使用一组全面的外泌体 RNA 内参对照对人生物体液中的细胞外 RNA 进行信使 RNA 捕获测序。

Center for Medical Genetics, Department of Biomolecular Medicine, OncoRNALab, Ghent University, C. Heymanslaan 10, 9000 Ghent, Belgium.

Cancer Research Institute Ghent (CRIG), Ghent University, C. Heymanslaan 10, 9000 Ghent, Belgium.

STAR Protoc. 2021 Apr 14;2(2):100475. doi: 10.1016/j.xpro.2021.100475. eCollection 2021 Jun 18.

Comprehensive transcriptome analysis of extracellular RNA (exRNA) purified from human biofluids is challenging because of the low RNA concentration and compromised RNA integrity. Here, we describe an optimized workflow to (1) isolate exRNA from different types of biofluids and (2) to prepare messenger RNA (mRNA)-enriched sequencing libraries using complementary hybridization probes. Importantly, the workflow includes 2 sets of synthetic spike-in RNA molecules as processing controls for RNA purification and sequencing library preparation and as an alternative data normalization strategy. For complete details on the use and execution of this protocol, please refer to Hulstaert et al. (2020).

从人体生物体液中纯化的细胞外 RNA (exRNA) 进行综合转录组分析具有挑战性，因为其 RNA 浓度低且 RNA 完整性受损。在这里，我们描述了一种优化的工作流程，用于 (1) 从不同类型的生物体液中分离 exRNA，以及 (2) 使用互补杂交探针来制备信使 RNA (mRNA) 富集测序文库。重要的是，该工作流程包括 2 组合成的 Spike-in RNA 分子，作为 RNA 纯化和测序文库制备的处理对照，以及替代数据归一化策略。有关此方案的使用和执行的完整详细信息，请参阅 Hulstaert 等人 (2020)。