Briefings in Functional Genomics Advance Access originally published online on August 20, 2007
Briefings in Functional Genomics 2007 6(3):186-192; doi:10.1093/bfgp/elm018
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Special Issue Papers |
EMMA—The European mouse mutant archive
Corresponding author. Martin Hrabè de Angelis, GSF National Research Center for Environment and Health, Institute of Experimental Genetics, Ingolstaedter Landstr. 1, D - 85764 Neuherberg, Germany. Tel: 089 3187 3302; Fax: 089 3187 3500; E-mail: hrabe{at}gsf.de
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ABSTRACT |
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 TOP ABSTRACT BACKGROUNDTHE EMMA NETWORKEMMA SERVICES AND PROCEDURESADDITIONAL EMMA SERVICESTHE EMMA ARCHIVEEMMA GOES GLOBAL--FIMREVALUE FOR RESEARCH COMMUNITYREFERENCES |
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The European Mouse Mutant Archive (EMMA) offers the worldwide scientific community a free archiving service for its mutant mouse lines and access to a wide range of disease models and other research tools. EMMA is currently comprised of seven partners who operate as the primary mouse repository in Europe. EMMA' s primary objectives are to establish and manage a unified repository for maintaining mouse mutations and to make them available to the scientific community. In addition to these core services, the consortium can generate germ-free (axenic) mice for its customers and also hosts courses in cryopreservation. EMMA is a founder member of the Federation of International Mouse Resources (FIMRe). The EMMA network is funded by the participating institutes, national research programmes and the European Commission Research Infrastructures Programme.
Keywords: cryopreservation, IVF, functional genomics, mouse mutants, disease models, mouse repository
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BACKGROUND |
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TOPABSTRACTBACKGROUNDTHE EMMA NETWORKEMMA SERVICES AND PROCEDURESADDITIONAL EMMA SERVICESTHE EMMA ARCHIVEEMMA GOES GLOBAL--FIMREVALUE FOR RESEARCH COMMUNITYREFERENCES |
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The European Mouse Mutant Archive (EMMA) is the main centre for the archiving and distribution of mutant mouse lines essential for biomedical research in Europe. The objective of EMMA is to provide a centralized access to mouse mutant lines for the scientific community. The laboratory mouse is the most important mammalian model for studying genetic and multi-factorial diseases in man. Based on its excellent genetics and long history in biomedical research numerous mouse mutants were identified. In the last two decades, technological improvements provided the ability to produce mouse mutants for virtually any gene that has been cloned by gene-specific approaches (knock-outs, knock-ins and conditional mutagenesis). This is complemented by random approaches such as large scale, genome-wide ENU mutagenesis and gene trapping. It is expected that more than 30 000 mutations in ES cells will be engineered and thousands of mouse disease models will become available over the next years by the collaborative effort of the International Mouse Knockout Consortium [1, 2]. Since it is impossible to retain all of these mutant mouse strains it is essential that all mutants that have been created are held in central repositories from which mutant mice can readily be made available to interested investigators [3, 4]. To meet these needs in Europe the European Mouse Mutant Archive was established and implemented. Six partners and the EBI formed the EMMA consortium. In addition to its financial support from the EU, EMMA is supported by the partner institutes and by national research programs.
Objectives of Emma
EMMA's primary objectives are to establish and manage a unified repository for maintaining mouse mutations and to make them available to the scientific community. In addition to these core services, the consortium can generate germ-free (axenic) mice for its customers and hosts courses in cryopreservation to promote the use and dissemination of frozen embryos and spermatozoa. Dissemination of knowledge is further facilitated by a dedicated EMMA resource database.
