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WNT Signaling Pathway Copy Number PCR Array

Human
 
Human WNT Signaling Pathway Copy Number PCR Array
The Human WNT Signaling Pathway qBiomarker Copy Number PCR Array profiles the copy number of 23 WNT signaling-related genes reported to undergo frequent genomic alterations. Genes selected include WNT pathway components, molecules/pathways that cross-talk with the WNT pathway, and WNT pathway effectors. These genes encode cell adhesion molecules, receptors, transcription factors and other signaling proteins that regulate processes such as the cell cycle, migration, cancer, and cellular differentiation, as well as developmental disorders. DNA copy number changes in WNT-related genes have been reported in multiple cancers. Genes were chosen from the most frequently amplified or deleted genes relevant to WNT signaling based on the primary literature and public databases. This array may serve as a useful tool to help classify samples by genotype and help verify phenotypic biomarkers. The array analyzes each gene in each sample in quadruplicate and includes a stable multi-copy reference assay for accurate copy number determination via appropriate DNA input normalization. The simplicity of the product format and operating procedure allow routine and reliable copy number profiling in any research laboratory with access to a real-time PCR instrument.

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

96-well Plate, 384-well (4 × 96) Plate, and 100-well Disc formats are available.

 

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Modify this Array    Gene Table

Catenin: CTNND1.

Cell Adhesion: CDH1, CDH13.

Cell Cycle: CCND1.

Cytoskeleton Regulators: GRB7, MARK4.

Growth Factor & Hormone Receptors: AR, ERBB2, FGFR2.

Intracellular Signaling Molecules: APC, DVL1, NKD2, PTK6.

MicroRNA Genes: MIR17HG, MIR517C.

Positive Regulator of WNT Signaling: C8orf4.

Transcription Factors: AR, FOSL1, IRX2, JUN, MYC, MYCN, PLAGL2, TP53.

WNT Target Genes: CCND1, CDH1, FOSL1, JUN, MYC.

Modify this Array    Gene Table
 

Functional Gene Grouping How It Works Manual & Resources Reagents & Software
 

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

qBiomarker Copy Number PCR Arrays are the most reliable and sensitive copy number profiling technology for analyzing a panel of loci in signal transduction pathways or disease related gene networks. The Copy Number PCR Arrays can be used for research on cancer, neurobiology, immunology, biomarker discovery and validation. See the complete list of Copy Number PCR Arrays.

You can use any 96-well, 384-well plate or 100-well disc real-time PCR instrument. See instrument-specific setup instructions

How it Works Performance Data Application Data

How it Works

The Copy Number PCR array is a set of optimized real-time PCR primer assays on 96-well, 100-tube or 384-well plates for measuring changes in copy number. The PCR array performs copy number analysis with real-time PCR sensitivity and the multi-loci profiling capability of a microarray. Simply mix the genomic DNA sample with the appropriate ready-to-use PCR master mix, aliquot equal volumes to each well of the same plate, and then run the real-time PCR cycling program. (Download user manual)

Figure 1: How Copy Number PCR Arrays Work - Protocol Chart

The procedure involves isolating genomic DNA (QIAGEN QIAamp DNA Mini Kit or FFPE Tissue Kit is recommended), qPCR detection on qBiomarker Copy Number PCR Arrays or Assays, and data analysis (using the qBiomarker Copy Number Data Analysis). An optional DNA sample quality control step immediately before the detection array or assay setup allows the user to qualify the DNA samples.
To complete the Copy Number PCR Array procedure, start with 400 to1000 nanograms of genomic DNA isolated from fresh tissues, or as low as 800 nanograms of DNA from FFPE sections. Then, mix your DNA with the ready-to-use qBiomarker SYBR Green Mastermixes and aliquot the mixture into each well of the same plate containing pre-dispensed locus-specific primer assays. By performing real-time PCR, the copy number status of a particular sample is determined by comparing the CT values between your test sample and a wild-type control sample (see qBiomarker Copy Number Data Analysis section for detailed principles).

Why qBiomarker Copy Number Arrays?
     Guaranteed Performance* - ready-to-use for copy number analysis
     Time and cost saving - less than 30 minutes hands-on time for analyzing 23 loci
     Ease of data analysis using our easy-to-use Excel-based data analysis template or web-based analysis tool

Layout and Controls: The PCR Arrays are available in both 96-, 384-well plates and 100-well discs and are used to measure the copy number of 23 or 95 genes related to a disease state or pathway. Each gene/locus-specific assay is repeated in technical quadruplicate as indicated in the figure above. As an example, the first gene in the array is measured in wells A1, C1, E1 and G1. The qBiomarker Multicopy Reference Assay (MRef) is used to normalize the data for differences in the amounts of genomic DNA. The Multicopy Reference Assay is shown in purple in the above figure in wells B12, D12, F12, and H12.

You can easily perform a Copy Number PCR Array experiment in your own laboratory, or send your samples to us and take advantage of our PCR Array Services.

*:  when using qBiomarker SYBRŪ Green Mastermix.

Performance Data:
The qBiomarker Copy Number PCR Array System yields a more accurate representation of changes in copy number by using a multi-copy reference assay to normalize the raw target specific data.

qBiomarker Multicopy Reference Assay
The qBiomarker Copy Number Arrays have an integrated multi-copy reference assay for data normalization. The multi-copy reference assay  recognizes a stable sequence that appears in the human genome over 60 times, and whose copy number is not affected or minimally affected by local genomic changes. Inclusion of this reference assay during testing allows use of the ΔΔCT method to accurately make copy number calls or relative copy number change calls for specific targets.

