This guide will assist you in using CisMols Analyzer, a program that identifies compositionally predicted cis-clusters, which we call CisMols (Cis-regulatory Modules), that occur in groups of co-regulated genes within each of their ortholog-pair evolutionarily conserved cis-regulatory regions. CisMols Analyzer is based on the hypothesis that the presence of a cluster of ortholog-conserved known cis-acting elements in all or many of the co-expressed or functionally related genes predicts that the genes were co-regulated by these elements. Designed to search for regulatory clusters not just in the upstream region of co-expressed genes but also in the non-coding intronic and 5' and 3' flanking genomic regions, CisMols Analyzer could lead to the discovery of probes for genome-wide identification of regulatory regions.
To assist you in using CisMols Analyzer, we have divided this help file into eight sections that reflect the main steps to using the system:
If you need additional support at any time, contact the CisMols administrator.
To create projects and gene lists, you need a login account. You can request one by emailing the CisMols administrator. Once you have an account, you can log in by completing the following steps:
1. If the login screen does not already appear, go to the left-hand menu, and click Login:
2. Type your User Name and Password.
3. Click Login:
You should now see a screen for selecting a data source. If you don't see this screen or receive an error message, try logging in again, remembering that user names and passwords are case-sensitive. If you still are unable to log in, contact the CisMols administrator.
Projects are used to group gene lists for analysis. They can contain public gene lists, private gene lists or both (for more information about gene lists, see Managing gene lists). When projects contain public gene lists, anyone can access them and search their public gene lists without obtaining a login account. If you are only interested in searching public gene lists in existing projects, you may disregard the instructions in this section and the next section, and go to the instructions for searching for cis-clusters.
If you are interested in creating your own projects, you need to obtain a login account. Once your account is set up, you can create multiple projects and assign relevant gene lists to each. Available gene lists include those you have created as well as those that other users have created and made publicly available. For more information about gene list creation and management, see Managing gene lists.
This section gives you step-by-step instructions for performing project management tasks. To complete any of these tasks, you need to access the Create or Edit Project screen by completing the following steps:
1. Log in if you have not done so already.
2. Select a data source:
3. Click CisMols Analyzer:
4. Click Projects:
The Create or Edit Project screen opens. From this screen, you can complete the following tasks:
1. In the upper-left section of the Create or Edit Project screen, click Create New Project:
2. Type the Project Name and a short Description.
3. Click Create:
The Create or Edit Project screen again appears. Your project should now be listed under My Projects. To assign one or more gene lists to the project, follow the steps in the next section.
To assign one or more gene lists to a project, complete the following steps.
1. From the Create or Edit Project screen, select the project by going to My Projects and clicking the project name:
2. Do one of the following:
3. Enter as many criteria as you wish, and click Search.
The View Gene Lists screen appears, displaying all private gene lists matching your search criteria.
4. Go to the table of available gene lists, and select the list(s) that you want to add to your project.
5. Click Add:
The Edit Projects screen again appears, indicating that the selected gene list(s) are now included in your project.
6. To add more gene lists to the project, repeat steps 2-5.
To remove one or more gene lists from a project, complete the following steps.
1. From the Create or Edit Project screen, select the project by going to My Projects and clicking the project name:
2. Under My Genes, select the the check box next to the gene(s) you want to remove.
3. Click Remove:
4. To confirm that you want to remove the selected gene(s), click OK.
If a project contains one or more public gene lists, any user can access the project and search its public gene list(s) without obtaining a login account. However, each project can have multiple owners and users with special privileges. Owners and users are able to view private gene lists in the project. In addition, owners are able to modify and delete projects. By default, you become the sole owner of each project you create. If you wish, you can add and remove owners and users by completing the following steps:
1. From the Create or Edit Project screen, select the project by going to My Projects and clicking the project name:
2. In the upper-right section of the screen, click Manage Users:
A screen appears listing associated and non-associated users.
3. Do one of the following:
4. To exit the screen, go to the upper-right section, and click Done:
To change the name of a project, complete the following steps.
1. From the Create or Edit Project screen, select the project by going to My Projects and clicking the project name:
2. For Name, type a different name for the project.
3. Click Rename:
To remove a project completely from the system, complete the following steps.
1. From the Create or Edit Project screen, select the project by going to My Projects and clicking the project name:
2. Click Delete:
3. To confirm that you want to delete the selected project, click OK.
CisMols Analyzer works with groups of co-expressed or related genes called gene lists, which further can be grouped into projects (see Managing projects). By default, the system contains a number of public gene lists that can be searched and analyzed without a login account. If you are only interested in searching public gene lists, you may disregard the information in this section and go to the instructions for searching for cis-clusters.
If you are interested in creating your own gene lists, you need to obtain a login account. Once your account is set up, you can follow the instructions in this section to create and manage gene lists.
