of Core projects active during FY2017: In FY2018, the Core was used by 12 investigators from all nine NHGRI branches for a number of projects as described below. Generation of mutant and transgenic lines, microinjections and training are some of our most popular services. As we completed projects for Burgess, Gahl and Kastner labs, we started new projects for Pavan, Brody and Muenke labs. A brief summary of the projects and number of services provided is described below. Microinjections of morpholinos and mRNA to evaluate patient variants in NAA10 to determine their pathogenicity (Biesecker lab). Microinjections of lung fibroblasts from HPS patients to monitor their migration in response to treatment (Gahl lab). Microinjections of transgenes, sgRNAs and morpholinos for targeted knockout, knockin and rescue projects for Burgess and Liu labs Generation of knockout mutants for 20 genes (Pavan, Muenke, Venditti and Chandrasekharappa labs). Pavan lab is investigating the role of ten genes in pigmentation, building upon their successful publication of the msfd12a zebrafish model in Science. Mutant generation for 5 genes is in progress for Brody and Muenke labs. CRISPR sgRNA design and injections for 5 genes for Chandrasekharappa lab to investigate genetic compensation. Hands-on training to 15 new users in microinjections, zebrafish handling, breeding, euthanasia, anesthesia, fin clips, genotyping, sgRNA and primer design, CRISPR-STAT, fluorescent PCR, sequence analysis, WISH, o-Dianisidine staining, bone and cartilage staining, imaging, tissue extraction and IVF. Founder screening to generate a transgenic line to visualize xanthopores (Pavan lab). Cloned, sequence validated and injected 2 non-coding fragments for evaluation of their role as regulatory elements (Wilson lab) Participated in characterization of the phenotypes of knockout fish for >35 genes involved in a variety of diseases and biological processes (Fanconi anemia, hematopoiesis, inflammatory diseases, leukemia, pigmentation and thyroid development) by analysis of embryonic and larval phenotypes by fin clips, fluorescent PCR, genotyping, WISH, imaging and histology. Performed cryopreservation and IVF of all new mutant and transgenic lines. Sequencing to genotype mutant lines with ENU-induced mutations for the Burgess lab. Facilitated importing and exporting fish lines for collaborations. Maintained 15 lines (WT and transgenic) by breeding, screening for the appropriate reporter using microscope, and genotyping. Technology development by testing various knock-in and base editing approaches. To generate above-mentioned mutant lines, we processed 35,000 DNA samples for genotyping by fluorescent PCR and performed 8,000 sequencing reactions using our capillary electrophoresis method.
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