کد مقاله کد نشریه سال انتشار مقاله انگلیسی نسخه تمام متن
5513596 1541206 2017 45 صفحه PDF دانلود رایگان
عنوان انگلیسی مقاله ISI
Gene editing and clonal isolation of human induced pluripotent stem cells using CRISPR/Cas9
موضوعات مرتبط
علوم زیستی و بیوفناوری بیوشیمی، ژنتیک و زیست شناسی مولکولی زیست شیمی
پیش نمایش صفحه اول مقاله
Gene editing and clonal isolation of human induced pluripotent stem cells using CRISPR/Cas9
چکیده انگلیسی
Human induced pluripotent stem cells (hiPSCs) represent an ideal in vitro platform to study human genetics and biology. The recent advent of programmable nucleases makes also the human genome amenable to experimental genetics through either the correction of mutations in patient-derived iPSC lines or the de novo introduction of mutations into otherwise healthy iPSCs. The production of specific and sometimes complex genotypes in multiple cell lines requires efficient and streamlined gene editing technologies. In this article we provide protocols for gene editing in hiPSCs. We presently achieve high rates of gene editing at up to three loci using a modified iCRISPR system. This system includes a doxycycline inducible Cas9 and sgRNA/reporter plasmids for the enrichment of transfected cells by fluorescence-activated cell sorting (FACS). Here we cover the selection of target sites, vector construction, transfection, and isolation and genotyping of modified hiPSC clones.
ناشر
Database: Elsevier - ScienceDirect (ساینس دایرکت)
Journal: Methods - Volumes 121–122, 15 May 2017, Pages 29-44
نویسندگان
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