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THE EMMA NETWORK |
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TOPABSTRACTBACKGROUNDTHE EMMA NETWORKEMMA SERVICES AND PROCEDURESADDITIONAL EMMA SERVICESTHE EMMA ARCHIVEEMMA GOES GLOBAL--FIMREVALUE FOR RESEARCH COMMUNITYREFERENCES |
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The EMMA network is a partnership of several laboratories and other institutions throughout Europe [5] (Table 1). The current membership includes the CNR Istituto di Biologia Cellulare in Monterotondo, Italy; the CNRS Centre de Distribution de Typage et d’Archivage Animale in Orleans, France; the MRC Mammalian Genetics Unit in Harwell, UK; the Karolinska Institute in Stockholm, Sweden; the Instituto Gulbenkian de Ciência in Oeiras, Portugal; the GSF Institute of Experimental Genetics in Munich, Germany and the EMBL European Bioinformatics Institute in Hinxton, UK, which is responsible for the development of the EMMA resource database. Recently, the Sanger Institute in Hinxton and the ICS in Straßbourg joined EMMA. Every EMMA partner is a major player in mouse genetics in their countries. In addition, EMMA is a founding member of FIMRe (Federation of International Mouse Resources, www.fimre.org) a collaborating group of mouse repository and resource centres worldwide. The EMMA network is directed by Professor Martin Hrabé de Angelis who also heads the GSF/IEG in Munich.
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The management structure of the EMMA consortium was set up to ensure an effective operation of such a large and international enterprise and consists of several components (Figure 1). EMMA is headed by the director who leads the Board of Participating Directors (BPD), the decision-making level within the consortium. The Technical Working Group develops and agrees on EMMA SOP's for quality control, archiving and distribution. To guarantee an effective communication within the virtual network, an internal webpage was set up which constitutes a fundamental tool for the management of scientific, technical and logistical activities carried out by each EMMA partner. Furthermore, the EMMA resource database provides up to date information about the archiving status of mice and describes the genetic and phenotypic properties of all the mutant strains that EMMA stocks. The available strain list is displayed on the public EMMA website (www.emmanet.org). This website is the link to the scientific community and facilitates the submission of mice to EMMA and requests of mice from EMMA. In addition, information on the SOP's used by EMMA is provided on the public website.
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EMMA SERVICES AND PROCEDURES |
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TOPABSTRACTBACKGROUNDTHE EMMA NETWORKEMMA SERVICES AND PROCEDURESADDITIONAL EMMA SERVICESTHE EMMA ARCHIVEEMMA GOES GLOBAL--FIMREVALUE FOR RESEARCH COMMUNITYREFERENCES |
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Core EMMA services—archiving and distribution
To control the quality of the EMMA processes Standard Operation Protocols (SOPs) regarding freezing procedures, health status and animal handling were developed (www.emmanet.org). The first control step after a mouse is taken into an EMMA facility is the geno- and/or phenotype verification of the respective mutation (Figure 2). This is followed by a test freezing of either sperm or embryos. The ability to reconstitute the stock from frozen material will be ascertained at the time the strain is being frozen. A mutant line is considered to be reconstituted as soon as live offspring bearing the appropriate geno- and/or phenotype has been raised to maturity. Health monitoring is performed on all in- and out-going mouse lines. All mice supplied to the customer have a specific-pathogen-free (SPF) status according to FELASA guidelines.
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Cryopreservation of germplasm is well recognized for providing an efficient management of genetic resources and freezing of in vivo produced embryos is still regarded as the method of choice. However, up to 500 embryos are normally required for a reliable re-establishment of a breeding colony from frozen embryos [6]. Therefore, preservation of a mutant strain may demand a time and space consuming expansion of the breeding colony before abolition. Since the first reports of successful cryopreservation of mouse spermatozoa were published [7–9], a large number of inbred mouse and hybrid strains as well as mutant mouse lines have been used for the cryopreservation of spermatozoa [10–14]. Compared to embryo freezing the cryopreservation of spermatozoa offers a number of advantages. Sperm from a single male could potentially give rise to as many as 20 times more offspring than embryos from a single female and the collection and manipulation of spermatozoa is faster [15]. Additionally, this method will facilitate the preservation of strains in which female reproductive problems are characteristics [16]. However, the difference between freezing haplotypes (spermatozoa) and full genomes (embryos) has to be kept in mind. Sperm freezing is problematic in situations where the genotype of the total genome is of interest e.g. in congenic or inbred strains. Recovering frozen embryos results in animals with the genetic background of their parents. To reach the latter after recovery of frozen spermatozoa, oocyte donors from the same strain have to be used. This might be important because several mutations lead to altered phenotypes in different genetic backgrounds. Sperm freezing of strains with maternally inherited alterations will be inappropriate if suitable oocyte donors are not available.
