基因组测序研究已经确定了广泛肿瘤类型中大量癌症突变，但是确定这些突变的表型影响仍然是一个挑战。本文中，我们开发了一种名为TISCC-seq的高通量、多重单细胞技术，使用CRISPR碱基编辑酶工程预先设计的突变，并直接在单个细胞中描绘其基因型，并确定每个突变的转录表型。目标基因转录的长读测序鉴定了工程突变，同时使用短读测序分析相同细胞集合的转录组剖析。通过整合，我们确定了基因型和表达表型的细胞分辨率的突变。使用细胞系，我们对超过100个TP53突变的基因表达影响进行工程和评估。基于单细胞基因表达，我们将这些突变分为具有功能显著表型的突变。© 2023. 作者。

Genome sequencing studies have identified numerous cancer mutations across a wide spectrum of tumor types, but determining the phenotypic consequence of these mutations remains a challenge. Here, we developed a high-throughput, multiplexed single-cell technology called TISCC-seq to engineer predesignated mutations in cells using CRISPR base editors, directly delineate their genotype among individual cells and determine each mutation's transcriptional phenotype. Long-read sequencing of the target gene's transcript identifies the engineered mutations, and the transcriptome profile from the same set of cells is simultaneously analyzed by short-read sequencing. Through integration, we determine the mutations' genotype and expression phenotype at single-cell resolution. Using cell lines, we engineer and evaluate the impact of >100 TP53 mutations on gene expression. Based on the single-cell gene expression, we classify the mutations as having a functionally significant phenotype.© 2023. The Author(s).