3’Pool-seq: an optimized cost-efficient and scalable method of whole-transcriptome gene expression profiling (Run0050). 3’Pool-seq: an optimized cost-efficient and scalable method of whole-transcriptome gene expression profiling (Run0050)

The advent of Next Generation Sequencing has allowed transcriptomes to be profiled with unprecedented accuracy, but the steep costs associated with full-length library sequencing have posed a limit on the accessibility and scalability of the technology. To address these limitations, we developed 3’Pool-seq, a simple, cost-effective, and scalable RNA-seq method that focuses sequencing to the 3’ end of mRNA transcripts. Overall design: Transcriptomes (3 replicates each) from homozygous GFAP-IL6 mice were compared to those from wild-type C57BL/6 for differential gene expression using different amounts of cDNA input during the tagmentation step of 3'Pool-seq. In this specific experiment, different amounts of cDNA were added to the Tagmentation reaction in order to optimize that step for 3'Pool-seq.

下一代测序（Next Generation Sequencing）技术的问世，使得转录组能够以前所未有的精度完成表征分析，但全长文库测序所伴随的高昂成本，对该技术的可及性与可扩展性构成了显著限制。
为解决上述局限，我们开发了3’Pool-seq技术——一种简便易用、成本可控且具备良好可扩展性的RNA测序方法，该方法将测序聚焦于信使RNA（mRNA）转录本的3'末端区域。
实验整体设计：将纯合GFAP-IL6小鼠（每组设置3次生物学重复）的转录组与野生型C57BL/6小鼠的转录组进行对比，通过在3’Pool-seq的转座标签化（tagmentation）步骤中设置不同的互补DNA（cDNA）投入量，开展差异基因表达分析。
本项特定实验中，为优化3’Pool-seq的转座标签化步骤，向转座标签化反应体系中加入了不同剂量的cDNA。