Department of Molecular Physiology and Biophysics, Vanderbilt University.
Division of Metabolic Health and Disease, Oregon National Primate Research Center.
J Vis Exp. 2024 Nov 8(213). doi: 10.3791/66849.
One challenge in studies using tissue collected from multiple cohorts is avoiding batch effects when preparing for large-scale multi-omic experiments, such as combined single-cell RNA sequencing and metabolomics. The method in the current study utilizes flash-frozen pancreatic islets from fetal non-human primates collected over a span of two years for input into single-nucleus RNA sequencing and ATAC sequencing assays. The cytosolic fraction generated during nuclear extraction was retained for downstream capillary electrophoresis-mass spectrometry and subsequent metabolite quantification. This method allows for bulk analysis of metabolites that contribute to the changing transcriptomic and epigenomic landscapes within experimental conditions. It is applicable to many tissue types and maximizes the amount of information that can be extracted from samples that are not readily available. As the contribution of metabolism to the establishment of cellular identity via epigenetic modifications becomes more appreciated, techniques that allow for identifying the contribution of metabolites in specific cell types are timely and necessary.
在使用来自多个队列的组织进行研究时，一个挑战是在为大规模多组学实验（如单细胞 RNA 测序和代谢组学的组合）做准备时避免批次效应。本研究中使用的方法利用了在两年时间内收集的来自胎儿非人类灵长类动物的冷冻胰岛组织，用于单细胞 RNA 测序和 ATAC 测序分析。核提取过程中产生的胞质部分被保留下来，用于下游毛细管电泳-质谱分析和随后的代谢物定量。这种方法允许对在实验条件下发生变化的转录组和表观基因组景观有贡献的代谢物进行批量分析。它适用于许多组织类型，并最大限度地从不易获得的样本中提取信息。随着代谢物通过表观遗传修饰对细胞身份建立的贡献得到更多的认识，能够识别特定细胞类型中代谢物贡献的技术是及时和必要的。
