Amplicon sequencing of genomically-encoded guide RNA sequences for the Brunello CRISPR library in eHap cells

eHap CRISPR-Cas9 mutagenized cells with no selection serves as a control. RPS6p antibody staining on the mutagenized eHap cells was used in a flow-sorting approach to isolate RPS6p high and low populations respectively. The genomic DNA from these different samples was isolated, subjected to PCR to amplify the genomically-encoded guide RNA sequences, the product(s) of which was then sequenced on a Hi-Seq X Ten instrument by Novogene.

未经筛选的eHap CRISPR-Cas9诱变细胞作为对照。对诱变后的eHap细胞进行RPS6p抗体染色，随后通过流式分选技术分别分离出RPS6p高表达与低表达的细胞群。提取上述不同样本的基因组DNA，通过PCR扩增基因组编码的向导RNA（guide RNA）序列，随后由诺禾致源（Novogene）在Hi-Seq X Ten测序仪上完成测序。