The so-called Cryo IVF (in vitro fertilization) method strikes a balance between advantages and disadvantages of both freezing methods. This approach provides the same security as compared to an in vivo production of embryos. Furthermore, a high number of embryos can be produced via IVF in a short time without expanding the colony, whereas the depositor only has to provide a small number of males of the respective mutant mouse strain (Figure 3).
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EMMA aims to constantly improve the established SOPs, in order to accelerate the cryopreservation procedures to provide customers a state of the art service and to ensure a high hygienic status of the mice [17, 18]. In addition, several of the partners in the EMMA consortium develop assisting reproductive techniques such as ICSI (intracytoplasmic sperm injection) and laser micro-dissection of oocytes as a means to rescue spermatozoa with low fertilization performance.
The reliability of the EMMA archive is therefore based on two major requirements. The chosen archiving methods have been proven extensively in the archiving units of the EMMA consortium [11, 18–20]. Nevertheless, the IVF rate, recovery rate after thawing and percentage of live offspring varies among strains [21]. Therefore, it is crucial to select accurately the most suitable archiving procedure or combination of different archiving procedures for each individual mouse mutant strain deposited in EMMA.
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ADDITIONAL EMMA SERVICES |
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TOPABSTRACTBACKGROUNDTHE EMMA NETWORKEMMA SERVICES AND PROCEDURESADDITIONAL EMMA SERVICESTHE EMMA ARCHIVEEMMA GOES GLOBAL--FIMREVALUE FOR RESEARCH COMMUNITYREFERENCES |
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Mouse production, axenic service, nomenclature service and cryocourses
ES cells and mouse production
Live mice can also be supplied to EMMA customers via micro-injection of ES cell lines into host blastocysts. Several EMMA partners (Table 1) are responsible for the production of up to 650 mouse mutant lines from the EUCOMM ES cell resource as partners in the EUCOMM and EUMODIC research programmes (http://www.eucomm.org/info/; http://www.eumodic.org/aboutus.html).
Axenic service
Multicellular organisms are colonized by symbiotic, parasitic or infectious micro-organisms. In the oral cavity as well as in the intestine, one can identify up to 500 different species of microbes. Fortunately, very few micro-organisms are pathogenic, and most microbes are either beneficial for the host or have no apparent effect on the host. The most effective strategy for establishing what microbes do for the host is to generate germ-free animals that are raised under sterile (axenic) conditions. To such axenic mice one can re-introduce specific microbes—these animals are referred to as gnotobiotic animals—and study the impact of particular microbial strains on a given biological phenomenon. In addition, analysis of animals raised under germ-free conditions has permitted the proper attribution of a phenotype to a gene or biological pathway, independently of the microbial flora. Numerous differences between germ-free and conventionally raised animals have been reported, and these include a decrease in specific tissue and organ size, altered metabolism and deficiencies for vitamins, as well as variation in a large array of immunologic characteristics. EMMA offers as a service the generation of germ-free (axenic) mice.
Nomenclature
EMMA also provides the pertinent nomenclature for archived mice according to the rules and guidelines established by the International Committee on Standardised Genetic Nomenclature for Mice.
Cryocourses
The EMMA cryopreservation courses cover the latest findings on advanced techniques for embryo handling, IVF, cryopreservation, thawing and culture, quality control and dedicated database management. Further topics are basic procedures for breeding mouse colonies under SPF conditions, principles of cryobiology, long-term storage systems and cryogenic equipment and the use of specific databases for storage and breeding programs. Three EMMA partners host four cryocourses each year which are advertised via the website.
