GeneGlobe Data Analysis Center
For analyzing and interpreting real-time PCR or NGS data
The GeneGlobe Data Analysis Center is a web resource for scientists analyzing their real-time PCR or NGS data. The real-time PCR modules transform threshold cycle (CT) values to calculated results for gene and miRNA expression, somatic mutation detection and copy number measurements. The NGS module supports the analysis of QIAGEN’s GeneRead target enrichment panels. Register on QIAGEN.com to access this complimentary suite of tools to accelerate your data analysis and interpretation.
Fold regulation comparison and p-value
Fold-Change (2^ (- Delta Delta CT)) is the normalized gene expression (2^(- Delta CT)) in the Test Sample divided the normalized gene expression (2^ (- Delta CT)) in the Control Sample. Fold-Regulation represents fold-change results in a biologically meaningful way. Fold-change values greater than one indicates a positive- or an up-regulation, and the fold-regulation is equal to the fold-change. Fold-change values less than one indicate a negative or down-regulation, and the fold-regulation is the negative inverse of the fold-change. Fold-change and fold-regulation values greater than 2 are indicated in red; fold-change values less than 0.5 and fold-regulation values less than -2 are indicated in blue. The p values are calculated based on a Student’s t-test of the replicate 2^ (- Delta CT) values for each gene in the control group and treatment groups, and p values less than 0.05 are indicated in red.
The scatter plot compares the normalized expression of every gene on the array between the two selected groups by plotting them against one another to quickly visualize large gene expression changes. The central line indicates unchanged gene expression. The dotted lines indicate the selected fold regulation threshold. Data points beyond the dotted lines in the upper left and lower right sections meet the selected fold regulation threshold.
The volcano plot helps quickly identify significant gene expression changes. The volcano plot displays statistical significance versus fold-change on the y- and x-axes, respectively. The volcano plot combines a p-value statistical test with the fold regulation change enabling identification of genes with both large and small expression changes that are statistically significant.
The clustergram performs non-supervised hierarchical clustering of the entire dataset to display a heat map with dendrograms indicating co-regulated genes across groups or individual samples.
The heat map provides a visualization of the fold changes in expression between the selected groups for every gene in the array in the context of the array layout. The table provides the fold regulation data used for the map as well as the Comments associated with each one.
The data analysis web portal calculates fold change/regulation using delta delta CT method, in which delta CT is calculated between gene of interest (GOI) and an average of reference genes (HKG), followed by delta-delta CT calculations (delta CT (Test Group)-delta CT (Control Group)). Fold Change is then calculated using 2^ (-delta delta CT) formula. The data analysis web portal also plots scatter plot, volcano plot, clustergram and heat map.
CT values are exported to an Excel file to create a table of CT values. This table is uploaded on to the GeneGlobe data analysis web portal. Samples are assigned to controls and test groups. CT values are normalized using a manual or automatic normalization method. Results are then analyzed by plots and charts (scatter, volcano, clustergram and heat map).