SEARCHING THE RNA Sequencing (RNA-Seq) DATA

Three search types are available: (1) Gene Search, to obtain gene expression data for specific genes of interest, (2) Differential Search, to compare expression between two sets of glioblastoma anatomic structures, and (3) Correlative Search, to find genes that have an expression pattern similar to a "seed gene" selected from the results of a Gene or a Differential search.

Clicking on the "?" button will take you to the appropriate help section.

Gene Search

With the Gene Search radio button selected (default), you can either select a gene only by typing the name of your gene of interest into the text box, or you can further restrict your selection criteria by selecting tumor features and/or tumors. Once you have made your selections, click the Search button. The search will not work unless a gene has been selected. You can further limit your search by selecting the box next to "Show exact matches only".

Results will be returned in a heatmap format with the genes on the y-axis (vertical) and the tumors/tumor features on the x-axis (horizontal). The heatmap represents the normalized gene level RNA-Seq expression data in FPKM (Fragments Per Kilobase Of Exon Per Million Fragments Mapped). By default, the columns in the heatmap are sorted first by tumor, then by tumor feature. To change sort order, click the toggle button in the top right hand corner of the heatmap.

Clicking on a data point in the heatmap will populate the metadata box above the heatmap with information specific to that gene and tumor. This meta-information includes information regarding the tumor, the gene and a search box to search for genes that show similar expression patterns as your gene of interest. There are two links available from the tumor box, clicking on the "Tumor Sub-Block" link will open up a new tab with the Specimen Detail Page and clicking the "SNI" link will take you to the Swedish Neuroscience Institute page (registration is required to access the patient metadata). Gene information includes metadata on the gene selected including links to ISH data for that gene (when available) and links to other Allen Brain Atlas resources that include that gene.

If there is ISH data available for a gene, there will be a link labeled "Yes" that will take you to the ISH Data for that gene.

You can adjust the window and the level of the color scale on the heatmap by adjusting the threshold bars or re-centering the color map scale bar. You can also zoom in or out to see the heatmap at different magnification levels.

You can select a set of genes for later viewing by selecting the check-boxes beside the list of gene symbols. Your choices are stored in a browser cookie in your computer and will remain in effect until you click the "Clear Selections" button or clear your web browser's cookie cache. Click the "View Selections" button to see your selections.

To restrict the columns that are displayed, select the "Filter Heatmap" button below the heatmap. Filtering your heatmap is a two-step process: first, select the "..." box to restrict your features, making sure to save your selections and then toggle the filter heatmap feature by clicking the "Filter Heatmap" button.

Correlative Search

In using gene expression databases, a "search by example" feature is highly desirable as genes with similar expression patterns may be related in function. Using the "Find Correlates" search utility will accomplish this function.

Once you have selected a gene of interest (by clicking on the heatmap that results from a Gene Search or Differential Search), you can find genes with similar expression patterns. All Features and All Tumors are included in your search by default, but you can select one or more features or tumors from the drop down menus. Once you have selected the tumor feature(s) and tumor(s) of interest, click "Find Correlates".

The search results highlight the genes whose expression profiles across the samples of interest resemble the expression profile of the example gene by ranking the genes according to the Pearson’s correlation coefficient (r) between their profile and that of the example gene.

If there is ISH data available for a gene, there will be a link labeled "Yes" that will take you to the ISH Data for that gene.

You can adjust the window and the level of the color scale on the heatmap by adjusting the threshold bars or re-centering the color map scale bar. You can also zoom in or out to see the heatmap at different magnification levels.

You can select a set of genes for later viewing by selecting the check-boxes beside the list of gene symbols. Your choices are stored in a browser cookie in your computer and will remain in effect until you click the "Clear Selections" button or clear your web browser's cookie cache. Click the "View Selections" button to see your selections.

To restrict the columns that are displayed, select the "Filter Heatmap" button below the heatmap. Filtering your heatmap is a two-step process: first, select the "..." box to restrict your features, making sure to save your selections and then toggle the filter heatmap feature by clicking the "Filter Heatmap" button.

You can see "anti-correlated" genes by toggling the sort order on column "r" or scrolling to the bottom of the heatmap.

Differential Search

Another common usage of gene expression databases is to find genes that show enrichment of expression in one set of samples compared to another set of samples. This type of query is supported by the "Differential Search" feature. Under the RNA-Seq tab, click on the examples in the “Browse by Differential Expression” box, or select the “Differential Search” radio button to start your own search. To find genes enriched in one "target set" of tumor features and/or tumors compared to a "contrast set" of tumor features and/or tumors, choose the target set using the "Target Tumor Features/Tumors" drop down menu, and the contrast set using the "Contrast Tumor Features/Tumors" drop down menu. The default setting selects all tumor features and tumors, but clicking in the "All Features" or "All Tumors" text boxes will allow you to select individual tumor feature(s)/tumor(s). Then, click "Search".

The search will return a list of genes enriched in the target domain(s) over the contrast domain(s) ranked by the Fold change in expression values. The results can be sorted also by the statistical significance of the evidence (p-Value) by clicking on the desired sort parameter.

If there is ISH data available for a gene, a link labeled "Yes" takes you to the ISH Data for that gene.

You can adjust the window and the level of the color scale on the heatmap by adjusting the threshold bars or re-centering the color map scale bar. You can also zoom in or out to see the heatmap at different magnification levels.

You can select a set of genes for later viewing by selecting the check-boxes beside the list of gene symbols. Your choices are stored in a browser cookie in your computer and will remain in effect until you click the "Clear Selections" button or clear your web browser's cookie cache. Click the "View Selections" button to see your selections.

To restrict the columns that are displayed, select the "Filter Heatmap" button below the heatmap. Filtering your heatmap is a two-step process: first, select the "..." box to restrict your features, making sure to save your selections and then toggle the filter heatmap feature by clicking the "Filter Heatmap" button.

Download Search Results

To download your search results, click the "Download this data" link at the bottom left of the heatmap. You will have the option of downloading up to 2000 genes at a time starting from any position in the heatmap. Your start position can be determined from the counter in the top left corner of the heatmap (see screenshot).

Your data will be downloaded as three separate files: 1) a Columns.csv file that lists the column headers with associated metadata for each sample, 2) a Rows.csv file that lists the row headers with associated metadata for each gene, and 3) an Expression.csv file that provides a matrix of the expression values for each data point.

The column headers, row headers and expression data will be consistent with the settings from which you downloaded the heatmap (i.e. color map).

To download the complete data set used in the heatmap, go to the Download tab and click the appropriate link.