EMMA procedures
Evaluation of submitted mouse strains
The external EMMA evaluation committee (Figure 1) is comprised of experts in the field of mouse genetics who are not members of the EMMA network. They evaluate each submission to the EMMA repository, thereby ensuring that the scientific community has access to valuable mouse mutant strains with significance for current and future genetic research.
Intellectual property rights
EMMA-maintained lines are supplied to qualified researchers as a service to the scientific community at large, solely for research purposes and not for commercial use. The EMMA conditions further state that the recipient individual, laboratory or institution may not transfer or sell these mice or their progeny to any third party outside the recipient institution. Any already existing Material Transfer Agreements will remain in full force and effect.
SOPs
EMMA has developed and implemented SOPs for quality control, archiving and distribution of living and frozen material. Detailed SOPs are important for the daily management of both the laboratory and logistical settings. The SOPs for cryopreservation of mutant mouse lines as sperm or embryo and for superovulation and IVF are displayed on the public EMMA website at www.emmanet.org.
Cost of EMMA services
The EMMA cryopreservation service is free of charge but EMMA imposes a charge of 200 
/strain when it supplies either live mice or frozen materials to customers. In all cases, customers have to cover the actual costs for shipping the material to and from the EMMA facilities.
Animal welfare
All EMMA partners implement and fulfil the requirements for animal protection and welfare according to their respective national regulations. By the nature of its operation EMMA significantly contributes to a reduction of experimental animals used, thereby supporting the 3R principles of replacement, refinement and reduction. Moreover, EMMA strongly promotes the shipment of frozen material. If mice are sent to EMMA or if live mice are requested from the archive, EMMA ensures that mice are shipped according to conditions that comply with LASA (Laboratory Animals Science Association) guidelines.
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THE EMMA ARCHIVE |
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TOPABSTRACTBACKGROUNDTHE EMMA NETWORKEMMA SERVICES AND PROCEDURESADDITIONAL EMMA SERVICESTHE EMMA ARCHIVEEMMA GOES GLOBAL--FIMREVALUE FOR RESEARCH COMMUNITYREFERENCES |
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Since its inception, the number of mutant mouse strains submitted to EMMA and the number of requests has steadily increased. Up-to-date nearly 1300 mouse mutant lines were submitted to EMMA from scientists of 14 different European countries. In the coming 5 years, we expect a triplication of the mouse lines held in the EMMA archive resulting in one of the largest mouse repositories worldwide. Nearly 60% of the requests for mutant mouse lines that EMMA fulfilled were from European scientists and about one-third from the USA and Canada. Requests from Asia are steadily increasing (Figure 4). So far, EMMA shipped mouse mutant lines to customers in 35 different countries. Among the mutants archived by EMMA are targeted mutant strains including numerous Cre-expressing lines and knock-outs, transgenic lines and ENU-induced mutant strains. Phenotypes of archived strains cover neurological defects, hearing defects and strains with skeletal malformations among many others. Several strains representing interesting disease models and valuable Cre-expressing lines that are in high demand are maintained as live stocks facilitating a fast delivery to the customers.
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500 requests) |
EMMA GOES GLOBAL—FIMRE |
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TOPABSTRACTBACKGROUNDTHE EMMA NETWORKEMMA SERVICES AND PROCEDURESADDITIONAL EMMA SERVICESTHE EMMA ARCHIVEEMMA GOES GLOBAL--FIMREVALUE FOR RESEARCH COMMUNITYREFERENCES |
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EMMA is a founding member of FIMRe, a collaborating group of mouse repository and resource centres worldwide [22]. The EMMA director, Prof. Hrabe de Angelis acts as vice chair of the FIMRe Board of Directors representing Europe. In practice, FIMRe aims to archive and provide strains of mice as cryopreserved embryos and gametes, ES cell lines and live breeding stock to the research community. The specific goals are to:
- coordinate the archiving of valuable genetically defined mice and ES cell lines in line with global research demand;
- establish consistent and the highest possible quality animal health standards in all resource centres;
- provide genetic verification of the mutations and quality control for genetic background;
- collaborate to ensure rapid distribution of archived strains worldwide and provide resource training to enable researchers to use existing mouse resources.
