Highly parallel and efficient single cell mRNA sequencing with paired picoliter chambers

单细胞转录组测序技术在单个细胞水平上对转录组进行高通量测序分析，从而揭示单个细胞内所有基因的表达情况，揭示细胞间的异质性，在发育生物学、免疫学、微生物学、神经科学、临床医学等领域有重要的应用前景。单细胞转录组测序的挑战在于如何高效地操控单个细胞，如何对大量的低拷贝数mRNA进行无偏倚扩增，如何避免背景游离mRNA的污染，以及如何同时对大量的单细胞进行并行测序以降低成本。化学化工学院杨朝勇教授课题组在高通量单细胞转录组测序新器件新方法研究方面取得重要进展.该工作由厦门大学、上海交通大学、美国斯坦福大学等多团队联合攻关完成。化学生物学系博士研究生张明霞、邹远和2011协同创新中心博士研究生许醒为论文的共同第一作者。ScRNA-seq has the ability to reveal accurate and precise cell types and states. Existing scRNA-seq platforms utilize bead-based technologies uniquely barcoding individual cells, facing practical challenges for precious samples with limited cell number. Here, we present a scRNA-seq platform, named Paired-seq, with high cells/beads utilization efficiency, cell-free RNAs removal capability, high gene detection ability and low cost. We utilize the differential flow resistance principle to achieve single cell/barcoded bead pairing with high cell utilization efficiency (95%). The integration of valves and pumps enables the complete removal of cell-free RNAs, efficient cell lysis and mRNA capture, achieving highest mRNA detection accuracy (R = 0.955) and comparable sensitivity. Lower reaction volume and higher mRNA capture and barcoding efficiency significantly reduce the cost of reagents and sequencing. The single-cell expression profile of mES and drug treated cells reveal cell heterogeneity, demonstrating the enormous potential of Paired-seq for cell biology, developmental biology and precision medicine.The authors thank the National Science Foundation of China (21927806, 21735004, 21521004, 21325522), the National Key R&D Program of China (2018YFC1602900), Innovative research team of high-level local universities in Shanghai, and the Program for Changjiang Scholars and Innovative Research Team in University (IRT13036) for their financial support.该研究工作得到国家重大科研仪器研制项目、国家基金委重点项目、创新研究群体项目等支持