2.1. Use Case 1
Are there mutant ES cells or mice available for my gene of interest?
What mutations does the gene carry in these mutants?
How can I obtain specific mutant ES cells or mice?
There are currently several large-scale programs underway to mutate all protein-coding genes in the mouse using gene trapping and gene targeting technologies: the Knockout Mouse Project (KOMP; USA) [
1]; the European Conditional Mouse Mutagenesis Program (EUCOMM) [
2]; the North American Conditional Mouse Mutagenesis Project (NorCOMM; Canada); and the high-throughput gene trapping effort by the Texas A&M Institute for Genomic Medicine (TIGM; USA) [
3]. In order to pursue their work in a collaborative and coordinated manner, these groups have formed the International Knockout Mouse Consortium (IKMC) [
4,
5]. The IKMC web portal (
www.knockoutmouse.org) is the central public web site for IKMC data, giving researchers world-wide access to up-to-date information on IKMC knockout vectors, ES cells and mice, and links to repositories from which these products can be ordered [
6]. The IKMC database also provides data and tools to the IKMC for selecting and prioritizing genes. For example, it indicates to the IKMC for what genes mutant ES cells or mice have already been generated by other researchers. Because all this external information and links to pertinent resources are also readily available through the web site, the IKMC web portal is an excellent central starting point for the queries listed above. In the following, we describe a workflow for performing these queries:
Enter the gene symbol(s) or gene ID(s) for your gene(s) of interest into the Search box and start the search.
Search results are listed as a summary with a record (row) for each gene that matches your query ( top). Each record includes (i) the gene symbol; (ii) a high-level summary of IKMC knockout attempts that states which programs are working on the gene, provides the status of the most advanced targeting effort per program with a link to more details, and indicates the availability of targeting vectors, mutant ES cells and mice, with links to the Repositories that distribute the respective IKMC products; and (iii) other resources that report on mutant ES cells or mice for the gene of interest, with the number of, and links to, corresponding entries at the respective sites.
If you are interested in IKMC products, follow the ‘Details’ link in the ‘IKMC Knockout Attempts’ column. This brings you to a page that lists all IKMC knockout attempts for your gene of interest ( bottom). One program might try different targeting vectors to mutate a gene, resulting in several targeting projects. The production pipeline status for each project is displayed, and the availability of products is indicated via order links that lead to the respective repositories. As soon as targeting vectors have been generated for a given project, an ‘Allele Details’ link is displayed to the right of the progress bar. Clicking on this link opens a graphical display that illustrates the salient molecular features of the targeted allele. On the top left side of this newly opened display is another link ‘View this project’ that leads to more comprehensive information about the mice, ES cells and targeting vector generated by a specific project. Order links guiding users to the respective repositories are provided on this page as well.
The search path just described leads to information about gene targeting products (vectors, ES cells and mice) from KOMP, EUCOMM and NorCOMM, and gene trap ES cell lines from TIGM. Information about gene trap ES cell lines from EUCOMM and NorCOMM will be fully integrated into the IKMC web portal in the near future. Then this data will be searchable and displayed in the same way as the other IKMC products described above.
Currently, if you are interested in EUCOMM and NorCOMM gene traps, or in non-IKMC products, follow the links provided in the ‘Other Resources’ section of the initial query summary to find the entries for your gene in these databases ( top). The International Mouse Strain Resource (IMSR) [
7] is a searchable database of inbred and mutant mouse strains and stocks available worldwide and provides access to the respective holder sites, including all the members of FIMRE, the Federation of International Mouse Resources. The International Gene Trap Consortium (IGTC) database [
8] records all gene trap lines available from the consortium and provides links to corresponding holder sites. The Mouse Genome Informatics (MGI) database [
9,
10] represents all mutants for a given gene that have been reported in the published literature, by the IKMC, and by the IMSR. If pertinent mutant products are recorded in MGI, the ‘Other Resources’ section provides two links, one to targeted mutations in MGI, one to other mutations in MGI. The latter category includes gene trap ES cell lines from EUCOMM and NorCOMM. All mutant records in MGI include a reference to the original publication. ES cells or mice that are not available through the IKMC or the IMSR might be obtained by contacting the authors.
IKMC data are also accessible through MGI and the IMSR; IKMC gene trap data are also available through the IGTC. However, the IKMC web portal provides the most up-to-date and the most detailed molecular information about its products, reports on work in progress, and maintains up-to-date links to pertinent external resources. Therefore, the search strategy presented here offers effective means to find, globally, mutant ES cells and mice together with detailed information about targeting vectors and mutant alleles.
2.2. Use Case 2
Are Recombinase driver lines available under promoters of my choice?
Are there Recombinase driver lines that show activity in tissues of my choice?
