides (dT) sequence bound to the Nucleo-Adembeads. Nucleo-Adembeads consists of superparamagnetic nanoparticles of uniform and a perfect spherical shape. Oligonucleotides (dT)25 are linked to the surface of the superparamagnetic nanoparticles via Biotin-Streptavidin system. The true spherical shape eliminates non-specific binding associates with irregulary shape particles. After processing the protocol using Nucleo- Adembeads, the mRNA is ready to use for downstream applications such as RT-PCR, Nothern Blotting, cDNA library construction, nuclease protection assay, in vitro translation, primer extension, subtractive cDNA cloning and reverse transcription.

• Prepare Biotinylated Oligo(dT)25 solution
• Prepare Beads
• Lysis step
• Bind to [Oligo (dT)25 – Bio-AdebBeads Streptavidin] complex
• Wash [Oligo(dT)25 - Bio-Adembeads Streptavidin] complex
• Elute mRNA

Beads	0.6mL Bio-Adembeads
Buffers	60 mL Lysis Buffer	4 mL Binding Buffer	12 mL Washing Buffer	2 mL Nuclease free Water
Other	Lyophilisate Biotinylated Oligo(dT)25
Storage	Store at 2-8°C.

• mRNA purification from cell lysate or whole blood (each from up to 1 Million Cells or 200 ul Blood)