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Oligotex mRNA Midi Kit

For mRNA purification from total RNA and for in vitro transcript cleanup
  • High recovery of pure mRNA in as little as 30 minutes
  • No oligo-dT cellulose or ethanol precipitation
  • Flexibility for use with widely varying amounts of starting RNA
Oligotex mRNA Kits enable purification of mRNA from total RNA and cleanup of poly A+ in vitro transcripts. Each Oligotex Kit includes Oligotex resin for binding poly A+ mRNA, and spin columns for washing and eluting bound mRNA. For purification of mRNA from cells and tissues, Oligotex Direct mRNA Kits are available.
Cat No./ID: 70042
Oligotex mRNA Midi Kit (12)
€866.00
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For 12 mRNA midipreps: 700 µl Oligotex Suspension, Small Spin Columns, Collection Tubes (1.5 ml), RNase-Free Reagents and Buffers
The Oligotex mRNA Midi Kit is intended for molecular biology applications. This product is not intended for the diagnosis, prevention, or treatment of a disease.
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Isolation of mRNA from total RNA.
Formaldehyde agarose gel and northern blot (GAPDH-probed) of poly A+ mRNA isolated from total RNA from various samples using Oligotex mRNA Kits.
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Oligotex particles and oligo-dT cellulose.
Scanning electron micrographs of Oligotex particles and oligo-dT cellulose at the same magnification. Note the uniform size and shape of Oligotex particles.
Performance

Oligotex mRNA kits are available for isolation of mRNA from less than 0.25 mg up to 1 mg of total RNA starting material (see Table).

Poly A+ mRNA purified with Oligotex resin is suitable for RT-PCR (see figure "RT-PCR of beta-actin mRNA") and poly A+ mRNA isolated from total RNA with Oligotex resin has been used for Northern, dot, or slot blotting (see figure "Isolation of mRNA from total RNA").

Table. Specifications of Oligotex mRNA kits
Amount of starting material (total RNA) per prep  Maximum number of preps per Oligotex mRNA Mini Kit* Maximum number of preps per Oligotex mRNA Midi Kit*
≤0.25 mg 12 (13) 12 (28)
0.25—0.5 mg 6 12 (14)
0.5—0.75 mg 4 12 (14)
0.75—1 mg 3 12
* Using the spin-column protocol. Batch protocols also available.
 Maximum number of preps per kit with purchase of additional spin columns. Additional 1.5 ml microcentrifuge tubes may be required.
Principle

Oligotex mRNA Kits contain spin columns and all necessary reagents and buffers for isolation of pure poly A+ mRNA from total RNA preparations or cleanup of in vitro transcripts. Oligotex resin consists of polystyrene–latex particles of uniform size (1.1 µm diameter) and a perfect spherical shape (see figure "Oligotex particles and oligo-dT cellulose") with dC10T30 oligonucleotides covalently linked to the surface. The size, composition, and surface structure of Oligotex particles have been optimized for uniform dispersion and minimal centrifugation time. The particles form a stable suspension that provides a large surface area for rapid and efficient binding of polyadenylic acids.

The high capacity and accuracy of hybridization provides superior purification of poly A+ mRNA through fast and efficient hybridization. mRNA recoveries are consistently greater than 90%. The Oligotex purification procedure minimizes loss of poly A+ RNA, eliminates the risk of degradation by RNases, and requires minimal hands-on time. This makes it ideal for simultaneous handling of multiple samples. Poly A+ mRNA is immediately ready for all standard applications, as well as for sensitive applications such as expression array, differential display, cDNA library construction, microinjection, and SAGE technology. Oligotex Suspension is also available separately.

Procedure

Optimized Oligotex purification procedures are convenient and straightforward (see flowchart "Oligotex mRNA procedure"). Total RNA preparations are incubated with Oligotex resin, and Oligotex:mRNA complexes are collected by a brief centrifugation step. Spin columns, supplied in the Oligotex Kits, provide the most convenient handling, and optional batch protocols are also provided for certain applications. After washing, the mRNA is eluted in a small volume of Tris buffer or water. Alcohol precipitation is not required.

Applications

Poly A+ mRNA purified with Oligotex resin is ready for use in any downstream application, including:

  • RT-PCR
  • cDNA synthesis
  • cDNA library construction
  • Subtractive hybridization
  • In vitro translation
Features
Specifications
Applications PCR, real-time PCR, microarray, transfection
Elution volume 20–100 µl
Format Spin column
Main sample type Total RNA
Processing Manual (centrifugation)
Purification of total RNA, miRNA, poly A+ mRNA, DNA or protein Poly A+ mRNA
Sample amount 250 µg – 1.0 mg
Stabilization Yes
Technology Polystyrene-latex particles
Time per run or per prep 32–50 minutes
Yield >90% recovery

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Kit Handbooks (1)
For Purification of poly A+ RNA from total RNA, purification of poly A+ RNA directly, from cultured cells or tissues, and purification of polyadenylated in vitro transcripts 
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User-Developed Protocols (1)
The procedure has been used successfully by customers for regeneration of Oligotex resin from spin columns following elution of poly A+ mRNA.
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