Partek Flow Documentation

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by variance: the analysis will give more emphasis to the features with higher variances. This is the default option for e.g. single cell RNA-seq data

In the advanced options settings, more detailed information generated from the analysis can be reported:

Generate PCA table: checking this option will output PC projections in a data node. 

Generate PC quality measures: checking this option will output eigenvalues, PC projections, and component loadings. Component loadings are the correlation coefficients between the features and PCs.

Generate mapping error statistics: checking this option will output the mapping error information for the first three PCs.

If the input data node is in linear scale, you can perform log transformation on PCA calculation. 


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SubtitleTextPCA setup dialog
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The PCA task creates a new task node, and to open it and see the result, do one of the following: select the PCA task node, proceed to the context sensitive menu and go to the Task result; or double-click on the PCA task node.

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SubtitleTextRNA-Seq workflow with principal components analysis (PCA) task invoked on the Gene counts data node
AnchorNamernaseq-pca

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Also note that PCA is available for ERCC assessment; open the QA/QC on ERCC controls task node and push View PCA in the lower left corner.

The results are presented as scatterplot  (Figure 3), with each dot on the plot being a sample, while the axes represent the PCs and the axes values correspond to the respective PC values. By default, The report containing eigenvalues, PC projections, component loadings, and mapping error information for the first three PCs. 

When open PCA node in Data viewer, by default, it is 3D scatterplot  (Figure 2), each dot on the plot represents an observation, while the first three PCs are shown on the X-, Y-, and Z-axis respectively, with the information content of an individual PC is in the parenthesis. 

As an exploratory tool, PCA scatterplot is applied to view any groupings in the data set and generate hypotheses based on the outcome, or to spot possible outliers.

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SubtitleTextPrincipal components analysis plot in 3D. Each dot is a sample. The axes show the first three principal components, with the fraction of explained variance in the parenthesis. The legend is on the right, showing effect of coloring and sizing on the appearance of the dots (an example)
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To rotate the plot left click & drag. To zoom in or out, use the mouse wheel. Click and drag the legend can move the legend to different location on the viewer.

The Detailed configuration on PCA plot can be customised by using the controls on the left (Figure 4). Color by shows the sample attributes as listed in the Data tab or you can set it to Fixed to have all the dots of the same color. Size by option works in the same way, but affects the dot sizes.found by clicking Help>How-to videos>Data viewer section.

In Data viewer, when select PCA data node is the Data section on the control panel (left panel), when drag the node to the plot section (Figure 3), you will have option to plot scree plot and tables. 

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SubtitleTextControl panel of principal components analysis Drag PCA data node to plot
AnchorNamepca-controlsdragPCA

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Connect by option is particularly useful for dependent study designs, where you can highlight the samples based on the same biological source by the connecting lines. Example on Figure 5 depicts results of a study where each RNA sample was processed by both RNA-seq and  gene expression array; the lines connect the same samples.

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SubtitleTextPrincipal components analysis plot in 3D. Each dot is a sample and samples originating from the same biological source (dependent study design) are connected by lines. The axes show the first three principal components, with the fraction of explained variance in the parenthesis
AnchorNamepca-3d

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When choose Scree plot icon Image Added, it will plot a 2D viewer, X-axis represents PCs, Y-axis represents eigenvalues (Figure 4)

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SubtitleTextCustomize colors dialog (default appearance). General tab (palette) is used for sample coloring and general sample attribute-based coloring in the chromosome view, hierarchical clustering, and general charts and graphs. Two-color numeric tab (palette) is used to color by numeric sample attribute in the hierarchical clustering, principal components analysis, and dot plot views. Save button saves your color preferences
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When click on a dot (sample), it will be selected and a label of the sample will be displayed, from the Label by drop-down list to select how to label the selected sample. To select more than one  samples,  press Ctrl & click.

Next, Show legend turns the legend on or off. Select all selects all the dots, while Show axis turns the coordinate axis on or off. To change the size of the dots, use the Dot size slider. Grid cells increases or decreases the size of the cells in the plot grid and the Major grid interval specifies the frequency of major grid lines (fat lines). E.g. setting the Major grid interval to 4 highlights every 4th grid line.

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Scree plot
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When mouse over on a point on the line, it will display detailed information of the PC. The scree plot shows how much variation each PC represents,  so it is often used to determine the number of principal components to keep for downstream analysis (e.g. tSNE, UMAP, graph-base clustering). The "elbow" point of the graph where the eigenvalues seem to level off should be considered as a cutoff point for downstream analysis.

PCA data node can also be draw as tables, when choose Table icon (Image Added), it will display the component loadings matrix in the viewer (Figure 5)

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SubtitleTextPrincipal components analysis plot in 2D. Each dot is a sample. The axes show the first two principal components, with the fraction of explained variance in the parenthesis (an example)
AnchorNamepca-2d

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Although first three PCs are shown by default, you can plot any of the first nine PCs, by using the X, Y, and Z drop-down lists.

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Component loadings are the correlation coefficients between the features and PCs.
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In the table, each row is a feature, the column represent PCs, the value is the correlation coefficient. The table can be downloaded as text file. In the configuration panel, the data table drop-down list in the content section, there is PCA projects option,  change to this option to display the projection table (Figure 6).

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SubtitleTextSave image dialog (default settings)PCA project table
AnchorNamesave-imageprojection

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In this table, each row is an observation, each column is a PC, the values are the PC scores. The table can be downloaded as text file.


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