Open AccessImproving single-cell transcriptome sequencing efficiency with a microfluidic phase-switch device
Author(s) -
Baoyue Zhang,
Hong Xu,
Yuqing Huang,
Shu Wang,
Hongtao Feng,
Jin Cai,
Jiang Zhong,
Chen Yan
Publication year - 2019
Publication title -
analyst (london. 1877. online)/analyst
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.998
H-Index - 153
eISSN - 1364-5528
pISSN - 0003-2654
DOI - 10.1039/c9an00823c
Subject(s) - microfluidics , transcriptome , phase (matter) , computational biology , chemistry , nanotechnology , materials science , biology , gene , gene expression , biochemistry , organic chemistry
In this paper, we present a novel method to improve the efficiency of single-cell transcriptome sequencing for analyzing valuable cell samples. The microfluidic device we designed integrates multiple single-cell isolation chambers with hydrodynamic traps and achieves a nearly 100% single-cell capture rate and minimal cell loss, making it particularly suitable for samples with limited numbers of cells. Single cells were encapsulated using a novel phase-switch method into picoliter-sized hydrogel droplets and easily recovered for subsequent reactions. Minimizing the reaction volume resulted in a high reverse transcription (RT) efficiency for RNA sequencing (RNA-Seq). With this novel microfluidic platform, we captured dozens of hESCs (H9) simultaneously and obtained live cells in individual picoliter volumes, thus allowing for the convenient construction of a high-quality library for deep single-cell RNA-Seq. Our single-cell RNA-Seq results confirmed that a spectrum of pluripotency existed within an H9 colony. This integrated microfluidic platform can be applied to various cell types for the investigation of rare cellular events, and the phase-switch single-cell processing strategy will improve the efficiency and accessibility of single-cell transcriptome sequencing analysis.