ValidPrime® replaces the need to perform no reverse transcriptase (RT(-)) controls to test for the presence of genomic DNA (gDNA) for all samples in your reverse transcriptase quantitative PCR (RT-qPCR) profiling.
In an expression profiling experiment based on m samples and n assays, traditional set up requires m RT(-) reactions plus m x n qPCR controls.
When using ValidPrime® only m n 1 controls are needed.
ValidPrime® will minimize the amount of control reactions and hence your costs as well as your efforts.
ValidPrime® is an assay to test for the presence of gDNA in test samples and when combined with a gDNA control sample, replaces all RT(-) controls. ValidPrime® is highly optimised and specific to a non-transcribed locus of gDNA that is present in exactly one copy per haploid normal genome. The sequence detected by the ValidPrime® assay has no transcriptional activity and is not present in pure cDNA preparations. Therefore, ValidPrime® measures the number of genomic copies present in a sample.
In an expression profiling experiment the ValidPrime® assay is added to the list of assays and the gDNA control is added to the list of samples. From the combined measurements of the ValidPrime® assay and the gene of interest (GOI) assays on all samples and on the gDNA control, the genomic background contribution to all RT-qPCR measurements can be assessed. ValidPrime® replaces the need to perform RT(-) controls for all reactions and makes RT-qPCR profiling easier and substantially cheaper.
The assay has very high PCR efficiency (E > 90% in tested commercial master mixes) and produces negligible amount of primer dimer products. Limit of detection (LOD) is estimated to 4 copies of DNA (0.01 ng of DNA) and limit of quantification (LOQ) is estimated to 32 copies
of DNA (0.08 ng of DNA).
For ValidPrime® Human, ValidPrime® Mouse, and ValidPrime® Rat, a gDNA standard is provided which can be used to test the sensitivity of qPCR assays for gDNA background. ValidPrime® Vertebrate does not include a gDNA standard, since the standard needs to be specific for the species analysed and arranged by the user.
If you cannot find the answer to your problem then please contact us or telephone +44 (0)1954 210 200
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