Research Report 2010 - MDC

Structure of the GroupGroup LeaderDr. Boris JerchowSecretariatChristine KrauseBoris JerchowTechnical AssistantsKatja BeckerTianwu QiaoAndrea LeschkeTransgenicsThe Transgenic Core Facility (TCF) offers expertise in all kinds of mouse Assisted ReproductiveTechnologies (ART). Starting at the planning phase, we support scientists from the MDCand their external collaborators with the design and layout of their projects. One focus of ourwork is the generation of genetically modified mouse lines. This is accomplished either bygene targeting in embryonic stem (ES) cells and subsequent generation of chimeric mice fromrecombinant ES cell clones or by microinjection of plasmid or BAC type transgenes into thepronuclei of fertilized oocytes. During the last year, we had the chance to develop a relatedtechnique together with Lajos Mates from the group of Zsuzsanna Izsvak: After coinjection of atransgene flanked by specific inverted repeats together with RNA encoding an optimizedversion of the Sleeping Beauty Transposase, SB100, into oocytes, an enzyme-mediatedintegration of the transgene takes place (refer to selected publication). Compared toconventional microinjection this technique leads to the integration of single transgenesinstead of the integration of an uncontrollable number of copies as a concatamer and yields ahigher rate of transgenic founders. The size of the transgene is a limitation of this technique,though, and only plasmid type sized transgenes can be integrated into the genome.A second focus of our work is the conservation of preciousmouse lines and the rederivation of conservedlines. This service has been continuously expanded duringthe last years both in quantity and in terms of thedifferent protocols that have been optimized in the lab.We can now offer the long term cryopreservation ofpre-implantation embryos as well as sperm in liquidnitrogen. Due to the growing number of mouse modelsworldwide, we see an increase in the number of organizations,who offer to send frozen material instead oflive mice. We therefore see an increased demand forrevitalization of lines conserved by third parties followinga variety of protocols. In this context we were successfulto establish a new protocol developed by theJackson Laboratories that make even recalcitrant lineslike C57BL/6 amenable for in vitro fertilization. In addition,we have a laser device at our disposal that can beused to penetrate the outer zona pellucida of the oocyteto facilitate sperm entry and thereby achieve fertilizationeven in cases, where this does not spontaneouslyhappen.In addition to the above, we have just taken on the rederivationof hygienically compromised lines by embryotransfer. Rederivation by hysterectomy is therefore discontinued.There are individual projects that do not fit into any ofthe above that can be supported by the TCF and we willhelp whenever it comes to the production, isolation,manipulation, culture or retransfer of pre-implantationembryos. Moreover, we can give advice on cloning andtargeting strategies, BAC preparation, ES cell culture, EScell strain background, coat color genetics, ES cell derivation,and more.Besides ongoing projects for our clients, the currentcenter of our work is the optimization of ES cell deriva-230 Technology Platforms