To complete all tasks in this section, you need to open the Welcome screen by completing the following steps:
1. Log in if you have not done so already.
2. Select a data source:
3. Click CisMols Analyzer:
The Welcome screen appears:
From this screen, you can complete the following tasks:
1. At the Welcome screen, click Create Gene List.
2. Enter any of the following criteria:
3. Click Search and Add Genes:
4. Go to the table of search results, and under Select, select the check box next to each gene that you want to add to your list. Or if you wish to add all the genes in the table to your list, go to the top of the table and select the Select All Genes check box:
5. Click Add to Gene Cart:
6. Repeat steps 2-5 as many times as you wish to add other genes to the list.
7. When you are ready to finalize the contents of your list, click View Genes and Proceed:
8. Verify the contents of your gene lists, and make any last-minute adjustments by using the Remove Selected Genes and Add More Genes buttons.
9. Once you have finalized the contents of your gene list, go to the top of the screen, and complete the following fields:
10. Click Submit Gene List:
You will receive several emails as your gene list is being created and uploaded. Once your list is ready, you can search the list for clusters and create a CisMolGram based on your search results.
When you create a gene list, you indicate whether it should be private or public. At any time, you can modify the access level of a gene list you have created by completing the following steps:
1. At the Welcome screen, click Search for Gene List.
2. Enter as many search criteria as you wish, and click Search:
3. Select one of your gene lists by clicking the Gene List Name:
4. Go to the top-right section of the screen, and select Private or Public.
5. Click Change Access Level:
The access level of your gene group is now modified.
When users search gene lists for cis-clusters, they can save their searches (see Saving your search). Once a search is saved, it can be retrieved by its owner and other users at any time (see Retrieving a saved search). When you log in and search for a gene list, all saved searches associated with this gene list appear at the bottom of the screen. If you are the owner of a saved search -- that is, if you are the user who created it originally -- you can delete it by completing the following steps.
1. At the Welcome screen, click Search for Gene List.
2. Enter as many search criteria as you wish, and click Search:
3. Select one of your gene lists by clicking the Gene List Name:
4. Under Saved Searches, select a search that you created.
5. In the same row, under Delete, click the trash can icon:
The saved search is now removed from the system.
You can rename a gene list you have created by completing the following steps:
1. At the Welcome screen, click Search for Gene List.
2. Enter as many search criteria as you wish, and click Search:
3. Select one of your gene lists by clicking the Gene List Name:
4. For Name, type a new name for the gene list.
5. Click Rename:
The name of your gene list is now changed.
You can delete a gene list you have created by completing the following steps:
1. At the Welcome screen, click Search for Gene List.
2. Enter as many search criteria as you wish, and click Search:
3. Go to the gene list you want to delete, and under Delete, click the trash can icon:
The gene list is now removed from the system, but all genes in the list remain in the database.
Whether you are searching an existing gene group or a group you created, the CisMols Analyzer enables you to customize the parameters of each search you perform. Specifically, you can use Boolean logic to restrict your search to known clusters and/or a combination of individual binding sites. You also can specify the minimum number of binding sites that must appear in each cluster, or the minimum number of genes in which each cluster must appear.
Continue reading this section for step-by-step instructions on selecting a gene list and performing one of the following tasks:
To select a gene list for analysis, complete the following steps:
1. Log in if you want to search private gene lists in addition to public gene lists.
2. From the homepage, select a data source:
3. Click CisMols Analyzer:
4. Click Search for Gene List:
5. Enter as many criteria as you wish, and click Search.
The View Gene Lists screen appears.
6. Go to the table of available gene lists, and select the list that you want to search.
You now are ready to perform a new search or retrieve a saved search.
To search a gene list from scratch, complete the following steps:
1. Select a gene list if you have not done so already.
2. Go to the bottom of the Edit Gene List Properties screen, and click View CisMols Clusters:
3. Under Min Genes and Sites in a Cluster, enter the following information:
4. For Order top 100 clusters by, indicate whether you want the system to rank results by the number of genes in which each cluster appears or by the number of binding sites in each cluster.
5. For View Region, type a sequence range if you wish to restrict your search of ALL genes to a range other than the default range of 10 kb upstream and downstream.
6. Under Select individual sites, indicate whether or not search results must contain specific binding sites.
7. Under Select from known Modules, indicate that search results must contain a known cis-module by selecting the check box next to the module. To select all modules, select the Select from known Modules check box.
8. Between the individual sites and known modules tables, select OR if search results should satisfy the conditions in EITHER table, or select AND if search results must satisfy the conditions in BOTH tables.
9. When you are finished defining your search criteria, click CisMols Search:
A screen appears for creating a CisMolGram.
