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Searching the Developmental Transcriptome

The Developmental Transcriptome search functionality can be found by clicking the corresponding button in the menu bar.

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 brain structures at desired developmental stages, 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 from any search type will take you to the appropriate help section.

Gene Search

With the Gene Search radio button selected (default) you can either select a category from the tag cloud or type the name of your gene of interest into the text box and then click Search. With the October 2013 release, sequence data from non-coding regions was also added to the data-base. You can find that data by entering the ncRNA Ensembl ID.

Results will be returned in a heat map format with the genes on the y-axis (vertical) and the donors/structures on the x-axis (horizontal). The heat map represents the normalized gene level RNA-Seq expression data in RPKM (Reads Per Kilobase of exon model per Million mapped reads). By default, the columns in the heat map are sorted first by donor, then by structure. To toggle the initial sort parameter, click the toggle button in the top right hand corner of the heat map.

Clicking on a data point in the heat map will bring up more information in the panel above the heat map. This information includes the structure (click to be taken to the reference atlas) and donor information, gene information including gene name and symbol (Click to be taken to the exon level data), Ensembl ID, gene expression (in log2 RPKM) and a link to related data in other Allen Brain Atlas resources.

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

You can select a set of probes for later use 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.

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" mode. Select the "Differential Search" radio button. To find genes enriched in one "target set" of brain regions and/or developmental stages compared to a "contrast set" of brain regions and/or stages, choose the target set using the "Target Structure(s)/Stage(s)" drop down menu, and the contrast set using the "Contrast Structure(s)/Stages(s)" drop down menu. The default setting selects all structures and stages, but clicking on the "All Structures" or "All Stages" drop down menu will allow you to select individual structure(s)/stages(s). Then click "Search".

The search will return a list of genes enriched in the target domain over the contrast domain. To see only the domains selected from your search criteria, select the "Filter Heatmap" button below the heatmap.

Once selected a window will open allowing you to select the structures and the developmental stages you are interested in visualizing.

Once you have identified a single gene and want to dig deeper into the exon level expression data, click on the blue colored gene name or symbol above the heat map. This link takes you to the transcriptome visualization page.

Correlative Search

In using gene expression databases, a "search by example" feature is also highly desirable as genes with similar expression patterns may be related in function. Using the "Find Correlates" search utility will accomplish this function. This search by example facility is also available in the Allen Human Brain Atlas as well as in the Allen Mouse Brain Atlas and in the Allen Developing Mouse Brain Atlas.

Once you have selected a gene of interest (by clicking on the heat map that results from a Gene or Differential search), you can find genes with similar expression patterns over the entire dataset or over a subset of structures and developmental stages. These subsets can be chosen using the drop down menus. All structures are checked by default, but clicking "All Structures" a second time will deselect all structures, similarly for "All Stages". Once you have selected the brain region(s) and development stage(s) of interest, click "Find Correlates".

The search will return a list of genes enriched in the target domain over the contrast domain. To see only the domains selected from your search criteria, select the "Filter Heatmap" button below the heatmap.

Once selected a window will open allowing you to select the structures and the developmental stages you are interested in visualizing.

Once you have identified a single gene and want to dig deeper into the exon level expression data, click on the blue colored gene name or symbol above the heat map. This link takes you to the transcriptome visualization page.

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

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).

Data Visualization

The transcriptome visualization page is divided into 3 sections.

The first section contains gene Information data including the Ensembl symbol and ID, the chromosome number, the NCBI Entrez symbol, ID and name, gene aliases, links to the USC and the Ensembl genome browsers which take you directly to gene locus data, and links to related data from other Allen Brain Atlas resources.

The second section provides a composite gene model where exons are represented by boxes and introns are represented by the green lines between exons. When you hover your mouse over an exon in either the RNA-Seq or exon array data, the exon is highlighted in this model.

The third section contains two tabs:
A tab labeled RNA-Seq with a heat map of the exon level expression data in RPKM (Reads Per Kilobase of exon model per Million mapped reads).
A second tab labeled Exon Array with a heat map of the exon probeset level expression data in log2 transformed normalized expression intensity values.

This data can be downloaded by clicking on the "Download this data" link at the bottom left of the heatmap.

Transcriptome Heat Maps

The heat map is a visualization of the exon expression values for the returned gene of interest. The heat map data is presented as a matrix with brain structure (by developmental stage) on the x-axis and gene exons on the y-axis. Brain structures are organized in ontological order. Clicking on the toggle button in the right hand corner will toggle the initial sorting parameter from structure to developmental stage.

Each row of the heat map in the RNA-Seq data represents an exon. Exons in the RNA-Seq data are labeled by the start position on the chromosome and the length of the exon. Each row of the heat map in the Exon Array data represents an exon probeset. Approximately 4 probes are selected for each putative exon region and called a probeset. Each row in the Exon Array data is labeled with the chromosome start position and the length of the probeset. Hovering the mouse over the exon in either heat map will highlight the corresponding exon in the composite gene model over the array data.

Each column of the heat map represents a tissue sample. The colors of the heat map are expression values, transformed to a log 2 scale. The color scale ranges from dark blue representing low expression and passes through cyan, yellow, orange and finally to dark red representing high expression. Hovering the mouse over a cell of the heat map will bring up donor and structure information in the area above the heat map.

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