Single-cell multi-gene identification of somatic mutations and gene rearrangements in cancer.
Single-cell multi-gene identification of somatic mutations and gene rearrangements in cancer.
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在这项概念验证研究中，我们开发了一种单细胞方法，该方法能提供在信使RNA编码区域发现的体细胞变异的基因型，并将这些基于转录本的变异与其匹配的细胞转录组整合在一起。我们在单细胞互补DNA文库上使用纳米孔自适应采样来验证靶基因转录本中的编码变异，并使用短读长测序来表征携带突变的细胞类型。利用一种癌细胞系鉴定出了16个靶点的CRISPR编辑，并且使用一个352基因 panel验证了该细胞系中的已知变异。使用包含161到529个基因的靶基因panel验证了原发性癌症样本中的变异。还在一名患者中鉴定出一种基因重排，这种重排发生在两个不同的肿瘤部位。
In this proof-of-concept study, we developed a single-cell method that provides genotypes of somatic alterations found in coding regions of messenger RNAs and integrates these transcript-based variants with their matching cell transcriptomes. We used nanopore adaptive sampling on single-cell complementary DNA libraries to validate coding variants in target gene transcripts, and short-read sequencing to characterize cell types harboring the mutations. CRISPR edits for 16 targets were identified using a cancer cell line, and known variants in the cell line were validated using a 352-gene panel. Variants in primary cancer samples were validated using target gene panels ranging from 161 to 529 genes. A gene rearrangement was also identified in one patient, with the rearrangement occurring in two distinct tumor sites.