CORE--MOUSE EMBRYONIC STEM (ES) CELL AND GENE TARGETING

点击翻译按钮获取中文摘要

Services provided by this Core take on various forms, in part tailored to meet the needs of the investigators who have been the users. In many instances, the service has been comprehensive and included advice on design of a targeting construct, providing selection cassettes for inclusion in the constructs, electroporation and selection of ES colonies, preparation of DMA, karyotype analysis of positive clones, and microinjection into blastocysts. Under these circumstances, the only task left to the investigator has been Southern blot analysis of DMA samples to identify targeted clones. In other instances, particularly with those in close proximity to the PI's laboratory, ES cells have been provided to the investigator for his/her own targeting. When positive clones are identified by Southern blot analysis, karyotyping and blastocyst microinjection are performed by this Core. Beyond the mechanics of gene targeting and blastocyst injection, this Core provides an additional, invaluable role--advice and assistance in the analysis of mouse phenotypes. this is largely managed by Dr. Fujiwara. The analysis of mutants requires use of specialized methods, including dissection of embryos, bleeding of embryos, hematopoietic colony assays from yolk sac, fetal liver, or peripheral blood samples, chimera analysis, and lacZ staining, in situ hybridization and histology. The experience and knowledge base of this Core has provided enormously useful advice and hands on assistance to many investigators, some who used no others services of the Core. Because the strengths became readily known in the Boston area, investigators throughout the Harvard Medical Area and elsewhere, approached this Core for guidance in evaluating their mouse knockout phenotypes. The Core has three simultaneous goals: First, the presence of this Core allows for highly efficient and successful production of gene targeted mice for members of the PI's laboratory. Second, because this Core had the appropriate expertise in both personnel and experience, it serves numerous investigators within the Harvard Medical Area and beyond. Third, this Core exports its products (largely mouse strains) to investigators elsewhere to support their own research projects and influence the field of hematology in the broadest sense.