 


qBiomarker Multicopy Reference Assay is superior to RNase P as a reference assay for copy number determination. Tumor cell line DNA (SKBR3) and reference genomic DNA were mixed in different ratios (100%, 87.5%, 75%, 50%, 25%, 12.5% and 0% SKBR3 cells respectively), and the DNA mixes were tested for GRB7 gene copy number, using either MRef assay or RNaseP assay as the reference. The GRB7 copy number in the reference genomic DNA is assumed to be 2. The "Expected" GRB7 gene copy numbers in 87.5%, 75%, 50%, 25%, 12.5% mixing ratio samples are calculated based on the GRB7 gene copy number in 100% SKBR3 genomic DNA sample, and the mixing ratio between the SKBR3 genomic DNA and Promega genomic DNA. The observed GRB7 gene copy numbers in general agree well with the expected values when using QIAGEN Multi-copy Reference Assay as the reference, while significant differences exist between the observed GRB7 gene copy numbers and the expected values when RNaseP is used as the reference assay.

RNaseP gene is not suitable as a normalizer of sample input in cancer cell line DNA samples. The absolute average copy numbers of RNaseP per normal genome copy amount of DNA were determined in two breast cancer cell line (SKBR3 and MCF7) genomic DNAs with the delta delta Ct method, using QIAGEN multi-copy reference assay as the normalization control of DNA input. The absolute copy number of RNaseP per normal genome in the genomic DNA  is assumed to be 2.

Universal Nature of Multicopy Reference Assay
The complete PCR Array System, with high quality input RNA, is guaranteed to yield single amplicons of the predicted size without primer dimers or other secondary products to ensure the most accurate real-time PCR results possible.

Stable Performance of the Multicopy Reference (MRef) Assay in 129 DNAs from 9 Major Human Populations. The qBiomarker multi-copy reference assay (MRef) and a qBiomarker copy number assay for RB1 were tested against DNAs from 129 healthy individuals from 9 major ethnic populations. Each assay and DNA sample combination was run in quadruple reactions. Delta CT between the average CT of the MRef assay and the RB1 assay was calculated for each individual DNA. The average delta Ct for samples within each ethnic population is plotted. Error bars show the standard deviation of the delta CT within each ethnic population. The RB1 gene is assumed to be present at 2 copies in all healthy individual DNAs.

Application Data

Aneuploidy Study


qBiomarker Copy Number Assays accurately identify aneuploidy. qBiomarker Copy Number Assays designed to target AR and MECP2 were tested against 4 cell line DNAs containing 1 copy (XY, Coriell NA13619), 2 copies (XX, Coriell NA01921), 3 copies (XXX, Coriell NA03623) and 4 copies (XXXX, Coriell NA11226) of X-chromosome. Chromosomal aberrations had been previously identified by cytogenetic methods. A control assay, targeting a stable, multi-copy region in the human genome, was used to normalize the amount of DNA input. ΔΔCT method was used to calculate the gene copy number, using XX (Coriell NA01921) as a 2-copy reference. Each assay was tested against each sample in quadruple replicate reactions, and a t-test was performed.

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Functional Gene Grouping How It Works Manual & Resources Reagents & Software
 
Handbook/User Manual qBiomarker Copy Number PCR Arrays and Assays (PDF)
Data Analysis Free Copy Number PCR Array Data Analysis Software (Web & Excel based)
Instrument Setup Instructions Materials & Equipment Required for our PCR products

PCR Array Instrument Setup Instructions & Protocol Files

FAQ FAQ for our Copy Number PCR products
Web Seminars Attend a live on-line seminar hosted by an Applications Scientist about PCR
Technical Brochures Copy Number PCR Array Brochure (PDF)
Powerpoint Presentations Copy Number Technology Overview Presentations
Pathway Central Pathway reviews and presentation-ready pathway maps
Individual Copy Number Assays Bench-Validated Copy Number qPCR assays for any locus in the genome
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Functional Gene Grouping How It Works Manual & Resources Reagents & Software
 

The complete qBiomarker Copy Number PCR Array System insures that you will have superior results from your pathway-focused copy number analysis experiment the first time, and every time guaranteed.

The Complete qBiomarker Copy Number PCR Array System Includes:

  • qBiomarker Copy Number PCR Arrays

  • qBiomarker SYBRŪ Green Mastermix
    Instrument-specific PCR master mixes insure high-efficiency, locus-specific amplification for the most accurate real-time PCR results.

  • qBiomarker Data Analysis Software (Excel & Web based)

Our convenient and high-quality PCR accessory products complete the qBiomarker PCR Array System and the qBiomarker Copy Number Assays

Other recommended accessory products:

QIAamp DNA Mini Kit (50)
Isolate genomic DNA from fresh or frozen samples for downstream real-time PCR array or assay analysis.

QIAamp DNA FFPE Tissue Kit
Isolate genomic DNA from fixed samples, including formalin-fixed paraffin-embedded (FFPE) blocks, paraffin blocks, sections, or slides, for downstream real-time PCR array or assay analysis.

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