A large number of targeted and gene trapped alleles produced by the IKMC, and an increasing number of targeted alleles generated by the biomedical research community, feature recombinase recognition sites that allow the generation of conditional mutants [
2,
11,
12,
13]. To take advantage of these conditional alleles, mouse lines that carry recombinases under specific promoters / enhancers are being developed [
14]. Time and tissue specificity of these recombinase lines is characterized, either by examining recombinase expression patterns, or, more accurately, by using reporter strains to determine the conditional allele pattern generated by the recombinase [
15]. Pertinent data about transgenic and knock-in recombinase lines are available through the recently established Cre-Portal (
www.creportal.org). The site lets you search for recombinase driver lines that display recombinase activity in specific tissues, or for mouse lines with recombinases under specific promoters / enhancers. As part of the larger MGI system, the portal provides the recombinase expression and reporter data, as well as phenotypic data obtained in conditional mutants. Further it indicates which mouse lines are available through the IMSR and provides links to corresponding entries in the IMSR database. The following query example illustrates how to find information about recombinase driver lines with specific promoters (see also ):
In the ‘Access Data’ section use the ‘Search for allele by promoter / driver specificity’. Select the promoter / driver of your choice from the selection list and click the ‘Go’ button.
The query result summary lists all the mouse lines / alleles that match your query. The ‘Recombinase Data’ column indicates whether recombinase activity data are available and in how many anatomical systems activity has been measured. The ‘Find Mice’ column indicates whether mouse lines are available and provides links to the IMSR. The ‘Refs’ column gives the number of publications that report on the allele / recombinase line, as one potential measure for the utility of the line, and links to the complete list of references.
To look up recombinase activity data for a specific allele, expand the corresponding information in the ‘Recombinase Data’ column by clicking on the triangle to the left of the phrase ‘Detected in x systems’. Then follow the link for a specific anatomical system to look up pertinent data, including the specific anatomical structures in which recombinase activity was reported, pertinent image data, as well as additional assay and genetic information.
To look up all recombinase and phenotype data for a given allele, on the query results summary, in the ‘Allele Symbol’ column, follow the link ‘phenotype data’.
The search and navigation path presented here is just one way to address this specific use case. For more information about the utility of the Cre Portal, you can consult the FAQs provided at the site and explore the database and its content in more detail.
2.3. Use Case 3
Are there mouse models available for a given human disease?
As pointed out above, mouse models are generated by forward and reverse genetic approaches. A key to both strategies is the phenotypic characterization of mouse mutants and the correlation of these data with human disease conditions. The MGI database is a primary resource for this type of information. MGI provides various means to search for genes and mouse mutants based on phenotypic and human disease information. Pertinent search forms and FAQs are available at
www.informatics.jax.org/phenotypes.shtml. Here we illustrate one example: finding mouse models based on human disease terms.
In the Access Data Section, follow the link ‘Human Disease (OMIM) Browser’. OMIM [
16] disease terms are listed in alphabetical order and the availability of mouse models is indicated, in parentheses, to the right of terms. Search or browse for your disease term of interest and click on the corresponding term ( top left).
(top right) shows the entry for Angelman Syndrome; AS (OMIM ID: 105830). As indicated on this page, human disease characteristics are associated with mutations in the human gene UBE3A and with mutations in the mouse orthologous gene Ube3a. However, mouse mutants for the Gabrb3 and Snrpn genes also show disease characteristics for Angelman Syndrome although OMIM data currently do not associate the disease with the orthologous human genes. Conversely, there are three additional human genes associated with Angelman Syndrome in OMIM: ANCR, CDKL5, and MECP2. The mouse ortholog for ANCR is not yet identified, and mouse mutants in Cdkl5 and Mecp2 do not show the disease characteristics. This clearly illustrates that whether a mouse mutant is a good model for a human disease conditions cannot be inferred simply by orthology. The detailed curation of mouse disease model information performed by MGI is needed to enable database queries such as the one described here.
The page illustrated by the top right of serves as an entry point to much more detailed information about the respective mouse models. By following the links in the ‘Mouse Models’ section, you can look up the detailed annotations for each allele, including information about the nature of the mutation and the phenotypes displayed on different genetic backgrounds. If MGI has recorded expression information for the respective mutant, links to these data are provided as well. To see the full detail of the phenotype information on the ‘Allele Detail’ pages, use the ‘show all annotated terms’ or ‘show all phenotypic details’ functions, or expand specific sections by clicking on the triangles.
Allele Detail pages in MGI can contain a lot of information just for one specific allele. Moreover, there can be many alleles, and thus many allele detail pages for a given gene. All the information that MGI holds for a given gene can be accessed from the respective Gene Detail page, including information about chromosomal location, mammalian orthologs, sequences, mutant alleles and phenotypes, Gene Ontology (GO) classifications, and expression data. Many links to external resources are also provided. ‘Gene Detail’ pages can be accessed by clicking on the gene symbol displayed on any of the pages described above, or via the Quick Search tool available on the top right corner of all MGI web pages.
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