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The qBiomarker Mutation PCR Arrays and Assays are designed to detect specific mutations reported in genes associated with disease processes. The mutations are selected from comprehensive curated databases (such as the COSMIC database), literature reviews, frequency of occurrence and functional relevance See the complete list of of Mutation Arrays and Assays.

"If you have access to a real-time PCR instrument, you can use PCR Arrays"

How it Works Performance Data Application Data

How it Works

Overview of the qBiomarker Somatic Mutation PCR Array / Assay Protocol.

The procedure involves DNA extraction (QIAGEN QIAamp DNA Mini Kit or FFPE Tissue Kit is recommended), an optional amplification (QIAGEN REPLI-g kit or REPLI-g UltraFast kit is recommended) step for DNA isolated from fresh samples, qPCR detection on qBiomarker Somatic Mutation PCR Arrays or Assays, and data analysis (using the qBiomarker Somatic Mutation Data Analysis Template). An optional DNA sample QC step immediately before the detection array or assay setup allows the user to qualify the DNA samples. (Download user manual)

How qBiomarker Somatic Mutation PCR Arrays/Assays Work

The procedure involves DNA extraction (QIAGEN QIAamp DNA Mini Kit or FFPE Tissue Kit is recommended), an optional amplification (QIAGEN REPLI-g kit or REPLI-g UltraFast kit is recommended) step for DNA isolated from fresh samples, qPCR detection on qBiomarker Somatic Mutation PCR Arrays or Assays, and data analysis (using the qBiomarker Somatic Mutation Data Analysis Template). An optional DNA sample QC step immediately before the detection array or assay setup allows the user to qualify the DNA samples.

To complete the Somatic Mutation PCR Array procedure, start with 5 to 10ng genomic DNA isolated from fresh tissues, or as low as 200 ng DNA from FFPE sections. DNA from fresh tissues can be uniformly amplified using QIAGEN REPLI-g UltraFast Kit. Then, mix your DNA with the included ready-to-use qBiomarker Probe PCR Master Mixes and aliquot the mixture into each well of the same plate containing pre-dispensed gene-specific primer and hydrolysis probe sets. By performing real-time PCR, the mutation status of a particular sample is determined by comparing the allele specific Ct values between your test sample and a wild-type control sample (see qBiomarker Somatic Mutation Data Analysis section for detailed principles).

Layout and Controls: Each array contains a panel of 5' nuclease assays for a stringently selected set of pathway focused somatic mutations, gene copy number controls, and PCR quality controls. The PCR Arrays are available in both 96-well and 384-well plate formats, containing either one or four replicates, respectively, of the 96-assay set panel. SABiosciences' qBiomarker Somatic Mutation Assays and qBiomarker Probe Master Mixes have been pre-optimized hand-in-hand for 5' nuclease based real-time RT-PCR detection. The simplicity of the qBiomarker Somatic Mutation PCR Array format and operating procedure allows routine somatic mutation profiling in any research laboratory with access to real-time PCR instruments.

Example Layout of Pathway-Focused qBiomarker Cancer Somatic Mutation PCR Array (96-well format)

Performance Data Sensitivity:

The complete PCR Array System yields a greater-than 85 percent present call with as little as 25 ng as much as 5 g of total RNA from a pathway representing genes expressed at a lower level (inflammatory cytokines and receptors).

Application Data

qBiomarker Somatic Mutation - Pathway-Focused Cancer Cell Profiling

Mutation profiling of cancer cell line DNAs on the qBiomarker Somatic Mutation EGFR Pathway PCR array. 200ng each of genomic DNA from a wildtype control cell line (WT) and 7 well characterized cancer cell lines were subject to profiling on the qBiomarker Somatic Mutation EGFR Pathway PCR array on an ABI 7900HT sequence detection system. Raw Ct data were analyzed with the qBiomarker Somatic Mutation PCR Array Data Analysis template using DDCt method. Each spike represents the presence of a mutation at each locus (x-axis) in each sample (y-axis).

Somatic Mutation Profiling of Cancer - Correlation with PyroSequencing

Human lung cancer FFPE samples profiled on qBiomarker Somatic Mutation EGFR Pathway PCR array. Genomic DNA from 10 FFPE tissue samples (adenocarcinoma) were subject to profiling on the qBiomarker Somatic Mutation EGFR Pathway PCR Array on an ABI 7900HT sequence detection system. Raw Ct data were analyzed with the qBiomarker Somatic Mutation PCR Array Data Analysis template using average Ct method. Each spike represents the presence of a mutation at each locus (x-axis) in each sample (y-axis).

(B) PyroMark pyrosequencing verifies the deletion and mutations identified by mutation PCR array. Two QIAGEN PyroMark pyrosequencing assays (one for EGFR c.2236_2250del15, one for KRas codon 12 and 13) confirmed the identities of the following mutant alleles in remaining DNA samples: samples 266-1 and 3237-1: EGFR c2236-2250del15 deletion; samples 276-2 and 296-1: KRas c.35G>T mutation. The pyrosequencing results further confirmed that the mutant alleles occur in only a fraction of the cells. A KRas c.37G>C (p.13G>R) mutation was detected by mutation profiler PCR array in sample 264-4, but the percentage of the mutation in this sample appears to be below the detection limit of the pyrosequencing method.

The qBiomarker Somatic Mutation PCR Arrays & Assays are intended for molecular biology applications. This product is not intended for the diagnosis, prevention, or treatment of a disease

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