Rapid Development and Distribution of Statistical Tools for High-Throughput Sequencing Data

Radiant Genome Browser: Capabilities and usage

Epigenetics

The Radiant Genome Browser is able to load and visualize methyl seq analysed data from the MethylPipe Package, available from Bioconductor (Kishore, 2014). MethylPipe analyses base resolution DNA methylation data in both the CpG and non-CpG sequence context.

Track loading instructions
To load a MethylPipe output file that represents information at single base resolution of methylated cytosine for a sample, start by choosing the correct reference genome used for our samples by clicking on the “File” menu, then select “Load Genome”. Savant has a selection of already pre-loaded genomes or you could also choose to load one from a file. You may want to select also one or more annotation tracks as shown below:


Now locate and load the MethylPipe tracks. From file menu, choose “Load Track from File…” locate the data file and open it; it is also possible to load multiple tracks at once.
For each data file opened will be displayed methyl-cytosine indication on the track and some descriptive statistics on the widget panel on the “Radiant” tab, usually located at the bottom of the program screen.
Data exploration
Once you have visualized your data, pin the widget panel by acting on the mini button on the right side of the “Radiant” panel, and explore it.
For example you can:
  • Zoom in and zoom out the track to get advantage of using a dynamic level of resolution: as you zoom out, on track indication of methyl-cytosines is calculated on the basis of progressively wider windows. As you zoom in enough, single methyl-cytosines will be displayed.
  • Note that different methyl-cytosine context are depicted in different colours, as on track and on the Radiant Widget Panel
  • Scroll the track view or enter the chromosome coordinates on the top of the window to trigger a real time update of the main display and of the widgets.
  • Use the slider to dynamically adjust the upper limit of the area chart, especially useful to explore distribution of methylation level in uncommon sequence contexts
  • Use the bookmarks panel (usually on the right) to load a set of genomic intervals from a tab separated or gff file or to add one of your own.
  • Point to a methylated cytosine on the track to display a popup with additional info.
The descriptive charts on the Radiant’s widget panel refer to the region displayed (interval defined in the ‘Location’ text box in the upper right) and represent the distribution of methylation level for each sequence context (area chart on the left) and the track methylation level (pie chart on the right). They are calculated as in Lister, Pelizzola et al. cited below.
The pie chart represents the percentage of methylated cytosines in each sequence context (mCG, mCHG and mCHH), calculated as a ratio.
Saving your work
Finally you may want to save the configuration of tracks, bookmarks and views you are working on: choose “Save Project…” from file menu. You will be able to load it again later by the same menu or you will find it in “Recent Projects” at the start of the program.
References
Kamal Kishore, Mattia Pelizzola (2014). methylPipe: Base resolution DNA methylation data analysis. R package version 0.99.9.
Lister, R. et al. Human DNA methylomes at base resolution show widespread epigenomic differences. Nature 462, 315–322 (2009).
Lister, R. et al. Hotspots of aberrant epigenomic reprogramming in human induced pluripotent stem cells. Nature 471, 68–73 (2011).

Physical interactions (HiC)

Radiant Genome Browser is able to load and visualize physical interactions and chromosome conformation capture data. HiC data seq files must be normalized and in the format of a sparse matrix file such those outputted from HiC-Pro.

Track loading instructions
To load a matrix file that represents a normalized contact maps, start by choosing the correct reference genome used for our samples by clicking on the “File” menu, then select “Load Genome”. Savant has a selection of already pre-loaded genomes or you could also choose to load one from a file. You may want to select also one or more annotation tracks as shown below:


Now locate and load the MethylPipe tracks. From file menu, choose “Load Track from File…” locate the data file and open it; it is also possible to load multiple tracks at once.
For each data file, a track displaying local interactions relative to the focused portion of the genome will be opened. In the “Radiant Panel” tab, usually located at the bottom of the program screen, on the left side a genome wide interaction map relative to inter chromosomal interactions (also called trans-interactions) will be depicted. On the right side, you will find a set of intra-chromosomal interaction maps (cis-interactions), one per chromosome of the reference genome. You can select them acting on the bar on the bottom.
Data exploration
Once you have visualized your data, pin the widget panel by acting on the mini button on the right side of the “Radiant” panel, and explore it.
For example you can:
  • Zooming in and zooming out, you will see the portions of the focused chromosome interacting with distal chromosomes, represented as glyphs on the upper side of the track. The length of the chromosome’s glyph is proportional to the actual length of the chromosomes; the distal part of the connector has a socket that is proportional to the length of the interacting DNA region and is located to reflect its position over the chromosome length. On the focused track you will also find a horizontal proximal socket.


  • Intra chromosomal relationships (cis) are represented horizontally as arcs which loop on the track itself.
  • Collapsing Algorithm feature: the HiC contact matrix is divided in windows of various lengths but the length of both proximal and distal sockets may be longer than the length of the window. This is due to the fact that the interval of windows on the focused chromosome which is connected seamlessly to the interval of windows on distal chromosomes is considered to be a bigger one.
  • The “Radiant Panel” tab on the bottom provides two Circos plot that are an useful overview of the inter-chromosomal and intra-chromosomal interactions. Those plots are interactive: clicking on the links you will navigate to the source interaction interval originating it. On the circos plot, the sources are represented with an increased distance between the link and the chart, whereas the sinks (i.e. the targets of the interaction) are closer to it.


  • HiC data explorations often involve large intervals and usually for those resolutions Savant does not display annotations or interval tracks. If you need to display these, you can configure it in the edit menu > preferences > Track Resolution.
  • Use the bookmarks panel (usually on the right) to load a set of genomic intervals from a tab separated or gff file or to add one of your own.

Saving your work
Finally you may want to save the configuration of tracks, bookmarks and views you are working on: choose “Save Project…” from file menu. You will be able to load it again later by the same menu or you will find it in “Recent Projects” at the start of the program.