Robust and high-performance hydrolysis probe-based detection
QuantiNova LNA Probe PCR Assays use FAM-labeled hydrolysis probe-based detection, which is highly sensitive and robust, because only the desired PCR product is detected. Probe-based detection still requires high PCR specificity to prevent amplification artifacts such as non-specific PCR products and primer–dimers that can compete for reaction components and compromise performance. However, we’ve already optimized the QuantiNova LNA Probe PCR system for you to eliminate non-specific amplification and give you accurate and sensitive detection every time.
Most comprehensive and specific coverage
Our proprietary algorithm has been used to design over 1.3 million QuantiNova LNA Probe PCR Assays to provide the most sensitive, accurate and effective mRNA and lncRNA analysis. The predesigned assays cover most transcripts in the Ensembl database for human, mouse and rat genes, enabling PCR-based gene expression studies in the greatest depth possible. Most of the assays are intron-spanning when possible and detect only RNA. Assays that do not span an intron are designated as such, and if there is one exon in the target, unwanted signals can be easily eliminated using the QuantiNova Reverse Transcription Kit with the integrated gDNA removal step.
Choosing the right assay for your target
Predesigned QuantiNova LNA Probe PCR Assays let you accurately and sensitively detect any human, mouse or rat mRNA or lncRNA, no matter what level of analysis you need: general transcript detection, detection or a specific transcript or differentiation of transcript isoforms.
Most human, mouse and rat genes have only one transcript, but for those with multiple transcripts, we selected up to three assays per transcript using the same algorithm. The design and primer positioning of these assays differ to match various usage requirements. Our assay selection guide helps you quickly identify the best assay from each category. After searching for assays for your target, simply review the results and look for the recommended assay that matches your intended use:
- Best coverage: for general transcript detection
- Best transcript assay: for detection of a specific transcript
- Best transcript-specific assay: for differentiation between specific transcript isoforms
Refer to the table below for more details.
| ||Gene-covering assays ||Transcript-specific assays |
|QIAGEN’s recommended assay ||Marked as “Best coverage” ||Marked as “Best transcript assay” ||Marked as “Best transcript-specific assay” |
|Description of assay design and coverage ||Covers most of the biologically relevant transcripts of the given gene ||Highly optimized to specifically detect the transcript of interest, targeting more isoforms of the transcript ||Splice variant transcript-specific |
|When to choose ||For general transcript screening and determining whether the gene of interest is expressed in a sample; for detection of as many transcripts and isoforms of a gene as possible ||For detection of a specific transcript ||For differentiation between specific isoforms of a transcript |
Complimentary data analysis
The complimentary, web-based data analysis tool in the GeneGlobe Data Analysis Center includes a user-friendly wizard to guide you step-by-step through the normalization and analysis of your data and generates publication-ready results figures.
Reference Gene Assays for any study
A wide selection of human, mouse and rat Reference Gene Assays are available to enable high-quality data normalization and ensure reliable results. These assays target endogenous coding RNAs, long non-coding RNA and small nucleolar RNA molecules that are typically constitutively expressed in a wide variety of tissues. The Reference Gene Assays are FAM labeled and have been functionally validated as reference genes for the QuantiNova LNA PCR system and work optimally with the QuantiNova RT and PCR reagents.
Normalization of mRNA/lncRNA qPCR results
Normalization removes technical and biological inter-sample variation unrelated to the biological changes under investigation. Proper normalization is critical for correct analysis and interpretation of results from real-time PCR experiments. Most commonly, stably expressed reference genes are used for normalization.
It is generally recommended to test several endogenous control reference gene candidates before setting up your actual mRNA/lncRNA expression analysis. These candidates should be chosen from genes expected to be stably expressed over the whole range of samples under investigation. They could be stably expressed mRNAs or lncRNAs selected based on literature or preexisting data (e.g., NGS or qPCR panel screening). The QuantiNova LNA PCR system offers validated reference gene assays for RNAs that tend to be stably expressed and are therefore good candidates as reference genes.
All reference gene candidates should be empirically validated for each study. One option for normalizing PCR panel when profiling a large number of mRNAs/lncRNAs is to normalize against the global mean – the average of all expressed mRNAs/lncRNAs. This can be a good option in samples with a high call rate (expressed genes) but should be used with caution in samples with low call rates. It is also not a good option in samples for which the general gene expression level is changed. Further guidance on normalization can also be found in the GeneGlobe Data Analysis Center.