RNA Extraction for Real-Time PCR
RNA real-time PCR is the most sensitive technique for mRNA detection and quantitation currently available. Compared to the two other commonly used techniques for quantifying mRNA levels—northern blot analysis and RNase protection assay—real-time PCR can be used to quantify mRNA levels from much smaller samples. One key variable to the success of any real-time PCR experiment is optimal RNA extraction. The one issue all RNA isolation methods have, however, is the inability to completely remove genomic DNA contamination from the RNA sample. DNA removal is critical especially for RT-PCR applications since DNA can serve as a template during the PCR portion of the experiment, resulting in false positives, background, etc. Ambion® offers products to simplify your RNA real-time PCR experiments and make the data more quantitative without the risk of genomic DNA contamination.
Eliminate Genomic DNA Contamination
Genomic DNA contamination can lead to false positive RT-PCR results. We offer a variety of Ambion® tools for minimizing genomic DNA contamination from RNA samples prior to RT-PCR. For example, DNA-free™ DNase Treatment and Removal Reagents are designed for removing contaminating DNA from RNA samples and for the removal of DNase after treatment without Proteinase K treatment and organic extraction. In addition, we offer TURBO DNase™ enzyme kits, a hyperactive enzyme engineered from wild-type bovine DNase. The proficiency of TURBO DNase™ enzyme in binding very low concentrations of DNA means that the enzyme is particularly effective in removing trace quantities of DNA contamination. To prevent cross contamination during PCR experiments, we also offer DNAZap™ DNA Degradation Solution and RNase-free barrier pipette tips.
- Quantitating Gene Expression Directly from Cell Lysates Using TaqMan® Real-Time PCR Analysis - TechNotes 14(1)
- Real-Time RT-PCR Directly from Cell Lysates: A Complete Workflow without RNA Purification - TechNotes 14(3)
- RecoverAll™ Total Nucleic Acid Isolation Kit