Single cell RNA-seq data of human hESCs comparing experiments with cDNA library equalization. Single cell RNA-seq data of human hESCs comparing experiments with cDNA library equalization

Considerable effort has been devoted to refining experimental protocols having reduced levels of technical variability and artifacts in single-cell RNA-sequencing data (scRNA-seq). We here present evidence that equalizing the concentration of cDNA libraries prior to pooling, a step not consistently performed in single-cell experiments, improves gene detection rates, enhances biological signals, and reduces technical artifacts in scRNA-seq data. In this submission the original single-cell cDNA is from GSE75748 cells labelled EC2 and TB2 in which the cDNA was not equalized prior to library preparation. We performed a set of experiments with the equalization step and provide three count matrices - 1) EQ (equalized cDNA prior to pooling in equal amounts); 2) EQ Vary (same as EQ but pooled at unequal amounts); 3) EQ-75% (equalized cDNA and pooled equally, sequeced to lower total depth). Overall design: Single cell RNA-seq data of human hESCs comparing experiments with cDNA library equalization

已有大量研究致力于优化实验方案，以降低单细胞RNA测序（single-cell RNA-sequencing, scRNA-seq）数据中的技术变异与伪影。本研究证实，在混合测序文库前均一化互补DNA（complementary DNA, cDNA）文库浓度——这一步骤在单细胞实验中并非普遍执行——可提升基因检出率、增强生物学信号，并降低scRNA-seq数据中的技术伪影。本提交数据集的原始单细胞cDNA来自标注为EC2与TB2的GSE75748细胞，该批样本在文库制备前未进行浓度均一化处理。我们通过增设浓度均一化步骤开展了一系列对照实验，并提供三份计数矩阵：1）EQ组：混合前将cDNA均一化至等量浓度；2）EQ Vary组：处理流程与EQ组一致，但混合时采用不等量比例；3）EQ-75%组：cDNA经均一化且等量混合，测序总深度较低。整体实验设计：针对人类胚胎干细胞（human embryonic stem cells, hESCs）的单细胞RNA测序数据，对比开展与未开展cDNA文库均一化的实验结果。