Laboratory of Adaptive Immunity, Institute of Molecular Genetics of the Czech Academy of Sciences, Prague, Czechia.
Front Immunol. 2024 Oct 9;15:1446931. doi: 10.3389/fimmu.2024.1446931. eCollection 2024.
Single-cell RNA sequencing (scRNA-seq) methods are widely used in life sciences, including immunology. Typical scRNA-seq analysis pipelines quantify the abundance of particular transcripts without accounting for alternative splicing. However, a well-established pan-leukocyte surface marker, CD45, encoded by the gene, presents alternatively spliced variants that define different immune cell subsets. Information about some of the splicing patterns in particular cells in the scRNA-seq data can be obtained using isotype-specific DNA oligo-tagged anti-CD45 antibodies. However, this requires generation of an additional sequencing DNA library. Here, we present IDEIS, an easy-to-use software for CD45 isoform quantification that uses single-cell transcriptomic data as the input. We showed that IDEIS accurately identifies canonical human CD45 isoforms in datasets generated by 10× Genomics 5' sequencing assays. Moreover, we used IDEIS to determine the specificity of the splicing pattern in mouse leukocyte subsets.
单细胞 RNA 测序 (scRNA-seq) 方法广泛应用于生命科学领域,包括免疫学。典型的 scRNA-seq 分析流程定量特定转录本的丰度,但不考虑可变剪接。然而,一个成熟的泛白细胞表面标志物 CD45,由 基因编码,存在定义不同免疫细胞亚群的可变剪接变体。可以使用同种型特异性 DNA 寡核苷酸标记的抗 CD45 抗体从 scRNA-seq 数据中的特定细胞中获得有关某些剪接模式的信息。然而,这需要生成额外的测序 DNA 文库。在这里,我们提出了 IDEIS,这是一种用于 CD45 异构体定量的易于使用的软件,它将单细胞转录组数据作为输入。我们表明,IDEIS 可以准确识别 10× Genomics 5' 测序测定生成的数据集中原核人类 CD45 异构体。此外,我们使用 IDEIS 来确定小鼠白细胞亚群中 剪接模式的特异性。