Instructions

CoNTExT takes as input the full transcriptomic data of your neural progenitor and differentiated cell cultures, measured via microarray chip, and compares it to in vivo transcriptomic atlases of brain development.  Your data is uploaded to the server, processed, and the results files are emailed to you.

Step 1: Acquire transcriptomic data of neural progenitor and differentiated cultures.  Importantly, 3 biological replicates each of neural progenitor and differentiated cultures are needed for the pipeline (minimum 6 total samples). These samples should be profiled on a microarray chip, and either run simultaneously on one chip, or randomized to a chip and batch corrected via ComBat or other method.

Step 2: Format the data.  CoNTExT requires 3 files to run, which must be formatted in the appropriate way.  All three files must be comma separated value files (*.csv).  You can make these files in MS Excel but we caution you that this program often changes values inadvertently.  We advise you to make these files in the R package.

Annotation file: This file contains the mapping of probes to genes. The rownames are the probes and should match exactly to the rownames of the expression file. The SYMBOL column represents the HGNC gene symbol.  The ENTREZ_GENE_ID is the Entrez gene ID from NCBI.  All of this information will be found on the website of the chip manufacturer, but must be organized in the following way.  Here is an example file:

Metadata file: This file contains the mapping of samples to their culture status and line. The rownames are the sample identifiers and should match exactly to the columns of the expression file. The Status column should be labeled with either “Progenitor” or “Differentiated” indicating the differentiation state of the culture. The Replicate column should be used to describe which samples came from the same line which can be defined as any of the following: same subject, same clone, or same line.  Replicate is used to properly account for samples which are more similar to each other than other samples as a random effect in a mixed effect smodel.  Here is an example file:

Expression file: This file contains the expression of each probe for each sample. Prior to input, expression files should be log2 transformation, quantile normalized, and batch corrected (if necessary). The columns of the expression file should correspond exactly to the rows of the metadata file. The rows of the expression file should correspond exactly to the rows of the annotation file. Here is an example file:

Step 3: Register. In order to prevent overuse or misuse of the servers, we ask that each contributor register with their name, email address, and institution. Registrations are manually approved, so we’ll try to get back to you within a day to approve your account.  Registration is only necessary one time. If you are a netbot, you must demonstrate devoted interest to neural stem cells and pass a Turing test.

Step 4: Upload. After logging in, the user can upload the three files to the server.  After clicking submit, the job will be executed on the server.  Running time is approximately 3-4 hours but may be longer if others are also running.  The user will receive an email with the results attached when the job is completed. If you do not receive an email within 4 hours, please first check the FAQs page and if your problem is not solved, send an email to the developers.