Department of Genome Sciences, University of Washington, Seattle, Washington, USA.
Howard Hughes Medical Institute, Seattle, Washington, USA.
Nat Methods. 2018 Apr;15(4):271-274. doi: 10.1038/nmeth.4604. Epub 2018 Feb 19.
Several groups recently coupled CRISPR perturbations and single-cell RNA-seq for pooled genetic screens. We demonstrate that vector designs of these studies are susceptible to ∼50% swapping of guide RNA-barcode associations because of lentiviral template switching. We optimized a published alternative, CROP-seq, in which the guide RNA also serves as the barcode, and here confirm that this strategy performs robustly and doubled the rate at which guides are assigned to cells to 94%.
最近有几个研究小组将 CRISPR 扰动和单细胞 RNA-seq 结合起来进行 pooled 遗传筛选。我们证明，由于慢病毒模板转换，这些研究的载体设计容易发生约 50%的向导 RNA-条码关联交换。我们对已发表的替代方案 CROP-seq 进行了优化，其中向导 RNA 也充当条码，在此我们确认该策略表现稳健，将向导分配给细胞的速度提高了一倍，达到 94%。
