Department of Bioengineering, University of California, Berkeley, California Institute for Quantitative Biosciences, Berkeley, CA, USA.
Fluent Biosciences, Watertown, MA, USA.
Nat Biotechnol. 2023 Nov;41(11):1557-1566. doi: 10.1038/s41587-023-01685-z. Epub 2023 Mar 6.
Current single-cell RNA-sequencing approaches have limitations that stem from the microfluidic devices or fluid handling steps required for sample processing. We develop a method that does not require specialized microfluidic devices, expertise or hardware. Our approach is based on particle-templated emulsification, which allows single-cell encapsulation and barcoding of cDNA in uniform droplet emulsions with only a vortexer. Particle-templated instant partition sequencing (PIP-seq) accommodates a wide range of emulsification formats, including microwell plates and large-volume conical tubes, enabling thousands of samples or millions of cells to be processed in minutes. We demonstrate that PIP-seq produces high-purity transcriptomes in mouse-human mixing studies, is compatible with multiomics measurements and can accurately characterize cell types in human breast tissue compared to a commercial microfluidic platform. Single-cell transcriptional profiling of mixed phenotype acute leukemia using PIP-seq reveals the emergence of heterogeneity within chemotherapy-resistant cell subsets that were hidden by standard immunophenotyping. PIP-seq is a simple, flexible and scalable next-generation workflow that extends single-cell sequencing to new applications.
现有的单细胞 RNA 测序方法存在一些局限性，这些局限性源于样本处理所需的微流控设备或流体处理步骤。我们开发了一种不需要专门的微流控设备、专业知识或硬件的方法。我们的方法基于基于粒子模板的乳化，它允许单细胞在均匀的液滴乳液中进行 cDNA 的封装和条形码化，只需要一个涡旋混合器。基于粒子模板的即时分区测序（PIP-seq）可以适应各种乳化格式，包括微孔板和大容量锥形管，使数千个样本或数百万个细胞可以在几分钟内处理。我们证明，PIP-seq 在小鼠-人类混合研究中产生了高纯度的转录组，与多组学测量兼容，并且与商业微流控平台相比，可以更准确地描述人乳腺组织中的细胞类型。使用 PIP-seq 对混合表型急性白血病进行单细胞转录组分析揭示了化疗耐药细胞亚群内异质性的出现，这些亚群的异质性被标准免疫表型分析所掩盖。PIP-seq 是一种简单、灵活和可扩展的下一代工作流程，将单细胞测序扩展到新的应用领域。
