VGAS Tutorials: TCR Analysis
Basic Analysis
CLICK the headings below to view the tutorial video.
GENE FREQUENCY
The Visual Genomics Analysis Suite (VGAS) can be used to compare the frequency of T-cell receptor V, D and J genes in a range of samples.
Once data files are loaded into the TCR/BCR homepage, user can select samples of interest and double-click on the “Gene Frequency” analysis in the right-hand column.
This opens a window displaying a Gene Frequency Histogram with genes on the x-axis and counts on the y-axis.
Axes can be displayed as linear or logarithmic scale, or as a percentage.
Gene sequences can be filtered for productive, out of frame, and/or has stop.
Options for V, D, or J gene chains.
Histogram size and block width can be modified using sliding scales.
Options to display or hide combinations of samples, legend can be added by selecting ‘Display Legend’ tick box.
Option to show results grid, showing gene and respective count numbers for different samples. Grid can be exported to Excel.
TOP CLONES
The Visual Genomics Analysis Suite (VGAS) can be used to highlight T-cell receptor (TCR) clones in a sample, to identify dominant clonotypes.
Once data files are loaded into the TCR/BCR homepage, user can select sample of interest and double-click on the “Top Clones” analysis in the right-hand column.
This opens a window displaying a Top Clones Histogram with Top 10 Clones on x-axis, and Clone Count on y-axis.
Hovering mouse over histogram bar displays information about clone (amino acid sequence, V and J chains, count and percent).
Scale can be set as linear or logarithmic, or as a percentage.
Gene sequences can be filtered for productive, out of frame, and/or has stop.
Number of top clones displayed on x-axis can be specified by typing a number into “Number of Top Clones”.
Histogram size and block width can be modified using sliding scales.
Options for display of axes including intervals or hiding scales via tick boxes.
Option to show results grid, showing top clone number, count, percent, amino acid, and V and J genes. Grid can be exported to Excel.
CLONE TRACKER
The Visual Genomics Analysis Suite (VGAS) can be used to highlight identical T-cell receptor clones between cell samples of interest.
Once data files are loaded into the TCR/BCR homepage, user can select samples of interest and double-click on the “Clone Tracker” analysis in the right-hand column.
This opens a window displaying a Clone Tracker Plot with sample names on x-axis, and Clone Count on y-axis, with each coloured line representing a DNA or amino acid sequence (changeable under “Compare Using”).
Plot size and track width can be modified using sliding scales.
Scale can be set as linear or logarithmic, or as a percentage.
Can display legend by selecting “Display Legend”.
Results Grid shows Amino Acid, Total Count, and then Counts per sample. Double-clicking on an amino acid/DNA sequence in results grid list will highlight the respective track on plot.
Sequence filters of Number of Top Clones or Clones Greater than % can be applied. Single counts can be removed.
CDR3 LENGTH
The Visual Genomics Analysis Suite (VGAS) can be used to analyse CDR3 lengths of T-cells within and between sample T-cell populations.
Once data files are loaded into the TCR/BCR homepage, user can select samples of interest and double-click on the “CDR3 Length” analysis in the right-hand column.
This opens a window displaying a CDR3 Length Histogram with CDR3 lengths on x-axis, and counts on y-axis. Legend can be displayed or hidden.
Plot size and block width can be modified using sliding scales.
Hovering over blocks shows information breaking down CDR3 length in total and in individual samples.
Scale can be linear or logarithmic.
Sequence filters including productive, out of frame and has stop can be applied.
Plot start and end can be specified to alter x-axis minimum and maximum.
Results grid indicating CDR3 length, and counts per sample can be shown and exported to Excel.
Individual samples can be included or excluded by selecting tick boxes under “Colour/Display”.
SCATTER PLOT
The Visual Genomics Analysis Suite (VGAS) can be used to compare T-cell DNA or amino acid sequences between samples of interest.
Once data files are loaded into the TCR/BCR homepage, user can select two samples of interest and double-click on the “Scatter Plot” analysis in the right-hand column.
This opens a window displaying a Linear Scatter plot with count of ‘Sample X’ on the x-axis, and count of ‘Sample Y’ on the y-axis.
Scale can be changed to linear, logarithmic or as a percentage. “-/+ Minimum Log Scale” can be used to alter logarithmic scale. Linear scale increments can be similarly altered using drop-down menu next to “Linear Scale”.
Samples can be compared on the basis of nucleotide (DNA) or amino acid sequence.
Plot size, point size and point depth can be modified using sliding scales.
Results Grid displays DNA or amino acid sequence, with counts in each sample.
Hovering over a point in the scatter plot shows information about sequence and counts.
Options to display or hide points for ‘Sample X’ series, ‘Sample Y’ series, and/or Sequences in both. Colours for series points can be selected by clicking on box next to series name.
Pearson Coefficient can be automatically calculated and plotted, with r and r-squared values recorded in legend.
SAMPLE OVERLAP
The Visual Genomics Analysis Suite (VGAS) can be used to compare T-cell DNA or amino acid sequences between samples of interest.
Once data files are loaded into the TCR/BCR homepage, user can select samples of interest and double-click on the “Sample Overlap” analysis in the right-hand column.
This opens a window displaying a Sample Overlap grid with squares coloured depending on level of overlap between samples. Samples listed down sides of grid, and r-squared and overlap (o) values printed inside squares. Values can be hidden by unselecting tick boxes.
User can compare samples based on nucleotide (DNA) or amino acid sequences.
Sequence filters of productive, out of frame, and has stop can be applied.
Grid block size and label size can be modified using sliding scales.
Heatmap colours can be changed to options displayed in left-hand panel, or removed altogether.
COMMON SEQUENCES
The Visual Genomics Analysis Suite (VGAS) can be used to compare T-cell DNA or amino acid sequences between samples of interest.
Once data files are loaded into the TCR/BCR homepage, user can select samples of interest and double-click on the “Common Sequences” analysis in the right-hand column.
This opens a window displaying a list of Common Sequences, total counts, and counts per sample. These can also be shown as a percentage.
Samples can be compared using nucleotide (DNA) or amino acid sequences.
Sequences can be filtered on basis of productive, out of frame, and has stop. Additionally, common sequences can be filtered based on intersection or union between samples.
SAMPLE SUMMARY
The Visual Genomics Analysis Suite (VGAS) can be used to compare T-cell clone sequences between samples of interest.
Once data files are loaded into the TCR/BCR homepage, user can select samples of interest and double-click on the “Sample Summary” analysis in the right-hand column.
This opens a window displaying a summary of the sample(s). Including a breakdown of productive, out of frame, has stop, and total sequences shown as number of sequences, % of total sequences, number of unique sequences, and % of unique sequences.
Additionally, this is further broken down into individual sample summaries
Advanced Analysis
CDR3 Well/Plate Explorer
Paired Gene Frequency
Setting Default Colours
