mRNA Capture Kit
For life science research only. Not for use in diagnostic procedures.
The mRNA Capture Kit enables preparation of highly purified poly (A)+ RNA, which may be used directly for qualitative or quantitative RT-PCR.
- Flexible – Use kit from small to high numbers of samples and for semi-automatic procedures. Use kit from small to high numbers of samples and for semi-automatic procedures.
- Efficient - Minimize sample loss, handling time, and contamination risk because mRNA isolation and RT-PCR can be performed in the same tube.
- Reliable - Produce excellent RT-PCR templates, even from small amounts of starting material, or material containing low-abundance mRNAs.
- Lysis Buffer
- Oligo (dT)20, biotin-labeled, 20x concentrated
- Water, double-distilled, PCR Grade
- Wash Buffer
- Streptavidin-Coated PCR Tubes
- Caps for PCR Tubes
The mRNA Capture Kit isolates highly purified poly (A)+ RNA and immobilizes it in a PCR tube. The immobilized mRNA is suitable for:
- Qualitative and quantitative RT-PCR (see Figure 1)
Figure 1: Detection of MCAD and beta-actin transcripts in K-562 cells. Lysates from 106 cells were serially diluted. mRNA from the dilutions was captured in streptavidin-coated PCR tubes. RT-PCR was performed in the same PCR tubes with reagents from the Titan One Tube PCR System and primers derived from the human MCAD (medium-chain acyl-CoA dehydrogenase) and beta-actin genes. Amplicons were run on a 1% agarose gel.
Lanes 1, 10, 18: DNA Molecular Weight Marker VI
Lane 2: 1 x 104 cells
Lanes 3, 11: 2 x 103 cells
Lanes 4, 12: 400 cells
Lanes 5, 13: 80 cells
Lanes 6, 14: 16 cells
Lanes 7, 15: 3.2 cells
Lanes 8, 16: 0.64 cells
Lanes 9, 17: Control (no RT performed)
Result: The mRNA Capture Kit and Titan One Tube RT-PCR System allow efficient detection of beta-actin and MCAD mRNAs in as few as 3.2 K-562 cells.
Affinity purification is a versatile and highly specific technique for the purification of virtually all classes of biomolecules utilizing differences in biological activities of chemical structures. The high selectivity of this technique results in good purification and high recovery. Often a concentrating effect is reached which enables large volumes to be conveniently processed.
Typical Total RNA/mRNA Recovery
Starting Material and Quantity
mRNA (µg )
|Time Required *||Number of Reactions|
|Cultured cells (10 7 )||30-500||0.3-25||15 minutes||192 reactions|
|Mouse brain (100 mg)||200||7|
|Mouse liver (100 mg)||
|Mouse lung (100 mg)||130||10|
* Does not include time required for sample preparation.
A lysis step releases RNA from cultured cells or tissues The poly (A)+ tail of mRNA in the lysate hybridizes to a biotin-labeled oligo (dT)20. The hybrid is then immobilized on a streptavidin-coated PCR tube, followed by washing steps to remove unbound contaminants. The oligo (dT) part of this immobilized hybrid also serves as an RT-PCR primer for reverse transcriptase. The entire process (including RT-PCR) takes place in a single tube. The capture process requires approximately 30 minutes.
The kit is function tested and tested for the absence of RNase according to the current quality control procedures.