(2)部分library要求细胞本身就表达Cas9,这本无可厚非,但部分细胞在高表达Cas9的情况下非常脆弱,这时候需要考虑使用inducible Cas9 system,而这个系统的构建本身就是有难度的实验,因此在设计可提前一定要做好全面分析。 (3)Library扩增工作量非常大,需要尽可能的严格按照protocol进行,并且配备质粒超大提试剂盒,部分Libra...
Here, we describe a general protocol for the generation of custom, pooled CRISPR sgRNA libraries for screening in cardiomyocyte cultures. This methodology can address a variety of lab-specific research questions in cardiomyocytes and other cell types, as the genes to be modified can be curated or...
(1)CRISPR screening在准备阶段需要使用电转的方式扩增文库,并需要NGS测序来确定文库丰度,尽管目前已经有一些library可以选用化学感受态细胞进行扩增,但选用library之前一定要查阅清楚; (2)CRISPR screening实验需要大量的细胞,因此对于一些无法扩增、数量有限且非常珍贵的细胞时,如原代细胞等,CRISPR screening则可能不适用; ...
Genome-scale CRISPR-Cas9 knockout and transcriptional activation screening | Nature Protocol | 基因组规模的 CRISPR-Cas9 敲除和转录激活筛选 Genome-wide CRISPR/Cas9 library screening identified PHGDH as a critical driver for Sorafenib resistance in HCC | 全基因组 CRISPR/Cas9 文库筛选确定 PHGDH 是 HCC...
CRISPRa is highly useful in screening for gain of functions, and CRISPRi is a more powerful tool than RNA interference (RNAi) libraries in screening for loss of functions. Positive selection using a CRISPR library can detect survival cells with specific conditions, such as drug treatment, and ...
response to oxidative stress, and genomic stability [135,136]. In engineered human T cells, the application of CRISPR–Cas9 library screening serves a pivotal role in the identification of primary regulators governing cell fate. This innovative approach enables the precise editing of these genes with...
By using genome-wide CRISPR/Cas9 library screening, we identify phosphoglycerate dehydrogenase (PHGDH), the first committed enzyme in the serine synthesis pathway (SSP), as a critical driver for Sorafenib resistance. Sorafenib treatment activates SSP by inducing PHGDH expression. With RNAi knockdown ...
CRISPRa is highly useful in screening for gain of functions, and CRISPRi is a more powerful tool than RNA interference (RNAi) libraries in screening for loss of functions. Positive selection using a CRISPR library can detect survival cells with specific conditions, such as drug treatment, and ...
Does IDT offer CRISPR screening analysis solutions? Does IDT offer CRISPR library design services? I want to target each of my genes with multiple guides. How many gRNAs do you suggest per gene? What synthetic arrayed library formats are available from IDT? Are additional customization options pos...
Sorafenib is the standard treatment for advanced hepatocellular carcinoma (HCC). However, the development of drug resistance is common. By using genome-wide CRISPR/Cas9 library screening, we identify phosphoglycerate dehydrogenase (PHGDH), the first committed enzyme in the serine synthesis pathway (SSP...