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Serenity, a senior studying Biology at Tufts University, assessed the efficacy of single-nuclei RNA sequencing technologies for profiling individual cell types in neonatal, pediatric, and adolescent post-mortem tissues to contribute towards the constriction of a molecular and data analysis resource portal, dGTEx for characterizing patterns of gene expression during pediatric development

The Developmental Genotype-Tissue Expression (dGTEx) project aims to build a public data resource and portal harboring single cell as well as bulk datasets from reference tissue samples across healthy neonatal, pediatric, and adolescent postmortem donors. I found it a privilege to be surrounded by wonderful scientists who were all exceptionally generous with me as I was navigating my way around Ó³»­´«Ã½ this summer. I immediately felt a connection to the project I was working on and became very comfortable with my mentor who showed me patience and ultimately independence while working in the lab. I am also grateful for the BSRP staff, especially Bruce and Frances, who took the time to mentor us in key lessons of how to be a more mindful, courageous, thoughtful, and goal-driven human. This is an experience I will never forget! It seeks to establish a molecular and data analysis resource as well as a tissue bank for the scientific community to study how gene expression and regulation varies across human developmental stages. Comprehensive transcriptional characterization of these tissues requires single-nuclei RNA-sequencing (snRNA-seq) but scaling this technology to thousands of samples poses challenges, including significant monetary costs and batch effects. Here our objective is to examine cost-effective and efficient snRNA-seq technologies to profile the tissues. In this study, nuclei were isolated from biobanked frozen tissue samples and subjected to the novel snRNA-seq technology, PIPseq (Fluent BioSciences) to test whether it could serve as an alternative to existing 10x Genomics technology for transcriptionally characterizing singular cells. Through the gene expression profile analysis, we compared sequencing outputs between a PIPseq V4 T20 and 10X Chromium 3’ v3.2 Chemistry reactions on the same samples.  We found that the PIPseq technology was highly efficient, capturing about twice the amount of cells compared to 10x, and was cost-effective in yielding a cost per cell 40% lower than 10x. Choosing a technology motivated by the goal of profiling and generating high quality data for a large number of cells encourages the lab to consider integrating the PIPseq workflow into the pipeline for processing tissue samples for the dGTEx project, which will facilitate the construction of a molecular portal for characterizing patterns of gene expression across pediatric development.

Project: Comparison of single nuclei gene expression profiling technologies for the dGTEx Projec

Mentor: Amrita Sule, Gene Regulation Observatory