Once users finish searching a gene list, they can save their search so that they and other users can retrieve the search at any time, perform it again or use it as the basis of a new search. To retrieve a saved search, complete the following steps:
1. Select a gene list if you have not done so already.
2. At the bottom of the Edit Gene List Properties screen, under Saved Searches, select the search you wish to perform.
3. Click Launch Search:
The CisMols search screen appears with all of the user's original selections and data. Click CisMols Search to proceed to the next screen, which shows how the user originally configured the CisMolGram, and from this screen, click View Peak Regions of Clusters to view the originally generated CisMolGram. Or modify the user's original selections and use them as the basis of your own search. Once you're finished, you can save this search under a new name, but you cannot save it under the same name as the original search.
Once your search results are returned, you can view the data not only in tabular form but also graphically. We call these images CisMolGrams. They depict the location of clusters within gene regions. In this section, you will learn how to create a CisMolGram -- specifically, how to complete the following tasks:
By default, all genes in the group are selected for inclusion in the CisMolGram. You can remove one or more genes by going to the top-left corner of the table, selecting On All But Selected and selecting the check boxes next to all genes that should NOT be included in the CisMolGram:
The total number of clusters matching your search criteria is indicated in the top-left corner of the screen. You can get a complete list in Excel format by clicking Download All:
In the table, only the top 100 clusters are displayed according to the number of genes in which they appear. By going to the table and clicking No. of Sites, you can sort clusters according to the number of sites they contain. You also can display a range other than the top 100 by going to the From and To text boxes in the upper-right corner of the screen, typing a range that's within the total number of clusters, and clicking Get Cluster Numbers:
Once you have finalized the contents of the clusters table, you must select at least one cluster. There are three ways to select clusters:
Go to the drop-down menu labeled Select No. of Clusters, and select the number of clusters to include in the CisMolGram. For example, to select the first 5 clusters in the table, select 5:
Next to the Select No. of Clusters drop-down menu, go to the From and To text boxes, type the range of clusters to include in the CisMolGram, and click Select Clusters. For example, to select the clusters in rows 10 through 25 in the table, type 10 in the first box and 25 in the second box:
To select individual clusters, go to the table and select the check box next to each cluster that you want to include in the CisMolGram:
Once you have specified which genes and clusters to include in the CisMolGram, you are ready to generate the image by clicking View Clusters:
A screen appears that graphically displays peaks, or regions containing one or more clusters, according to the conditions you specified. See the next section to learn how to manipulate and analyze your CisMolGram.
Once your CisMolGram is generated, you will see an image similar to the following, where each symbol represents a cluster:
This section outlines several options for modifying your CisMolGram and aiding your analysis including
A peak is a region of a gene containing one or more cis-cluster. By default, the system is set to display up to five clusters at any peak. If a peak includes more than five clusters, the following symbol appears:
Simply click the symbol to view all clusters in this peak.
To adjust the maximum number of clusters per peak, go to the Display options directly below the CisMolGram. For Max Clusters to display at a peak, type a whole number. Then click Refresh.
By default, the CisMolGram includes the region 10kb up- and downstream of the gene -- unless you specified a different region when you entered your search criteria (see Performing a new search).
At this point, you cannot modify the CisMolGram to include more clusters than specified by the gene group parameters or your original search criteria. However, you can zoom in on the existing CisMolGram by going to the Display options directly below the CisMolGram. For Viewable Region, type a region in base pairs. Then click Refresh. For all genes, peaks are now displayed for the region you specified.
To view peaks in orthologs of the human genes (i.e., mouse genes), go to the Display options directly below the graph. Click Show Ortholog Sequence View.
For each peak, you can view a TraFaC image, or depiction of the shared transcription factor binding sites that constitute the peak. To view a TraFaC image, click any cluster in the peak:
In the pop-up window, click View Trafac Image. Here is an example of the screen that appears, along with numbered annotations explaining its parts:
For each peak, you can view a regulogram, or a cis-element hit density image, which depicts a moving-window average of the number of shared cis-elements occurring in phylogenetically conserved regions. To view a regulogram, click any cluster in the peak:
In the pop-up window, click View regulogram. Here is an example of the screen that appears, along with numbered annotations explaining its parts:
Below the CisMolGram is a legend indicating (a) the number of genes containing each cluster and (b) the sites within each cluster. You can sort the items in this legend by clicking the buttons above it (grouped into three categories -- Gene Sorting, Site Sorting and Cluster Sorting). You also can sort items by clicking the row and column labels in the table, as outlined below:
At any time when modifying and analyzing a CisMolGram, you can export the image and legend to several common file formats. Simply go to the bottom-left corner of the screen, and for Export to, select a file format:
Before you exit the CisMols Analyzer, you can save your search, including all your selections on every screen, by completing the following steps.
1. Go to the bottom of the screen, and click Save Search:
2. For Search Name, type a one-word name for your search.
3. For Description, type a short description of your search.
4. Click Save:
A message should appear indicating your search has been saved. If it does not, go back to the CisMolGram and repeat these steps to try saving your search again.
For instructions on opening saved searches, see Retrieving a saved search.