The long term vision of the FIMRE network is to secure availability, to assure quality and to promote sharing of mutant mice and to disseminate information and resources to the global biomedical research community. Strains held at all the FIMRe resource centres are listed in the International Mouse Strain Resource (IMSR) (http://www.imsr.org) [23].
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VALUE FOR RESEARCH COMMUNITY |
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TOPABSTRACTBACKGROUNDTHE EMMA NETWORKEMMA SERVICES AND PROCEDURESADDITIONAL EMMA SERVICESTHE EMMA ARCHIVEEMMA GOES GLOBAL--FIMREVALUE FOR RESEARCH COMMUNITYREFERENCES |
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The mouse is the key model organism to study mammalian gene function mainly due to its excellent genetics and a plethora of available mutants modelling human diseases. These mutants offer the opportunity to decipher molecular disease mechanisms and provide in some instances a basis for the development of diagnostic, prognostic and therapeutic strategies. Therefore, it is essential that these valuable resources are retained in a central repository from which they can be readily made available to interested investigators. Thus, EMMA plays a crucial role in exploiting the potential benefits to human health presented by the research in mammalian genetics.
Outlook
The number of mutant mouse lines submitted to EMMA as well as the number of requests for mutants from EMMA is likely to increase significantly in the near future. This is due to large scale and systematic efforts worldwide to perform saturation mutagenesis of the mouse genome using gene targeting and gene trapping approaches. By the collaborative effort of the International Mouse Knockout Consortium more than 30 000 mutations in ES cells will be engineered and thousands of mouse disease models will become available over the next years [1]. In addition, we envision that humanized mice will be developed as disease models to mimic the human physiology more accurately and leading to an increased predictability of mouse models. All generated mouse mutants will ultimately require phenotyping, archiving and dissemination to the research laboratories. However, the current capacity to achieve this goal is limited. Indeed, existing facilities across Europe can offer capacity for the dissemination and analysis of around a few hundred disease models per year. It is thus necessary to organize phenotyping, archiving and distribution of mouse models on a well-concerted and large-scale level. To address these challenges EMMA will participate in Infrafrontier, a project of the European ESFRI Roadmap, which aims to organize two complementary and interlinked distributed infrastructures namely ‘Phenomefrontier’ and ‘Archivefrontier’ [24]. ‘Phenomefrontier’ will provide a European platform equipped with the latest technologies, in particular in vivo imaging and data management tools, for the phenotyping of medically relevant mouse models. ‘Archivefrontier’ will provide a European resource for state-of-the-art archiving and dissemination of those mouse models and will consist in a major upgrade of the EMMA.
As a founding member of FIMRE, a collaborating group of mouse repository and resource centres, EMMA will continue its efforts to contribute to the further development of this network and support the global distribution of the emerging mouse mutant resources. The long term goal and vision of the FIMRE network is to secure availability, to assure quality and to promote sharing of mutant mice and to disseminate information and resources to the global biomedical research community with EMMA being the European representative.
| Key Points The European Mouse Mutant Archive is a network of nine partners operating as the primary mouse repository in Europe. EMMA offers to the biomedical research community:
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FOOTNOTES |
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Martin Hrabè de Angelis, Susan Marschall and Michael Hagn work at the GSF National Research Center for Environment and Health, GmbH, Institute of Experimental Genetics.
Michael Hagn is the EMMA Project Manager.
Susan Marschall heads the Cryo-Unit at the Institute of Experimental Genetics.
Martin Hrabè de Angelis heads the Institute of Experimental Genetics and is director of EMMA and the German Mouse Clinic.
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TOPABSTRACTBACKGROUNDTHE EMMA NETWORKEMMA SERVICES AND PROCEDURESADDITIONAL EMMA SERVICESTHE EMMA ARCHIVEEMMA GOES GLOBAL--FIMREVALUE FOR RESEARCH COMMUNITYREFERENCES |
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